Genetics

From the University of Arizona Cancer Center, Tucson, AZ (Dr. Ehsani), and University of Wisconsin Carbone Cancer Center and School of Medicine and Public Health, Madison, WI (Dr. Wisinski).

Abstract

• Objectives: To describe common genomic tests being used clinically to assess prognosis and guide adjuvant chemotherapy and endocrine therapy decisions for early-stage breast cancer.
• Methods: Case presentation and review of the literature.
• Results: Hormone receptor–positive (HR-positive) breast cancers, which express the estrogen and/or progesterone receptor, account for the majority of breast cancers. Endocrine therapy can be highly effective for patients with these HR-positive tumors, and identification of HR-positive breast cancers that do not require the addition of chemotherapy is critical. Clinicopathological features of the breast cancer, including tumor size, nodal involvement, grading, and HR status, are insufficient in predicting the risk for recurrence or the need for chemotherapy. Furthermore, a portion of HR-positive breast cancers have an ongoing risk for late recurrence, and longer durations of endocrine therapy are being used to reduce this risk.
• Conclusion: There is sufficient evidence for use of genomic testing in early-stage HR-positive breast cancer to aid in chemotherapy recommendations. Further confirmation of genomic assays for prediction of benefit from prolonged endocrine therapy is needed.

Key words: molecular testing; decision aids; HR-positive cancer; recurrence risk; adjuvant chemotherapy; endocrine therapy.

Despite the increase in incidence of breast cancer, breast cancer mortality has decreased over the past several decades. This is likely due to both early detection and advances in systemic therapy. However, with more widespread use of screening mammography, there are increasing concerns regarding potential overdiagnosis of cancer [1]. One key challenge is that breast cancer is a heterogeneous disease. Thus, improved tools for determining breast cancer biology can help physicians individualize treatments, with low-risk cancers approached with less aggressive treatments, thus preventing unnecessary toxicities, and higher-risk cancers treated appropriately.

Traditionally, adjuvant chemotherapy was recommended based on tumor features such as stage (tumor size, regional nodal involvement), grade, expression of hormone receptors (estrogen receptor [ER] and progesterone receptor [PR]) and human epidermal growth factor receptor-2 (HER2), and patient features (age, menopausal status). However, this approach is not accurate enough to guide individualized treatment recommendations, which are based on the risk for recurrence and the reduction in this risk that can be achieved with various systemic treatments. In particular, there are individuals with low-risk HR-positive, HER2-negative breast cancers who could be spared the toxicities of cytotoxic chemotherapies without compromising the prognosis.

Beyond chemotherapy, endocrine therapies also have risks, especially when given for extended durations. Recently, extended endocrine therapy has been shown to prevent late recurrences of HR-positive breast cancers. In the MA.17R study, extended endocrine therapy with letrozole for a total of 10 years (beyond 5 years of an aromatase inhibitor [AI]) decreased the risk for breast cancer recurrence or the occurrence of contralateral breast cancer by 34% [2]. However, the overall survival was similar between the 2 groups and the results were not confirmed in other studies [3–5]. Identifying the subgroup of patients who benefit from this extended AI therapy is important in the era of personalized medicine. Several tumor genomic assays have been developed to provide additional prognostic and predictive information with the goal of individualizing adjuvant therapies for breast cancer. Although assays are also being evaluated in HER2-positive and triple negative breast cancer, this review will focus on HR-positive, HER2-negative breast cancer.

Case Study

Initial Presentation

A 54-year-old postmenopausal woman with no significant past medical history presents with an abnormal screening mammogram, which shows a focal asymmetry in the 10 o’clock position at middle depth of the left breast. Further work-up with a diagnostic mammogram and ultrasound of the left breast shows a suspicious hypoechoic solid mass with irregular margins measuring 17 mm. The patient undergoes an ultrasound-guided core needle biopsy of the suspicious mass, the results of which are consistent with an invasive ductal carcinoma, Nottingham grade 2, ER strongly positive (95%), PR weakly positive (5%), HER2 negative, and Ki-67 of 15%. She undergoes a left partial mastectomy and sentinel lymph node biopsy, with final pathology demonstrating a single focus of invasive ductal carcinoma, measuring 2.2 cm in greatest dimension with no evidence of lymphovascular invasion. Margins are clear and 2 sentinel lymph nodes are negative for metastatic disease (final pathologic stage IIA, pT2 pN0 cM0). She is referred to medical oncology to discuss adjuvant systemic therapy.

• Can additional testing be used to determine prognosis and guide systemic therapy rec-ommendations for early-stage HR-positive/HER2-negative breast cancer?

After a diagnosis of early-stage breast cancer, the key clinical question faced by the patient and medical oncologist is: what is the individual’s risk for a metastatic breast cancer recurrence and thus the risk for death due to breast cancer? Once the risk for recurrence is established, systemic adjuvant chemotherapy, endocrine therapy, and/or HER2-directed therapy are considered based on the receptor status (ER/PR and HER2) to reduce this risk. Hormone receptor (HR)–positive, HER2-negative breast cancer is the most common type of breast cancer. Although adjuvant endocrine therapy has significantly reduced the risk for recurrence and improved survival for HR-positive breast cancer [6], the role of adjuvant chemotherapy for this subset of breast cancer remains unclear. Prior to genomic testing, the recommendation for adjuvant chemotherapy for HR-positive/HER2-negative tumors was primarily based on patient age and tumor stage and grade. However, chemotherapy overtreatment remained a concern given the potential short- and long-term risks of chemotherapy. Further studies into HR-positive/HER2-negative tumors have shown that these tumors can be divided into 2 main subtypes, luminal A and luminal B [7]. These subtypes represent unique biology and differ in terms of prognosis and response to endocrine therapy and chemotherapy. Luminal A tumors are strongly endocrine responsive and have a good prognosis, while luminal B tumors are less endocrine responsive and are associated with a poorer prognosis; the addition of adjuvant chemotherapy is often considered for luminal B tumors [8]. Several tests, including tumor genomic assays, are now available to help with delineating the tumor subtype and aid in decision-making regarding adjuvant chemotherapy for HR-positive/HER2-negative breast cancers.

Tests for Guiding Adjuvant Chemotherapy Decisions

Ki-67 Assays, Including IHC4 and PEPI

Chronic proliferation is a hallmark of cancer cells [9]. Ki-67, a nuclear nonhistone protein whose expression varies in intensity throughout the cell cycle, has been used as a measurement of tumor cell proliferation [10]. Two large meta-analyses have demonstrated that high Ki-67 expression in breast tumors is independently associated with worse disease-free and overall survival rates [11,12]. Ki-67 expression has also been used to classify HR-positive tumors as luminal A or B. After classifying tumor subtypes based on intrinsic gene expression profiling, Cheang et al determined that a Ki-67 cut point of 13.25% differentiated luminal A and B tumors [13]. However, the ideal cut point for Ki-67 remains unclear, as the sensitivity and specificity in this study was 77% and 78%, respectively. Others have combined Ki-67 with standard ER, PR, and HER2 testing. This IHC4 score, which weighs each of these variables, was validated in postmenopausal patients from the ATAC (Arimidex, Tamoxifen, Alone or in Combination) trial who had ER-positive tumors and did not receive chemotherapy [14]. The prognostic information from the IHC4 was similar to that seen with the 21-gene recurrence score (Oncotype DX), which is discussed later in this article. The key challenge with Ki-67 testing currently is the lack of a validated test methodology, and intraobserver variability in interpreting the Ki-67 results [15]. Recent series have suggested that Ki-67 be considered as a continuous marker rather than a set cut point [16]. These issues continue to impact the clinical utility of Ki-67 for decision making for adjuvant chemotherapy.

Ki-67 and the preoperative endocrine prognostic index (PEPI) score have been explored in the neoadjuvant setting to separate postmenopausal women with endocrine-sensitive versus intrinsically resistant disease and identify patients at risk for recurrent disease [17]. The on-treatment levels of Ki-67 in response to endocrine therapy have been shown to be more prognostic than baseline values, and a decrease in Ki-67 as early as 2 weeks after initiation of neoadjuvant endocrine therapy is associated with endocrine-sensitive tumors and improved outcome. The PEPI score was developed through retrospective analysis of the P024 trial [18] to evaluate the relationship between post-neoadjuvant endocrine therapy tumor characteristics and risk for early relapse. This was subsequently validated in an independent data set from the IMPACT trial [19]. Patients with low pathological stage (0 or 1) and a favorable biomarker profile (PEPI score 0) at surgery had the best prognosis in the absence of chemotherapy. On the other hand, higher pathological stage at surgery and a poor biomarker profile with loss of ER positivity or persistently elevated Ki-67 (PEPI score of 3) identified de novo endocrine-resistant tumors which are at higher risk for early relapse [20]. The ongoing Alliance A011106 ALTERNATE trial (ALTernate approaches for clinical stage II or III Estrogen Receptor positive breast cancer NeoAdjuvant TrEatment in postmenopausal women, NCT01953588) is a phase 3 study to prospectively test this hypothesis.

21-Gene Recurrence Score (Oncotype DX Assay)

The 21-gene Oncotype DX assay is conducted on paraffin-embedded tumor tissue and measures the expression of 16 cancer-related genes and 5 reference genes using quantitative polymerase chain reaction. The genes included in this assay are mainly related to proliferation (including Ki-67), invasion, and HER2 or estrogen signaling [21]. Originally, the 21-gene recurrence score assay was analyzed as a prognostic biomarker tool in a prospective-retrospective biomarker substudy of the National Surgical Adjuvant Breast and Bowel Project (NSABP) B-14 clinical trial in which patients with node-negative, ER-positive tumors were randomly assigned to receive tamoxifen or placebo without chemotherapy [22]. Using the standard reported values of low risk (< 18), intermediate risk (18–30), or high risk (≥ 31) for recurrence, among the tamoxifen-treated patients, cancers with a high-risk recurrence score had a significantly worse rate of distant recurrence and overall survival [21]. Inferior breast cancer survival with a high recurrence score was also confirmed in other series of endocrine-treated patients with node-negative and node-positive disease [23–25].

The predictive utility of the 21-gene recurrence score for endocrine therapy has also been evaluated. A comparison of the placebo- and tamoxifen-treated patients from the NSABP B-14 trial demonstrated that the 21-gene recurrence score predicted benefit from tamoxifen in cancers with low- or intermediate-risk recurrence scores [26]. However, there was no benefit from the use of tamoxifen over placebo in cancers with high-risk recurrence scores. To date, this intriguing data has not been prospectively confirmed, and thus the 21-gene recurrence score is not used to avoid endocrine therapy.

The 21-gene recurrence score is primarily used by oncologists to aid in decision-making regarding adjuvant chemotherapy in patients with node-negative and node-positive (with up to 3 positive lymph nodes), HR-positive/HER2-negative breast cancers. The predictive utility of the 21-gene recurrence score for adjuvant chemotherapy was initially tested using tumor samples from the NSABP B-20 study. This study initially compared adjuvant tamoxifen alone with tamoxifen plus chemotherapy in patients with node-negative, HR-positive tumors. The prospective-retrospective biomarker analysis showed that the patients with high-risk 21-gene recurrence scores benefited from the addition of chemotherapy, whereas those with low- or intermediate-risk did not have an improved freedom from distant recurrence with chemotherapy [27]. Similarly, an analysis from the prospective phase 3 Southwest Oncology Group (SWOG) 8814 trial comparing tamoxifen to tamoxifen with chemotherapy showed that for node-positive tumors, chemotherapy benefit was only seen in those with high 21-gene recurrence scores [24].

Prospective studies are now starting to report results regarding the predictive role of the 21-gene recurrence score. The TAILORx (Trial Assigning Individualized Options for Treatment) trial includes women with node-negative, HR-positive and HER2-negative tumors measuring 0.6 to 5 cm. All patients were treated with standard of care endocrine therapy for at least 5 years. Chemotherapy was determined based on the 21-gene recurrence score results on the primary tumor. The 21-gene recurrence score cutoffs were changed to low (0–10), intermediate (11–25), and high (≥ 26). Patients with scores of 26 or higher were treated with chemotherapy, and those with intermediate scores were randomly assigned to hemotherapy or no chemotherapy; results from this cohort are still pending. However, excellent breast cancer outcomes with endocrine therapy alone were reported from the 1626 (15.9% of total cohort) prospectively followed patients with low-recurrence score tumors. The 5-year invasive disease-free survival was 93.8%, with overall survival of 98% [28]. Given that 5 years is appropriate follow-up to see any chemotherapy benefit, this data supports the recommendation for no chemotherapy in this cohort of patients with very low 21-gene recurrence scores.

The RxPONDER (Rx for Positive Node, Endocrine Responsive Breast Cancer) trial is evaluating women with 1 to 3 node-positive, HR-positive, HER2-negative tumors. In this trial, patients with 21-gene recurrence scores of 0 to 25 were assigned to adjuvant chemotherapy or none. Those with scores of 26 or higher were assigned to chemotherapy. All patients received standard adjuvant endocrine therapy. This study has completed accrual and results are pending. Of note, TAILORx and RxPONDER did not investigate the potential lack of benefit of endocrine therapy in cancers with high recurrence scores. Furthermore, despite data suggesting that chemotherapy may not even benefit women with 4 or more nodes involved but who have a low recurrence score [24], due to the lack of prospective data in this cohort and the quite high risk for distant recurrence, chemotherapy continues to be the standard of care for these patients.

PAM50 (Breast Cancer Prognostic Gene Signature)

Using microarray and quantitative reverse transcriptase PCR (RT-PCR) on formalin-fixed paraffin-embedded (FFPE) tissues, the Breast Cancer Prognostic Gene Signature (PAM50) assay was initially developed to identify intrinsic breast cancer subtypes, including luminal A, luminal B, HER2-enriched, and basal-like [7,29]. Based on the prediction analysis of microarray (PAM) method, the assay measures the expression levels of 50 genes, provides a risk category (low, intermediate, and high), and generates a numerical risk of recurrence score (ROR). The intrinsic subtype and ROR have been shown to add significant prognostic value to the clinicopathological characteristics of tumors. Clinical validity of PAM50 was evaluated in postmenopausal women with HR-positive, early-stage breast cancer treated in the prospective ATAC and ABCSG-8 (Austrian Breast and Colorectal Cancer Study Group 8) trials [30,31]. In 1017 patients with ER-positive breast cancer treated with anastrozole or tamoxifen in the ATAC trial, ROR added significant prognostic information beyond the clinical treatment score (integrated prognostic information from nodal status, tumor size, histopathologic grade, age, and anastrozole or tamoxifen treatment) in all patients. Also, compared with the 21-gene recurrence score, ROR provided more prognostic information in ER-positive, node-negative disease and better differentiation of intermediate- and higher-risk groups. Fewer patients were categorized as intermediate risk by ROR and more as high risk, which could reduce the uncertainty in the estimate of clinical benefit from chemotherapy [30]. The clinical utility of PAM50 as a prognostic model was also validated in 1478 postmenopausal women with ER-positive early-stage breast cancer enrolled in the ABCSG-8 trial. In this study, ROR assigned 47% of patients with node-negative disease to the low-risk category. In this low-risk group, the 10-year metastasis risk was less than 3.5 %, indicating lack of benefit from additional chemotherapy [31]. A key limitation of the PAM50 is the lack of any prospective studies with this assay.

PAM50 has been designed to be carried out in any qualified pathology laboratory. Moreover, the ROR score provides additional prognostic information about risk of late recurrence, which will be discussed in the next section.

70-Gene Breast Cancer Recurrence Assay (MammaPrint)

MammaPrint is a 70-gene assay that was initially developed using an unsupervised, hierarchical clustering algorithm on whole-genome expression arrays with early-stage breast cancer. Among 295 consecutive patients who had MammaPrint testing, those classified with a good-prognosis tumor signature (n = 115) had an excellent 10-year survival rate (94.5%) compared to those with a poor-prognosis signature (54.5%), and the signature remained prognostic upon multivariate analysis [32]. Subsequently, a pooled analysis comparing outcomes by MammaPrint score in patients with node-negative or 1 to 3 node-positive breast cancers treated as per discretion of their medical team with either adjuvant chemotherapy plus endocrine therapy or endocrine therapy alone reported that only those patients with a high-risk score benefited from chemotherapy [33]. Recently, a prospective phase 3 study (MINDACT [Microarray In Node negative Disease may Avoid ChemoTherapy]) evaluating the utility of MammaPrint for adjuvant chemotherapy decision-making reported results [34]. In this study, 6693 women with early-stage breast cancer were assessed by clinical risk and genomic risk using MammaPrint. Those with low clinical and genomic risk did not receive chemotherapy, while those with high clinical and genomic risk all received chemotherapy. The primary goal of the study was to assess whether forgoing chemotherapy would be associated with a low rate of recurrence in those patients with a low-risk prognostic MammaPrint signature but high clinical risk. A total of 1550 patients (23.2%) were in the discordant group, and the majority of these patients had HR-positive disease (98.1%). Without chemotherapy, the rate of survival without distant metastasis at 5 years in this group was 94.7% (95% confidence interval [CI] 92.5% to 96.2%), which met the primary endpoint. Of note, initially, MammaPrint was only available for fresh tissue analysis, but recent advances in RNA processing now allow for this analysis on FFPE tissue [35].

Summary

Case Continued

The patient undergoes 21-gene recurrence score testing, which shows a low recurrence score of 10, estimating the 10-year risk of distant recurrence to be approximately 7% with 5 years of tamoxifen. Chemo-therapy is not recommended. The patient completes adjuvant whole breast radiation therapy, and then, based on data supporting AIs over tamoxifen in postmenopausal women, she is started on anastrozole [36]. She initially experiences mild side effects from treatment, including fatigue, arthralgia, and vaginal dryness, but her symptoms are able to be managed. As she approaches 5 years of adjuvant endocrine therapy with anastrozole, she is struggling with rotator cuff injury and is anxious about recurrence, but has no evidence of recurrent cancer. Her bone density scan in the beginning of her fourth year of therapy shows a decrease in bone mineral density, with the lowest T score of –1.5 at the left femoral neck, consistent with osteopenia. She has been treated with calcium and vitamin D supplements.

• How long should this patient continue treatment with anastrozole?

The risk for recurrence is highest during the first 5 years after diagnosis for all patients with early breast cancer [37]. Although HR-positive breast cancers have a better prognosis than HR-negative disease, the pattern of recurrence is different between the 2 groups, and it is estimated that approximately half of the recurrences among patients with HR-positive early breast cancer occur after the first 5 years from diagnosis. Annualized hazard of recurrence in HR-positive breast cancer has been shown to remain elevated and fairly stable beyond 10 years, even for those with low tumor burden and node-negative disease [38]. Prospective trials showed that for women with HR-positive early breast cancer, 5 years of adjuvant tamoxifen could substantially reduce recurrence rates and improve survival, and this became the standard of care [39]. AIs are considered the standard of care for adjuvant endocrine therapy in most postmenopausal women, as they result in a significantly lower recurrence rate compared with tamoxifen, either as initial adjuvant therapy or sequentially following 2 to 3 years of tamoxifen [40].

However, extending AI therapy from 5 years to 10 years is not clearly beneficial. In the MA.17R trial, although longer AI therapy resulted in significantly better disease-free survival (95% versus 91%, hazard ratio 0.66; P = 0.01), this was primarily due to a lower incidence of contralateral breast cancer in those taking the AI compared with placebo. The distant recurrence risks were similar and low (4.4% versus 5.5%), and there was no overall survival difference [2]. Also, the NSABP B-42 study, which was presented at the 2016 San Antonio Breast Cancer Symposium, did not meet its predefined endpoint for benefit from extending adjuvant AI therapy with letrozole beyond 5 years [3]. Thus, the absolute benefit from extended endocrine therapy has been modest across these studies. Although endocrine therapy is considered relatively safe and well tolerated, side effects can be significant and even associated with morbidity. Ideally, extended endocrine therapy should be offered to the subset of patients who would benefit the most. Several genomic diagnostic assays, including the EndoPredict test, PAM50, and the Breast Cancer Index (BCI) tests, specifically assess the risk for late recurrence in HR-positive cancers.

Tests for Assessing Risk for Late Recurrence

PAM50

Studies suggest that the ROR score also has value in predicting late recurrences. Analysis of data in patients enrolled in the ABCSG-8 trial showed that ROR could identify patients with endocrine-sensitive disease who are at low risk for late relapse and could be spared from unwanted toxicities of extended endocrine therapies. In 1246 ABCSG-8 patients between years 5 and 15, the PAM50 ROR demonstrated an absolute risk of distant recurrence of 2.4% in the low-risk group, as compared with 17.5% in the high-risk group [44]. Also, a combined analysis of patients from both the ATAC and ABCSG-8 trials demonstrated the utility of ROR in identifying this subgroup of patients with low risk for late relapse [45].

EndoPredict

EndoPredict (EP) is another quantitative RT-PCR–based assay which uses FFPE tissues to calculate a risk score based on 8 cancer-related and 3 reference genes. The score is combined with clinicopathological factors including tumor size and nodal status to make a comprehensive risk score (EPclin). EPclin is used to dichotomize patients into EP low- and EP high-risk groups. EP has been validated in 2 cohorts of patients enrolled in separate randomized studies, ABCSG-6 and ABCSG-8. EP provided prognostic information beyond clinicopathological variables to predict distant recurrence in patients with HR-positive, HER2-negative early breast cancer [46]. More important, EP has been shown to predict early (years 0–5) versus late (> 5 years after diagnosis) recurrences and identify a low-risk subset of patients who would not be expected to benefit from further treatment beyond 5 years of endocrine therapy [47]. Recently, EP and EPclin were compared with the 21-gene (Oncotype DX) recurrence score in a patient population from the TransATAC study. Both EP and EPclin provided more prognostic information compared to the 21-gene recurrence score and identified early and late relapse events [48]. EndoPredict is the first multigene expression assay that could be routinely performed in decentral molecular pathological laboratories with a short turnaround time [49].

Breast Cancer Index

The BCI is a RT-PCR–based gene expression assay that consists of 2 gene expression biomarkers: molecular grade index (MGI) and HOXB13/IL17BR (H/I). The BCI was developed as a prognostic test to assess risk for breast cancer recurrence using a cohort of ER-positive patients (n = 588) treated with adjuvant tamoxifen versus observation from the prospective randomized Stockholm trial [50]. In this blinded retrospective study, H/I and MGI were measured and a continuous risk model (BCI) was developed in the tamoxifen-treated group. More than 50% of the patients in this group were classified as having a low risk of recurrence. The rate of distant recurrence or death in this low-risk group at 10 years was less than 3%. The performance of the BCI model was then tested in the untreated arm of the Stockholm trial. In the untreated arm, BCI classified 53%, 27%, and 20% of patients as low, intermediate, and high risk, respectively. The rate of distant metastasis at 10 years in these risk groups was 8.3% (95% CI 4.7% to 14.4%), 22.9% (95% CI 14.5% to 35.2%), and 28.5% (95% CI 17.9% to 43.6%), respectively, and the rate of breast cancer–specific mortality was 5.1% (95% CI 1.3% to 8.7%), 19.8% (95% CI 10.0% to 28.6%), and 28.8% (95% CI 15.3% to 40.2%) [50].

The prognostic and predictive values of the BCI have been validated in other large, randomized studies and in patients with both node-negative and node-positive disease [51,52]. The predictive value of the endocrine-response biomarker, the H/I ratio, has been demonstrated in randomized studies. In the MA.17 trial, a high H/I ratio was associated with increased risk for late recurrence in the absence of letrozole. However, extended endocrine therapy with letrozole in patients with high H/I ratios predicted benefit from therapy and decreased the probability of late disease recurrence [53]. BCI was also compared to IHC4 and the 21-gene recurrence score in the TransATAC study and was the only test to show prognostic significance for both early (0–5 years) and late (5–10 year) recurrence [54].

The impact of the BCI results on physicians’ recommendations for extended endocrine therapy was assessed by a prospective study. This study showed that the test result had a significant effect on both physician treatment recommendation and patient satisfaction. BCI testing resulted in a change in physician recommendations for extended endocrine therapy, with an overall decrease in recommendations for extended endocrine therapy from 74% to 54%. Knowledge of the test result also led to improved patient satisfaction and decreased anxiety [55].

Summary

Due to the risk for late recurrence, extended endocrine therapy is being recommended for many patients with HR-positive breast cancers. Multiple genomic assays are being developed to better understand an individual’s risk for late recurrence and the potential for benefit from extended endocrine therapies. However, none of the assays have been validated in prospective randomized studies. Further validation is needed prior to routine use of these assays.

Case Continued

A BCI test is done and the result shows 4.3% BCI low-risk category in years 5–10; low likelihood of benefit from extended endocrine therapy. After discussing the results of the BCI test in the context of no survival benefit from extending AIs beyond 5 years, both the patient and her oncologist feel comfortable with discontinuing endocrine therapy at the end of 5 years.

Conclusion

Reduction in breast cancer mortality is mainly the result of improved systemic treatments. With advances in breast cancer screening tools in recent years, the rate of cancer detection has increased. This has raised concerns regarding overdiagnosis. To prevent unwanted toxicities associated with overtreatment, better treatment decision tools are needed. Several genomic assays are currently available and widely used to provide prognostic and predictive information and aid in decisions regarding appropriate use of adjuvant chemotherapy in HR-positive/HER2-negative early-stage breast cancer. Ongoing studies are refining the cutoffs for these assays and expanding the applicability to node-positive breast cancers. Furthermore, with several studies now showing benefit from the use of extended endocrine therapy, some of these assays may be able to identify the subset of patients who are at increased risk for late recurrence and who might benefit from extended endocrine therapy. Advances in molecular testing has enabled clinicians to offer more personalized treatments to their patients, improve patient’s compliance, and decrease anxiety and conflict associated with management decisions. Although small numbers of patients with HER2-positive and triple negative breast cancers were also included in some of these studies, use of genomic assays in this subset of patients is very limited and currently not recommended.

Corresponding author: Kari Braun Wisinski, MD, 1111 Highland Avenue, 6033 Wisconsin Institute for Medical Research, Madison, WI 53705-2275, kbwisinski@medicine.wisc.edu.

Financial disclosures: This work was supported by the NCI Cancer Center Support Grant P30 CA014520.

From the University of Arizona Cancer Center, Tucson, AZ (Dr. Ehsani), and University of Wisconsin Carbone Cancer Center and School of Medicine and Public Health, Madison, WI (Dr. Wisinski).

Abstract

• Objectives: To describe common genomic tests being used clinically to assess prognosis and guide adjuvant chemotherapy and endocrine therapy decisions for early-stage breast cancer.
• Methods: Case presentation and review of the literature.
• Results: Hormone receptor–positive (HR-positive) breast cancers, which express the estrogen and/or progesterone receptor, account for the majority of breast cancers. Endocrine therapy can be highly effective for patients with these HR-positive tumors, and identification of HR-positive breast cancers that do not require the addition of chemotherapy is critical. Clinicopathological features of the breast cancer, including tumor size, nodal involvement, grading, and HR status, are insufficient in predicting the risk for recurrence or the need for chemotherapy. Furthermore, a portion of HR-positive breast cancers have an ongoing risk for late recurrence, and longer durations of endocrine therapy are being used to reduce this risk.
• Conclusion: There is sufficient evidence for use of genomic testing in early-stage HR-positive breast cancer to aid in chemotherapy recommendations. Further confirmation of genomic assays for prediction of benefit from prolonged endocrine therapy is needed.

Key words: molecular testing; decision aids; HR-positive cancer; recurrence risk; adjuvant chemotherapy; endocrine therapy.

Despite the increase in incidence of breast cancer, breast cancer mortality has decreased over the past several decades. This is likely due to both early detection and advances in systemic therapy. However, with more widespread use of screening mammography, there are increasing concerns regarding potential overdiagnosis of cancer [1]. One key challenge is that breast cancer is a heterogeneous disease. Thus, improved tools for determining breast cancer biology can help physicians individualize treatments, with low-risk cancers approached with less aggressive treatments, thus preventing unnecessary toxicities, and higher-risk cancers treated appropriately.

Traditionally, adjuvant chemotherapy was recommended based on tumor features such as stage (tumor size, regional nodal involvement), grade, expression of hormone receptors (estrogen receptor [ER] and progesterone receptor [PR]) and human epidermal growth factor receptor-2 (HER2), and patient features (age, menopausal status). However, this approach is not accurate enough to guide individualized treatment recommendations, which are based on the risk for recurrence and the reduction in this risk that can be achieved with various systemic treatments. In particular, there are individuals with low-risk HR-positive, HER2-negative breast cancers who could be spared the toxicities of cytotoxic chemotherapies without compromising the prognosis.

Beyond chemotherapy, endocrine therapies also have risks, especially when given for extended durations. Recently, extended endocrine therapy has been shown to prevent late recurrences of HR-positive breast cancers. In the MA.17R study, extended endocrine therapy with letrozole for a total of 10 years (beyond 5 years of an aromatase inhibitor [AI]) decreased the risk for breast cancer recurrence or the occurrence of contralateral breast cancer by 34% [2]. However, the overall survival was similar between the 2 groups and the results were not confirmed in other studies [3–5]. Identifying the subgroup of patients who benefit from this extended AI therapy is important in the era of personalized medicine. Several tumor genomic assays have been developed to provide additional prognostic and predictive information with the goal of individualizing adjuvant therapies for breast cancer. Although assays are also being evaluated in HER2-positive and triple negative breast cancer, this review will focus on HR-positive, HER2-negative breast cancer.

Case Study

Initial Presentation

A 54-year-old postmenopausal woman with no significant past medical history presents with an abnormal screening mammogram, which shows a focal asymmetry in the 10 o’clock position at middle depth of the left breast. Further work-up with a diagnostic mammogram and ultrasound of the left breast shows a suspicious hypoechoic solid mass with irregular margins measuring 17 mm. The patient undergoes an ultrasound-guided core needle biopsy of the suspicious mass, the results of which are consistent with an invasive ductal carcinoma, Nottingham grade 2, ER strongly positive (95%), PR weakly positive (5%), HER2 negative, and Ki-67 of 15%. She undergoes a left partial mastectomy and sentinel lymph node biopsy, with final pathology demonstrating a single focus of invasive ductal carcinoma, measuring 2.2 cm in greatest dimension with no evidence of lymphovascular invasion. Margins are clear and 2 sentinel lymph nodes are negative for metastatic disease (final pathologic stage IIA, pT2 pN0 cM0). She is referred to medical oncology to discuss adjuvant systemic therapy.

• Can additional testing be used to determine prognosis and guide systemic therapy rec-ommendations for early-stage HR-positive/HER2-negative breast cancer?

After a diagnosis of early-stage breast cancer, the key clinical question faced by the patient and medical oncologist is: what is the individual’s risk for a metastatic breast cancer recurrence and thus the risk for death due to breast cancer? Once the risk for recurrence is established, systemic adjuvant chemotherapy, endocrine therapy, and/or HER2-directed therapy are considered based on the receptor status (ER/PR and HER2) to reduce this risk. Hormone receptor (HR)–positive, HER2-negative breast cancer is the most common type of breast cancer. Although adjuvant endocrine therapy has significantly reduced the risk for recurrence and improved survival for HR-positive breast cancer [6], the role of adjuvant chemotherapy for this subset of breast cancer remains unclear. Prior to genomic testing, the recommendation for adjuvant chemotherapy for HR-positive/HER2-negative tumors was primarily based on patient age and tumor stage and grade. However, chemotherapy overtreatment remained a concern given the potential short- and long-term risks of chemotherapy. Further studies into HR-positive/HER2-negative tumors have shown that these tumors can be divided into 2 main subtypes, luminal A and luminal B [7]. These subtypes represent unique biology and differ in terms of prognosis and response to endocrine therapy and chemotherapy. Luminal A tumors are strongly endocrine responsive and have a good prognosis, while luminal B tumors are less endocrine responsive and are associated with a poorer prognosis; the addition of adjuvant chemotherapy is often considered for luminal B tumors [8]. Several tests, including tumor genomic assays, are now available to help with delineating the tumor subtype and aid in decision-making regarding adjuvant chemotherapy for HR-positive/HER2-negative breast cancers.

Tests for Guiding Adjuvant Chemotherapy Decisions

Ki-67 Assays, Including IHC4 and PEPI

Chronic proliferation is a hallmark of cancer cells [9]. Ki-67, a nuclear nonhistone protein whose expression varies in intensity throughout the cell cycle, has been used as a measurement of tumor cell proliferation [10]. Two large meta-analyses have demonstrated that high Ki-67 expression in breast tumors is independently associated with worse disease-free and overall survival rates [11,12]. Ki-67 expression has also been used to classify HR-positive tumors as luminal A or B. After classifying tumor subtypes based on intrinsic gene expression profiling, Cheang et al determined that a Ki-67 cut point of 13.25% differentiated luminal A and B tumors [13]. However, the ideal cut point for Ki-67 remains unclear, as the sensitivity and specificity in this study was 77% and 78%, respectively. Others have combined Ki-67 with standard ER, PR, and HER2 testing. This IHC4 score, which weighs each of these variables, was validated in postmenopausal patients from the ATAC (Arimidex, Tamoxifen, Alone or in Combination) trial who had ER-positive tumors and did not receive chemotherapy [14]. The prognostic information from the IHC4 was similar to that seen with the 21-gene recurrence score (Oncotype DX), which is discussed later in this article. The key challenge with Ki-67 testing currently is the lack of a validated test methodology, and intraobserver variability in interpreting the Ki-67 results [15]. Recent series have suggested that Ki-67 be considered as a continuous marker rather than a set cut point [16]. These issues continue to impact the clinical utility of Ki-67 for decision making for adjuvant chemotherapy.

Ki-67 and the preoperative endocrine prognostic index (PEPI) score have been explored in the neoadjuvant setting to separate postmenopausal women with endocrine-sensitive versus intrinsically resistant disease and identify patients at risk for recurrent disease [17]. The on-treatment levels of Ki-67 in response to endocrine therapy have been shown to be more prognostic than baseline values, and a decrease in Ki-67 as early as 2 weeks after initiation of neoadjuvant endocrine therapy is associated with endocrine-sensitive tumors and improved outcome. The PEPI score was developed through retrospective analysis of the P024 trial [18] to evaluate the relationship between post-neoadjuvant endocrine therapy tumor characteristics and risk for early relapse. This was subsequently validated in an independent data set from the IMPACT trial [19]. Patients with low pathological stage (0 or 1) and a favorable biomarker profile (PEPI score 0) at surgery had the best prognosis in the absence of chemotherapy. On the other hand, higher pathological stage at surgery and a poor biomarker profile with loss of ER positivity or persistently elevated Ki-67 (PEPI score of 3) identified de novo endocrine-resistant tumors which are at higher risk for early relapse [20]. The ongoing Alliance A011106 ALTERNATE trial (ALTernate approaches for clinical stage II or III Estrogen Receptor positive breast cancer NeoAdjuvant TrEatment in postmenopausal women, NCT01953588) is a phase 3 study to prospectively test this hypothesis.

21-Gene Recurrence Score (Oncotype DX Assay)

The 21-gene Oncotype DX assay is conducted on paraffin-embedded tumor tissue and measures the expression of 16 cancer-related genes and 5 reference genes using quantitative polymerase chain reaction. The genes included in this assay are mainly related to proliferation (including Ki-67), invasion, and HER2 or estrogen signaling [21]. Originally, the 21-gene recurrence score assay was analyzed as a prognostic biomarker tool in a prospective-retrospective biomarker substudy of the National Surgical Adjuvant Breast and Bowel Project (NSABP) B-14 clinical trial in which patients with node-negative, ER-positive tumors were randomly assigned to receive tamoxifen or placebo without chemotherapy [22]. Using the standard reported values of low risk (< 18), intermediate risk (18–30), or high risk (≥ 31) for recurrence, among the tamoxifen-treated patients, cancers with a high-risk recurrence score had a significantly worse rate of distant recurrence and overall survival [21]. Inferior breast cancer survival with a high recurrence score was also confirmed in other series of endocrine-treated patients with node-negative and node-positive disease [23–25].

The predictive utility of the 21-gene recurrence score for endocrine therapy has also been evaluated. A comparison of the placebo- and tamoxifen-treated patients from the NSABP B-14 trial demonstrated that the 21-gene recurrence score predicted benefit from tamoxifen in cancers with low- or intermediate-risk recurrence scores [26]. However, there was no benefit from the use of tamoxifen over placebo in cancers with high-risk recurrence scores. To date, this intriguing data has not been prospectively confirmed, and thus the 21-gene recurrence score is not used to avoid endocrine therapy.

The 21-gene recurrence score is primarily used by oncologists to aid in decision-making regarding adjuvant chemotherapy in patients with node-negative and node-positive (with up to 3 positive lymph nodes), HR-positive/HER2-negative breast cancers. The predictive utility of the 21-gene recurrence score for adjuvant chemotherapy was initially tested using tumor samples from the NSABP B-20 study. This study initially compared adjuvant tamoxifen alone with tamoxifen plus chemotherapy in patients with node-negative, HR-positive tumors. The prospective-retrospective biomarker analysis showed that the patients with high-risk 21-gene recurrence scores benefited from the addition of chemotherapy, whereas those with low- or intermediate-risk did not have an improved freedom from distant recurrence with chemotherapy [27]. Similarly, an analysis from the prospective phase 3 Southwest Oncology Group (SWOG) 8814 trial comparing tamoxifen to tamoxifen with chemotherapy showed that for node-positive tumors, chemotherapy benefit was only seen in those with high 21-gene recurrence scores [24].

Prospective studies are now starting to report results regarding the predictive role of the 21-gene recurrence score. The TAILORx (Trial Assigning Individualized Options for Treatment) trial includes women with node-negative, HR-positive and HER2-negative tumors measuring 0.6 to 5 cm. All patients were treated with standard of care endocrine therapy for at least 5 years. Chemotherapy was determined based on the 21-gene recurrence score results on the primary tumor. The 21-gene recurrence score cutoffs were changed to low (0–10), intermediate (11–25), and high (≥ 26). Patients with scores of 26 or higher were treated with chemotherapy, and those with intermediate scores were randomly assigned to hemotherapy or no chemotherapy; results from this cohort are still pending. However, excellent breast cancer outcomes with endocrine therapy alone were reported from the 1626 (15.9% of total cohort) prospectively followed patients with low-recurrence score tumors. The 5-year invasive disease-free survival was 93.8%, with overall survival of 98% [28]. Given that 5 years is appropriate follow-up to see any chemotherapy benefit, this data supports the recommendation for no chemotherapy in this cohort of patients with very low 21-gene recurrence scores.

The RxPONDER (Rx for Positive Node, Endocrine Responsive Breast Cancer) trial is evaluating women with 1 to 3 node-positive, HR-positive, HER2-negative tumors. In this trial, patients with 21-gene recurrence scores of 0 to 25 were assigned to adjuvant chemotherapy or none. Those with scores of 26 or higher were assigned to chemotherapy. All patients received standard adjuvant endocrine therapy. This study has completed accrual and results are pending. Of note, TAILORx and RxPONDER did not investigate the potential lack of benefit of endocrine therapy in cancers with high recurrence scores. Furthermore, despite data suggesting that chemotherapy may not even benefit women with 4 or more nodes involved but who have a low recurrence score [24], due to the lack of prospective data in this cohort and the quite high risk for distant recurrence, chemotherapy continues to be the standard of care for these patients.

PAM50 (Breast Cancer Prognostic Gene Signature)

Using microarray and quantitative reverse transcriptase PCR (RT-PCR) on formalin-fixed paraffin-embedded (FFPE) tissues, the Breast Cancer Prognostic Gene Signature (PAM50) assay was initially developed to identify intrinsic breast cancer subtypes, including luminal A, luminal B, HER2-enriched, and basal-like [7,29]. Based on the prediction analysis of microarray (PAM) method, the assay measures the expression levels of 50 genes, provides a risk category (low, intermediate, and high), and generates a numerical risk of recurrence score (ROR). The intrinsic subtype and ROR have been shown to add significant prognostic value to the clinicopathological characteristics of tumors. Clinical validity of PAM50 was evaluated in postmenopausal women with HR-positive, early-stage breast cancer treated in the prospective ATAC and ABCSG-8 (Austrian Breast and Colorectal Cancer Study Group 8) trials [30,31]. In 1017 patients with ER-positive breast cancer treated with anastrozole or tamoxifen in the ATAC trial, ROR added significant prognostic information beyond the clinical treatment score (integrated prognostic information from nodal status, tumor size, histopathologic grade, age, and anastrozole or tamoxifen treatment) in all patients. Also, compared with the 21-gene recurrence score, ROR provided more prognostic information in ER-positive, node-negative disease and better differentiation of intermediate- and higher-risk groups. Fewer patients were categorized as intermediate risk by ROR and more as high risk, which could reduce the uncertainty in the estimate of clinical benefit from chemotherapy [30]. The clinical utility of PAM50 as a prognostic model was also validated in 1478 postmenopausal women with ER-positive early-stage breast cancer enrolled in the ABCSG-8 trial. In this study, ROR assigned 47% of patients with node-negative disease to the low-risk category. In this low-risk group, the 10-year metastasis risk was less than 3.5 %, indicating lack of benefit from additional chemotherapy [31]. A key limitation of the PAM50 is the lack of any prospective studies with this assay.

PAM50 has been designed to be carried out in any qualified pathology laboratory. Moreover, the ROR score provides additional prognostic information about risk of late recurrence, which will be discussed in the next section.

70-Gene Breast Cancer Recurrence Assay (MammaPrint)

MammaPrint is a 70-gene assay that was initially developed using an unsupervised, hierarchical clustering algorithm on whole-genome expression arrays with early-stage breast cancer. Among 295 consecutive patients who had MammaPrint testing, those classified with a good-prognosis tumor signature (n = 115) had an excellent 10-year survival rate (94.5%) compared to those with a poor-prognosis signature (54.5%), and the signature remained prognostic upon multivariate analysis [32]. Subsequently, a pooled analysis comparing outcomes by MammaPrint score in patients with node-negative or 1 to 3 node-positive breast cancers treated as per discretion of their medical team with either adjuvant chemotherapy plus endocrine therapy or endocrine therapy alone reported that only those patients with a high-risk score benefited from chemotherapy [33]. Recently, a prospective phase 3 study (MINDACT [Microarray In Node negative Disease may Avoid ChemoTherapy]) evaluating the utility of MammaPrint for adjuvant chemotherapy decision-making reported results [34]. In this study, 6693 women with early-stage breast cancer were assessed by clinical risk and genomic risk using MammaPrint. Those with low clinical and genomic risk did not receive chemotherapy, while those with high clinical and genomic risk all received chemotherapy. The primary goal of the study was to assess whether forgoing chemotherapy would be associated with a low rate of recurrence in those patients with a low-risk prognostic MammaPrint signature but high clinical risk. A total of 1550 patients (23.2%) were in the discordant group, and the majority of these patients had HR-positive disease (98.1%). Without chemotherapy, the rate of survival without distant metastasis at 5 years in this group was 94.7% (95% confidence interval [CI] 92.5% to 96.2%), which met the primary endpoint. Of note, initially, MammaPrint was only available for fresh tissue analysis, but recent advances in RNA processing now allow for this analysis on FFPE tissue [35].

Summary

Case Continued

The patient undergoes 21-gene recurrence score testing, which shows a low recurrence score of 10, estimating the 10-year risk of distant recurrence to be approximately 7% with 5 years of tamoxifen. Chemo-therapy is not recommended. The patient completes adjuvant whole breast radiation therapy, and then, based on data supporting AIs over tamoxifen in postmenopausal women, she is started on anastrozole [36]. She initially experiences mild side effects from treatment, including fatigue, arthralgia, and vaginal dryness, but her symptoms are able to be managed. As she approaches 5 years of adjuvant endocrine therapy with anastrozole, she is struggling with rotator cuff injury and is anxious about recurrence, but has no evidence of recurrent cancer. Her bone density scan in the beginning of her fourth year of therapy shows a decrease in bone mineral density, with the lowest T score of –1.5 at the left femoral neck, consistent with osteopenia. She has been treated with calcium and vitamin D supplements.

• How long should this patient continue treatment with anastrozole?

The risk for recurrence is highest during the first 5 years after diagnosis for all patients with early breast cancer [37]. Although HR-positive breast cancers have a better prognosis than HR-negative disease, the pattern of recurrence is different between the 2 groups, and it is estimated that approximately half of the recurrences among patients with HR-positive early breast cancer occur after the first 5 years from diagnosis. Annualized hazard of recurrence in HR-positive breast cancer has been shown to remain elevated and fairly stable beyond 10 years, even for those with low tumor burden and node-negative disease [38]. Prospective trials showed that for women with HR-positive early breast cancer, 5 years of adjuvant tamoxifen could substantially reduce recurrence rates and improve survival, and this became the standard of care [39]. AIs are considered the standard of care for adjuvant endocrine therapy in most postmenopausal women, as they result in a significantly lower recurrence rate compared with tamoxifen, either as initial adjuvant therapy or sequentially following 2 to 3 years of tamoxifen [40].

However, extending AI therapy from 5 years to 10 years is not clearly beneficial. In the MA.17R trial, although longer AI therapy resulted in significantly better disease-free survival (95% versus 91%, hazard ratio 0.66; P = 0.01), this was primarily due to a lower incidence of contralateral breast cancer in those taking the AI compared with placebo. The distant recurrence risks were similar and low (4.4% versus 5.5%), and there was no overall survival difference [2]. Also, the NSABP B-42 study, which was presented at the 2016 San Antonio Breast Cancer Symposium, did not meet its predefined endpoint for benefit from extending adjuvant AI therapy with letrozole beyond 5 years [3]. Thus, the absolute benefit from extended endocrine therapy has been modest across these studies. Although endocrine therapy is considered relatively safe and well tolerated, side effects can be significant and even associated with morbidity. Ideally, extended endocrine therapy should be offered to the subset of patients who would benefit the most. Several genomic diagnostic assays, including the EndoPredict test, PAM50, and the Breast Cancer Index (BCI) tests, specifically assess the risk for late recurrence in HR-positive cancers.

Tests for Assessing Risk for Late Recurrence

PAM50

Studies suggest that the ROR score also has value in predicting late recurrences. Analysis of data in patients enrolled in the ABCSG-8 trial showed that ROR could identify patients with endocrine-sensitive disease who are at low risk for late relapse and could be spared from unwanted toxicities of extended endocrine therapies. In 1246 ABCSG-8 patients between years 5 and 15, the PAM50 ROR demonstrated an absolute risk of distant recurrence of 2.4% in the low-risk group, as compared with 17.5% in the high-risk group [44]. Also, a combined analysis of patients from both the ATAC and ABCSG-8 trials demonstrated the utility of ROR in identifying this subgroup of patients with low risk for late relapse [45].

EndoPredict

EndoPredict (EP) is another quantitative RT-PCR–based assay which uses FFPE tissues to calculate a risk score based on 8 cancer-related and 3 reference genes. The score is combined with clinicopathological factors including tumor size and nodal status to make a comprehensive risk score (EPclin). EPclin is used to dichotomize patients into EP low- and EP high-risk groups. EP has been validated in 2 cohorts of patients enrolled in separate randomized studies, ABCSG-6 and ABCSG-8. EP provided prognostic information beyond clinicopathological variables to predict distant recurrence in patients with HR-positive, HER2-negative early breast cancer [46]. More important, EP has been shown to predict early (years 0–5) versus late (> 5 years after diagnosis) recurrences and identify a low-risk subset of patients who would not be expected to benefit from further treatment beyond 5 years of endocrine therapy [47]. Recently, EP and EPclin were compared with the 21-gene (Oncotype DX) recurrence score in a patient population from the TransATAC study. Both EP and EPclin provided more prognostic information compared to the 21-gene recurrence score and identified early and late relapse events [48]. EndoPredict is the first multigene expression assay that could be routinely performed in decentral molecular pathological laboratories with a short turnaround time [49].

Breast Cancer Index

The BCI is a RT-PCR–based gene expression assay that consists of 2 gene expression biomarkers: molecular grade index (MGI) and HOXB13/IL17BR (H/I). The BCI was developed as a prognostic test to assess risk for breast cancer recurrence using a cohort of ER-positive patients (n = 588) treated with adjuvant tamoxifen versus observation from the prospective randomized Stockholm trial [50]. In this blinded retrospective study, H/I and MGI were measured and a continuous risk model (BCI) was developed in the tamoxifen-treated group. More than 50% of the patients in this group were classified as having a low risk of recurrence. The rate of distant recurrence or death in this low-risk group at 10 years was less than 3%. The performance of the BCI model was then tested in the untreated arm of the Stockholm trial. In the untreated arm, BCI classified 53%, 27%, and 20% of patients as low, intermediate, and high risk, respectively. The rate of distant metastasis at 10 years in these risk groups was 8.3% (95% CI 4.7% to 14.4%), 22.9% (95% CI 14.5% to 35.2%), and 28.5% (95% CI 17.9% to 43.6%), respectively, and the rate of breast cancer–specific mortality was 5.1% (95% CI 1.3% to 8.7%), 19.8% (95% CI 10.0% to 28.6%), and 28.8% (95% CI 15.3% to 40.2%) [50].

The prognostic and predictive values of the BCI have been validated in other large, randomized studies and in patients with both node-negative and node-positive disease [51,52]. The predictive value of the endocrine-response biomarker, the H/I ratio, has been demonstrated in randomized studies. In the MA.17 trial, a high H/I ratio was associated with increased risk for late recurrence in the absence of letrozole. However, extended endocrine therapy with letrozole in patients with high H/I ratios predicted benefit from therapy and decreased the probability of late disease recurrence [53]. BCI was also compared to IHC4 and the 21-gene recurrence score in the TransATAC study and was the only test to show prognostic significance for both early (0–5 years) and late (5–10 year) recurrence [54].

The impact of the BCI results on physicians’ recommendations for extended endocrine therapy was assessed by a prospective study. This study showed that the test result had a significant effect on both physician treatment recommendation and patient satisfaction. BCI testing resulted in a change in physician recommendations for extended endocrine therapy, with an overall decrease in recommendations for extended endocrine therapy from 74% to 54%. Knowledge of the test result also led to improved patient satisfaction and decreased anxiety [55].

Summary

Due to the risk for late recurrence, extended endocrine therapy is being recommended for many patients with HR-positive breast cancers. Multiple genomic assays are being developed to better understand an individual’s risk for late recurrence and the potential for benefit from extended endocrine therapies. However, none of the assays have been validated in prospective randomized studies. Further validation is needed prior to routine use of these assays.

Case Continued

A BCI test is done and the result shows 4.3% BCI low-risk category in years 5–10; low likelihood of benefit from extended endocrine therapy. After discussing the results of the BCI test in the context of no survival benefit from extending AIs beyond 5 years, both the patient and her oncologist feel comfortable with discontinuing endocrine therapy at the end of 5 years.

Conclusion

Reduction in breast cancer mortality is mainly the result of improved systemic treatments. With advances in breast cancer screening tools in recent years, the rate of cancer detection has increased. This has raised concerns regarding overdiagnosis. To prevent unwanted toxicities associated with overtreatment, better treatment decision tools are needed. Several genomic assays are currently available and widely used to provide prognostic and predictive information and aid in decisions regarding appropriate use of adjuvant chemotherapy in HR-positive/HER2-negative early-stage breast cancer. Ongoing studies are refining the cutoffs for these assays and expanding the applicability to node-positive breast cancers. Furthermore, with several studies now showing benefit from the use of extended endocrine therapy, some of these assays may be able to identify the subset of patients who are at increased risk for late recurrence and who might benefit from extended endocrine therapy. Advances in molecular testing has enabled clinicians to offer more personalized treatments to their patients, improve patient’s compliance, and decrease anxiety and conflict associated with management decisions. Although small numbers of patients with HER2-positive and triple negative breast cancers were also included in some of these studies, use of genomic assays in this subset of patients is very limited and currently not recommended.

Corresponding author: Kari Braun Wisinski, MD, 1111 Highland Avenue, 6033 Wisconsin Institute for Medical Research, Madison, WI 53705-2275, kbwisinski@medicine.wisc.edu.

Financial disclosures: This work was supported by the NCI Cancer Center Support Grant P30 CA014520.

From the University of Arizona Cancer Center, Tucson, AZ (Dr. Ehsani), and University of Wisconsin Carbone Cancer Center and School of Medicine and Public Health, Madison, WI (Dr. Wisinski).

Abstract

• Objectives: To describe common genomic tests being used clinically to assess prognosis and guide adjuvant chemotherapy and endocrine therapy decisions for early-stage breast cancer.
• Methods: Case presentation and review of the literature.
• Results: Hormone receptor–positive (HR-positive) breast cancers, which express the estrogen and/or progesterone receptor, account for the majority of breast cancers. Endocrine therapy can be highly effective for patients with these HR-positive tumors, and identification of HR-positive breast cancers that do not require the addition of chemotherapy is critical. Clinicopathological features of the breast cancer, including tumor size, nodal involvement, grading, and HR status, are insufficient in predicting the risk for recurrence or the need for chemotherapy. Furthermore, a portion of HR-positive breast cancers have an ongoing risk for late recurrence, and longer durations of endocrine therapy are being used to reduce this risk.
• Conclusion: There is sufficient evidence for use of genomic testing in early-stage HR-positive breast cancer to aid in chemotherapy recommendations. Further confirmation of genomic assays for prediction of benefit from prolonged endocrine therapy is needed.

Key words: molecular testing; decision aids; HR-positive cancer; recurrence risk; adjuvant chemotherapy; endocrine therapy.

Despite the increase in incidence of breast cancer, breast cancer mortality has decreased over the past several decades. This is likely due to both early detection and advances in systemic therapy. However, with more widespread use of screening mammography, there are increasing concerns regarding potential overdiagnosis of cancer [1]. One key challenge is that breast cancer is a heterogeneous disease. Thus, improved tools for determining breast cancer biology can help physicians individualize treatments, with low-risk cancers approached with less aggressive treatments, thus preventing unnecessary toxicities, and higher-risk cancers treated appropriately.

Traditionally, adjuvant chemotherapy was recommended based on tumor features such as stage (tumor size, regional nodal involvement), grade, expression of hormone receptors (estrogen receptor [ER] and progesterone receptor [PR]) and human epidermal growth factor receptor-2 (HER2), and patient features (age, menopausal status). However, this approach is not accurate enough to guide individualized treatment recommendations, which are based on the risk for recurrence and the reduction in this risk that can be achieved with various systemic treatments. In particular, there are individuals with low-risk HR-positive, HER2-negative breast cancers who could be spared the toxicities of cytotoxic chemotherapies without compromising the prognosis.

Beyond chemotherapy, endocrine therapies also have risks, especially when given for extended durations. Recently, extended endocrine therapy has been shown to prevent late recurrences of HR-positive breast cancers. In the MA.17R study, extended endocrine therapy with letrozole for a total of 10 years (beyond 5 years of an aromatase inhibitor [AI]) decreased the risk for breast cancer recurrence or the occurrence of contralateral breast cancer by 34% [2]. However, the overall survival was similar between the 2 groups and the results were not confirmed in other studies [3–5]. Identifying the subgroup of patients who benefit from this extended AI therapy is important in the era of personalized medicine. Several tumor genomic assays have been developed to provide additional prognostic and predictive information with the goal of individualizing adjuvant therapies for breast cancer. Although assays are also being evaluated in HER2-positive and triple negative breast cancer, this review will focus on HR-positive, HER2-negative breast cancer.

Case Study

Initial Presentation

A 54-year-old postmenopausal woman with no significant past medical history presents with an abnormal screening mammogram, which shows a focal asymmetry in the 10 o’clock position at middle depth of the left breast. Further work-up with a diagnostic mammogram and ultrasound of the left breast shows a suspicious hypoechoic solid mass with irregular margins measuring 17 mm. The patient undergoes an ultrasound-guided core needle biopsy of the suspicious mass, the results of which are consistent with an invasive ductal carcinoma, Nottingham grade 2, ER strongly positive (95%), PR weakly positive (5%), HER2 negative, and Ki-67 of 15%. She undergoes a left partial mastectomy and sentinel lymph node biopsy, with final pathology demonstrating a single focus of invasive ductal carcinoma, measuring 2.2 cm in greatest dimension with no evidence of lymphovascular invasion. Margins are clear and 2 sentinel lymph nodes are negative for metastatic disease (final pathologic stage IIA, pT2 pN0 cM0). She is referred to medical oncology to discuss adjuvant systemic therapy.

• Can additional testing be used to determine prognosis and guide systemic therapy rec-ommendations for early-stage HR-positive/HER2-negative breast cancer?

After a diagnosis of early-stage breast cancer, the key clinical question faced by the patient and medical oncologist is: what is the individual’s risk for a metastatic breast cancer recurrence and thus the risk for death due to breast cancer? Once the risk for recurrence is established, systemic adjuvant chemotherapy, endocrine therapy, and/or HER2-directed therapy are considered based on the receptor status (ER/PR and HER2) to reduce this risk. Hormone receptor (HR)–positive, HER2-negative breast cancer is the most common type of breast cancer. Although adjuvant endocrine therapy has significantly reduced the risk for recurrence and improved survival for HR-positive breast cancer [6], the role of adjuvant chemotherapy for this subset of breast cancer remains unclear. Prior to genomic testing, the recommendation for adjuvant chemotherapy for HR-positive/HER2-negative tumors was primarily based on patient age and tumor stage and grade. However, chemotherapy overtreatment remained a concern given the potential short- and long-term risks of chemotherapy. Further studies into HR-positive/HER2-negative tumors have shown that these tumors can be divided into 2 main subtypes, luminal A and luminal B [7]. These subtypes represent unique biology and differ in terms of prognosis and response to endocrine therapy and chemotherapy. Luminal A tumors are strongly endocrine responsive and have a good prognosis, while luminal B tumors are less endocrine responsive and are associated with a poorer prognosis; the addition of adjuvant chemotherapy is often considered for luminal B tumors [8]. Several tests, including tumor genomic assays, are now available to help with delineating the tumor subtype and aid in decision-making regarding adjuvant chemotherapy for HR-positive/HER2-negative breast cancers.

Tests for Guiding Adjuvant Chemotherapy Decisions

Ki-67 Assays, Including IHC4 and PEPI

Chronic proliferation is a hallmark of cancer cells [9]. Ki-67, a nuclear nonhistone protein whose expression varies in intensity throughout the cell cycle, has been used as a measurement of tumor cell proliferation [10]. Two large meta-analyses have demonstrated that high Ki-67 expression in breast tumors is independently associated with worse disease-free and overall survival rates [11,12]. Ki-67 expression has also been used to classify HR-positive tumors as luminal A or B. After classifying tumor subtypes based on intrinsic gene expression profiling, Cheang et al determined that a Ki-67 cut point of 13.25% differentiated luminal A and B tumors [13]. However, the ideal cut point for Ki-67 remains unclear, as the sensitivity and specificity in this study was 77% and 78%, respectively. Others have combined Ki-67 with standard ER, PR, and HER2 testing. This IHC4 score, which weighs each of these variables, was validated in postmenopausal patients from the ATAC (Arimidex, Tamoxifen, Alone or in Combination) trial who had ER-positive tumors and did not receive chemotherapy [14]. The prognostic information from the IHC4 was similar to that seen with the 21-gene recurrence score (Oncotype DX), which is discussed later in this article. The key challenge with Ki-67 testing currently is the lack of a validated test methodology, and intraobserver variability in interpreting the Ki-67 results [15]. Recent series have suggested that Ki-67 be considered as a continuous marker rather than a set cut point [16]. These issues continue to impact the clinical utility of Ki-67 for decision making for adjuvant chemotherapy.

Ki-67 and the preoperative endocrine prognostic index (PEPI) score have been explored in the neoadjuvant setting to separate postmenopausal women with endocrine-sensitive versus intrinsically resistant disease and identify patients at risk for recurrent disease [17]. The on-treatment levels of Ki-67 in response to endocrine therapy have been shown to be more prognostic than baseline values, and a decrease in Ki-67 as early as 2 weeks after initiation of neoadjuvant endocrine therapy is associated with endocrine-sensitive tumors and improved outcome. The PEPI score was developed through retrospective analysis of the P024 trial [18] to evaluate the relationship between post-neoadjuvant endocrine therapy tumor characteristics and risk for early relapse. This was subsequently validated in an independent data set from the IMPACT trial [19]. Patients with low pathological stage (0 or 1) and a favorable biomarker profile (PEPI score 0) at surgery had the best prognosis in the absence of chemotherapy. On the other hand, higher pathological stage at surgery and a poor biomarker profile with loss of ER positivity or persistently elevated Ki-67 (PEPI score of 3) identified de novo endocrine-resistant tumors which are at higher risk for early relapse [20]. The ongoing Alliance A011106 ALTERNATE trial (ALTernate approaches for clinical stage II or III Estrogen Receptor positive breast cancer NeoAdjuvant TrEatment in postmenopausal women, NCT01953588) is a phase 3 study to prospectively test this hypothesis.

21-Gene Recurrence Score (Oncotype DX Assay)

The 21-gene Oncotype DX assay is conducted on paraffin-embedded tumor tissue and measures the expression of 16 cancer-related genes and 5 reference genes using quantitative polymerase chain reaction. The genes included in this assay are mainly related to proliferation (including Ki-67), invasion, and HER2 or estrogen signaling [21]. Originally, the 21-gene recurrence score assay was analyzed as a prognostic biomarker tool in a prospective-retrospective biomarker substudy of the National Surgical Adjuvant Breast and Bowel Project (NSABP) B-14 clinical trial in which patients with node-negative, ER-positive tumors were randomly assigned to receive tamoxifen or placebo without chemotherapy [22]. Using the standard reported values of low risk (< 18), intermediate risk (18–30), or high risk (≥ 31) for recurrence, among the tamoxifen-treated patients, cancers with a high-risk recurrence score had a significantly worse rate of distant recurrence and overall survival [21]. Inferior breast cancer survival with a high recurrence score was also confirmed in other series of endocrine-treated patients with node-negative and node-positive disease [23–25].

The predictive utility of the 21-gene recurrence score for endocrine therapy has also been evaluated. A comparison of the placebo- and tamoxifen-treated patients from the NSABP B-14 trial demonstrated that the 21-gene recurrence score predicted benefit from tamoxifen in cancers with low- or intermediate-risk recurrence scores [26]. However, there was no benefit from the use of tamoxifen over placebo in cancers with high-risk recurrence scores. To date, this intriguing data has not been prospectively confirmed, and thus the 21-gene recurrence score is not used to avoid endocrine therapy.

The 21-gene recurrence score is primarily used by oncologists to aid in decision-making regarding adjuvant chemotherapy in patients with node-negative and node-positive (with up to 3 positive lymph nodes), HR-positive/HER2-negative breast cancers. The predictive utility of the 21-gene recurrence score for adjuvant chemotherapy was initially tested using tumor samples from the NSABP B-20 study. This study initially compared adjuvant tamoxifen alone with tamoxifen plus chemotherapy in patients with node-negative, HR-positive tumors. The prospective-retrospective biomarker analysis showed that the patients with high-risk 21-gene recurrence scores benefited from the addition of chemotherapy, whereas those with low- or intermediate-risk did not have an improved freedom from distant recurrence with chemotherapy [27]. Similarly, an analysis from the prospective phase 3 Southwest Oncology Group (SWOG) 8814 trial comparing tamoxifen to tamoxifen with chemotherapy showed that for node-positive tumors, chemotherapy benefit was only seen in those with high 21-gene recurrence scores [24].

Prospective studies are now starting to report results regarding the predictive role of the 21-gene recurrence score. The TAILORx (Trial Assigning Individualized Options for Treatment) trial includes women with node-negative, HR-positive and HER2-negative tumors measuring 0.6 to 5 cm. All patients were treated with standard of care endocrine therapy for at least 5 years. Chemotherapy was determined based on the 21-gene recurrence score results on the primary tumor. The 21-gene recurrence score cutoffs were changed to low (0–10), intermediate (11–25), and high (≥ 26). Patients with scores of 26 or higher were treated with chemotherapy, and those with intermediate scores were randomly assigned to hemotherapy or no chemotherapy; results from this cohort are still pending. However, excellent breast cancer outcomes with endocrine therapy alone were reported from the 1626 (15.9% of total cohort) prospectively followed patients with low-recurrence score tumors. The 5-year invasive disease-free survival was 93.8%, with overall survival of 98% [28]. Given that 5 years is appropriate follow-up to see any chemotherapy benefit, this data supports the recommendation for no chemotherapy in this cohort of patients with very low 21-gene recurrence scores.

The RxPONDER (Rx for Positive Node, Endocrine Responsive Breast Cancer) trial is evaluating women with 1 to 3 node-positive, HR-positive, HER2-negative tumors. In this trial, patients with 21-gene recurrence scores of 0 to 25 were assigned to adjuvant chemotherapy or none. Those with scores of 26 or higher were assigned to chemotherapy. All patients received standard adjuvant endocrine therapy. This study has completed accrual and results are pending. Of note, TAILORx and RxPONDER did not investigate the potential lack of benefit of endocrine therapy in cancers with high recurrence scores. Furthermore, despite data suggesting that chemotherapy may not even benefit women with 4 or more nodes involved but who have a low recurrence score [24], due to the lack of prospective data in this cohort and the quite high risk for distant recurrence, chemotherapy continues to be the standard of care for these patients.

PAM50 (Breast Cancer Prognostic Gene Signature)

Using microarray and quantitative reverse transcriptase PCR (RT-PCR) on formalin-fixed paraffin-embedded (FFPE) tissues, the Breast Cancer Prognostic Gene Signature (PAM50) assay was initially developed to identify intrinsic breast cancer subtypes, including luminal A, luminal B, HER2-enriched, and basal-like [7,29]. Based on the prediction analysis of microarray (PAM) method, the assay measures the expression levels of 50 genes, provides a risk category (low, intermediate, and high), and generates a numerical risk of recurrence score (ROR). The intrinsic subtype and ROR have been shown to add significant prognostic value to the clinicopathological characteristics of tumors. Clinical validity of PAM50 was evaluated in postmenopausal women with HR-positive, early-stage breast cancer treated in the prospective ATAC and ABCSG-8 (Austrian Breast and Colorectal Cancer Study Group 8) trials [30,31]. In 1017 patients with ER-positive breast cancer treated with anastrozole or tamoxifen in the ATAC trial, ROR added significant prognostic information beyond the clinical treatment score (integrated prognostic information from nodal status, tumor size, histopathologic grade, age, and anastrozole or tamoxifen treatment) in all patients. Also, compared with the 21-gene recurrence score, ROR provided more prognostic information in ER-positive, node-negative disease and better differentiation of intermediate- and higher-risk groups. Fewer patients were categorized as intermediate risk by ROR and more as high risk, which could reduce the uncertainty in the estimate of clinical benefit from chemotherapy [30]. The clinical utility of PAM50 as a prognostic model was also validated in 1478 postmenopausal women with ER-positive early-stage breast cancer enrolled in the ABCSG-8 trial. In this study, ROR assigned 47% of patients with node-negative disease to the low-risk category. In this low-risk group, the 10-year metastasis risk was less than 3.5 %, indicating lack of benefit from additional chemotherapy [31]. A key limitation of the PAM50 is the lack of any prospective studies with this assay.

PAM50 has been designed to be carried out in any qualified pathology laboratory. Moreover, the ROR score provides additional prognostic information about risk of late recurrence, which will be discussed in the next section.

70-Gene Breast Cancer Recurrence Assay (MammaPrint)

MammaPrint is a 70-gene assay that was initially developed using an unsupervised, hierarchical clustering algorithm on whole-genome expression arrays with early-stage breast cancer. Among 295 consecutive patients who had MammaPrint testing, those classified with a good-prognosis tumor signature (n = 115) had an excellent 10-year survival rate (94.5%) compared to those with a poor-prognosis signature (54.5%), and the signature remained prognostic upon multivariate analysis [32]. Subsequently, a pooled analysis comparing outcomes by MammaPrint score in patients with node-negative or 1 to 3 node-positive breast cancers treated as per discretion of their medical team with either adjuvant chemotherapy plus endocrine therapy or endocrine therapy alone reported that only those patients with a high-risk score benefited from chemotherapy [33]. Recently, a prospective phase 3 study (MINDACT [Microarray In Node negative Disease may Avoid ChemoTherapy]) evaluating the utility of MammaPrint for adjuvant chemotherapy decision-making reported results [34]. In this study, 6693 women with early-stage breast cancer were assessed by clinical risk and genomic risk using MammaPrint. Those with low clinical and genomic risk did not receive chemotherapy, while those with high clinical and genomic risk all received chemotherapy. The primary goal of the study was to assess whether forgoing chemotherapy would be associated with a low rate of recurrence in those patients with a low-risk prognostic MammaPrint signature but high clinical risk. A total of 1550 patients (23.2%) were in the discordant group, and the majority of these patients had HR-positive disease (98.1%). Without chemotherapy, the rate of survival without distant metastasis at 5 years in this group was 94.7% (95% confidence interval [CI] 92.5% to 96.2%), which met the primary endpoint. Of note, initially, MammaPrint was only available for fresh tissue analysis, but recent advances in RNA processing now allow for this analysis on FFPE tissue [35].

Summary

Case Continued

The patient undergoes 21-gene recurrence score testing, which shows a low recurrence score of 10, estimating the 10-year risk of distant recurrence to be approximately 7% with 5 years of tamoxifen. Chemo-therapy is not recommended. The patient completes adjuvant whole breast radiation therapy, and then, based on data supporting AIs over tamoxifen in postmenopausal women, she is started on anastrozole [36]. She initially experiences mild side effects from treatment, including fatigue, arthralgia, and vaginal dryness, but her symptoms are able to be managed. As she approaches 5 years of adjuvant endocrine therapy with anastrozole, she is struggling with rotator cuff injury and is anxious about recurrence, but has no evidence of recurrent cancer. Her bone density scan in the beginning of her fourth year of therapy shows a decrease in bone mineral density, with the lowest T score of –1.5 at the left femoral neck, consistent with osteopenia. She has been treated with calcium and vitamin D supplements.

• How long should this patient continue treatment with anastrozole?

The risk for recurrence is highest during the first 5 years after diagnosis for all patients with early breast cancer [37]. Although HR-positive breast cancers have a better prognosis than HR-negative disease, the pattern of recurrence is different between the 2 groups, and it is estimated that approximately half of the recurrences among patients with HR-positive early breast cancer occur after the first 5 years from diagnosis. Annualized hazard of recurrence in HR-positive breast cancer has been shown to remain elevated and fairly stable beyond 10 years, even for those with low tumor burden and node-negative disease [38]. Prospective trials showed that for women with HR-positive early breast cancer, 5 years of adjuvant tamoxifen could substantially reduce recurrence rates and improve survival, and this became the standard of care [39]. AIs are considered the standard of care for adjuvant endocrine therapy in most postmenopausal women, as they result in a significantly lower recurrence rate compared with tamoxifen, either as initial adjuvant therapy or sequentially following 2 to 3 years of tamoxifen [40].

However, extending AI therapy from 5 years to 10 years is not clearly beneficial. In the MA.17R trial, although longer AI therapy resulted in significantly better disease-free survival (95% versus 91%, hazard ratio 0.66; P = 0.01), this was primarily due to a lower incidence of contralateral breast cancer in those taking the AI compared with placebo. The distant recurrence risks were similar and low (4.4% versus 5.5%), and there was no overall survival difference [2]. Also, the NSABP B-42 study, which was presented at the 2016 San Antonio Breast Cancer Symposium, did not meet its predefined endpoint for benefit from extending adjuvant AI therapy with letrozole beyond 5 years [3]. Thus, the absolute benefit from extended endocrine therapy has been modest across these studies. Although endocrine therapy is considered relatively safe and well tolerated, side effects can be significant and even associated with morbidity. Ideally, extended endocrine therapy should be offered to the subset of patients who would benefit the most. Several genomic diagnostic assays, including the EndoPredict test, PAM50, and the Breast Cancer Index (BCI) tests, specifically assess the risk for late recurrence in HR-positive cancers.

Tests for Assessing Risk for Late Recurrence

PAM50

Studies suggest that the ROR score also has value in predicting late recurrences. Analysis of data in patients enrolled in the ABCSG-8 trial showed that ROR could identify patients with endocrine-sensitive disease who are at low risk for late relapse and could be spared from unwanted toxicities of extended endocrine therapies. In 1246 ABCSG-8 patients between years 5 and 15, the PAM50 ROR demonstrated an absolute risk of distant recurrence of 2.4% in the low-risk group, as compared with 17.5% in the high-risk group [44]. Also, a combined analysis of patients from both the ATAC and ABCSG-8 trials demonstrated the utility of ROR in identifying this subgroup of patients with low risk for late relapse [45].

EndoPredict

EndoPredict (EP) is another quantitative RT-PCR–based assay which uses FFPE tissues to calculate a risk score based on 8 cancer-related and 3 reference genes. The score is combined with clinicopathological factors including tumor size and nodal status to make a comprehensive risk score (EPclin). EPclin is used to dichotomize patients into EP low- and EP high-risk groups. EP has been validated in 2 cohorts of patients enrolled in separate randomized studies, ABCSG-6 and ABCSG-8. EP provided prognostic information beyond clinicopathological variables to predict distant recurrence in patients with HR-positive, HER2-negative early breast cancer [46]. More important, EP has been shown to predict early (years 0–5) versus late (> 5 years after diagnosis) recurrences and identify a low-risk subset of patients who would not be expected to benefit from further treatment beyond 5 years of endocrine therapy [47]. Recently, EP and EPclin were compared with the 21-gene (Oncotype DX) recurrence score in a patient population from the TransATAC study. Both EP and EPclin provided more prognostic information compared to the 21-gene recurrence score and identified early and late relapse events [48]. EndoPredict is the first multigene expression assay that could be routinely performed in decentral molecular pathological laboratories with a short turnaround time [49].

Breast Cancer Index

The BCI is a RT-PCR–based gene expression assay that consists of 2 gene expression biomarkers: molecular grade index (MGI) and HOXB13/IL17BR (H/I). The BCI was developed as a prognostic test to assess risk for breast cancer recurrence using a cohort of ER-positive patients (n = 588) treated with adjuvant tamoxifen versus observation from the prospective randomized Stockholm trial [50]. In this blinded retrospective study, H/I and MGI were measured and a continuous risk model (BCI) was developed in the tamoxifen-treated group. More than 50% of the patients in this group were classified as having a low risk of recurrence. The rate of distant recurrence or death in this low-risk group at 10 years was less than 3%. The performance of the BCI model was then tested in the untreated arm of the Stockholm trial. In the untreated arm, BCI classified 53%, 27%, and 20% of patients as low, intermediate, and high risk, respectively. The rate of distant metastasis at 10 years in these risk groups was 8.3% (95% CI 4.7% to 14.4%), 22.9% (95% CI 14.5% to 35.2%), and 28.5% (95% CI 17.9% to 43.6%), respectively, and the rate of breast cancer–specific mortality was 5.1% (95% CI 1.3% to 8.7%), 19.8% (95% CI 10.0% to 28.6%), and 28.8% (95% CI 15.3% to 40.2%) [50].

The prognostic and predictive values of the BCI have been validated in other large, randomized studies and in patients with both node-negative and node-positive disease [51,52]. The predictive value of the endocrine-response biomarker, the H/I ratio, has been demonstrated in randomized studies. In the MA.17 trial, a high H/I ratio was associated with increased risk for late recurrence in the absence of letrozole. However, extended endocrine therapy with letrozole in patients with high H/I ratios predicted benefit from therapy and decreased the probability of late disease recurrence [53]. BCI was also compared to IHC4 and the 21-gene recurrence score in the TransATAC study and was the only test to show prognostic significance for both early (0–5 years) and late (5–10 year) recurrence [54].

The impact of the BCI results on physicians’ recommendations for extended endocrine therapy was assessed by a prospective study. This study showed that the test result had a significant effect on both physician treatment recommendation and patient satisfaction. BCI testing resulted in a change in physician recommendations for extended endocrine therapy, with an overall decrease in recommendations for extended endocrine therapy from 74% to 54%. Knowledge of the test result also led to improved patient satisfaction and decreased anxiety [55].

Summary

Due to the risk for late recurrence, extended endocrine therapy is being recommended for many patients with HR-positive breast cancers. Multiple genomic assays are being developed to better understand an individual’s risk for late recurrence and the potential for benefit from extended endocrine therapies. However, none of the assays have been validated in prospective randomized studies. Further validation is needed prior to routine use of these assays.

Case Continued

A BCI test is done and the result shows 4.3% BCI low-risk category in years 5–10; low likelihood of benefit from extended endocrine therapy. After discussing the results of the BCI test in the context of no survival benefit from extending AIs beyond 5 years, both the patient and her oncologist feel comfortable with discontinuing endocrine therapy at the end of 5 years.

Conclusion

Reduction in breast cancer mortality is mainly the result of improved systemic treatments. With advances in breast cancer screening tools in recent years, the rate of cancer detection has increased. This has raised concerns regarding overdiagnosis. To prevent unwanted toxicities associated with overtreatment, better treatment decision tools are needed. Several genomic assays are currently available and widely used to provide prognostic and predictive information and aid in decisions regarding appropriate use of adjuvant chemotherapy in HR-positive/HER2-negative early-stage breast cancer. Ongoing studies are refining the cutoffs for these assays and expanding the applicability to node-positive breast cancers. Furthermore, with several studies now showing benefit from the use of extended endocrine therapy, some of these assays may be able to identify the subset of patients who are at increased risk for late recurrence and who might benefit from extended endocrine therapy. Advances in molecular testing has enabled clinicians to offer more personalized treatments to their patients, improve patient’s compliance, and decrease anxiety and conflict associated with management decisions. Although small numbers of patients with HER2-positive and triple negative breast cancers were also included in some of these studies, use of genomic assays in this subset of patients is very limited and currently not recommended.

Corresponding author: Kari Braun Wisinski, MD, 1111 Highland Avenue, 6033 Wisconsin Institute for Medical Research, Madison, WI 53705-2275, kbwisinski@medicine.wisc.edu.

Financial disclosures: This work was supported by the NCI Cancer Center Support Grant P30 CA014520.

A 30-year-old man presented with multiple asymptomatic skin-colored and yellowish papules and nodules over the elbows, wrists, feet, buttocks, hands, and forearms (Figure 1) that had appeared suddenly 2 weeks before.

His medical history included diabetes mellitus, Hansen disease diagnosed 6 years earlier and treated with a multibacillary regimen for 1 year, and pancreatitis diagnosed 6 months earlier.

Laboratory testing. When a serum sample was centrifuged at 1,500 rpm for 15 minutes, a large lipid layer formed at the top (Figure 2). Other results:

• Triglycerides 5,742 mg/dL (reference range < 160)
• Total cholesterol 293 mg/dL (< 240 mg/dL)
• Fasting blood glucose 473 mg/dL
• Low-density and high-density lipoprotein cholesterol levels normal
• Complete blood cell count within normal limits.

Other testing. The electrocardiogram was normal. Retinal examination showed evidence of lipemia retinalis. Histologic study of a biopsy specimen from one of the lesions showed foamy macrophages in the superficial dermis with lymphocytic infiltrate, confirming the diagnosis of eruptive xanthoma.

An urgent medical referral was sought. The patient was started on statins and fibrates, and the treatment resulted in remarkable improvement.

ERUPTIVE XANTHOMA

Cutaneous manifestations can be warning signs of systemic disease, and physicians should be aware of the dermatologic presentations of common medical conditions.

Eruptive xanthoma is a sign of severe hypertriglyceridemia and is almost always due to an acquired cause such as hyperchylomicro­nemia or an inherited lipoprotein disorder (eg, lipoprotein lipase deficiency in children, common familial hypertriglyceridemia type 5 in adults).¹

Chylomicronemia syndrome is characterized by triglyceride levels greater than 1,000 mg/dL in a patient with eruptive xanthoma, lipemia retinalis, or abdominal pain or pancreatitis. The syndrome has a prevalence of 1.7 out of 10,000 patients.² Treatment is a strict low-fat diet, with a minimal role for fibrates and nicotinic acid.

Eruptive xanthoma is seen at the time of presentation in 8.5% of patients with severe hypertriglyceridemia (serum level > 1,772 mg/dL).³ In patients with serum triglyceride levels greater than 1,000 mg/dL, the risk of acute pancreatitis is 5%, and at levels above 2,000 mg/dL, the risk is 10% to 20%.⁴

In our patient, the diagnosis of chylomicronemia was based on the appearance of the skin lesions, his history of pancreatitis and diabetes mellitus, and the results of the initial workup, which revealed severe hypertriglyceridemia and lipemia retinalis. The presence of eruptive xanthomas should raise suspicion for a grossly elevated triglyceride level. Therefore, in view of the increased risk of acute pancreatitis and pancreatic necrosis associated with chylomicronemic syndrome,² recognizing the cutaneous signs is mandatory.

A 30-year-old man presented with multiple asymptomatic skin-colored and yellowish papules and nodules over the elbows, wrists, feet, buttocks, hands, and forearms (Figure 1) that had appeared suddenly 2 weeks before.

His medical history included diabetes mellitus, Hansen disease diagnosed 6 years earlier and treated with a multibacillary regimen for 1 year, and pancreatitis diagnosed 6 months earlier.

Laboratory testing. When a serum sample was centrifuged at 1,500 rpm for 15 minutes, a large lipid layer formed at the top (Figure 2). Other results:

• Triglycerides 5,742 mg/dL (reference range < 160)
• Total cholesterol 293 mg/dL (< 240 mg/dL)
• Fasting blood glucose 473 mg/dL
• Low-density and high-density lipoprotein cholesterol levels normal
• Complete blood cell count within normal limits.

Other testing. The electrocardiogram was normal. Retinal examination showed evidence of lipemia retinalis. Histologic study of a biopsy specimen from one of the lesions showed foamy macrophages in the superficial dermis with lymphocytic infiltrate, confirming the diagnosis of eruptive xanthoma.

An urgent medical referral was sought. The patient was started on statins and fibrates, and the treatment resulted in remarkable improvement.

ERUPTIVE XANTHOMA

Cutaneous manifestations can be warning signs of systemic disease, and physicians should be aware of the dermatologic presentations of common medical conditions.

Eruptive xanthoma is a sign of severe hypertriglyceridemia and is almost always due to an acquired cause such as hyperchylomicro­nemia or an inherited lipoprotein disorder (eg, lipoprotein lipase deficiency in children, common familial hypertriglyceridemia type 5 in adults).¹

Chylomicronemia syndrome is characterized by triglyceride levels greater than 1,000 mg/dL in a patient with eruptive xanthoma, lipemia retinalis, or abdominal pain or pancreatitis. The syndrome has a prevalence of 1.7 out of 10,000 patients.² Treatment is a strict low-fat diet, with a minimal role for fibrates and nicotinic acid.

Eruptive xanthoma is seen at the time of presentation in 8.5% of patients with severe hypertriglyceridemia (serum level > 1,772 mg/dL).³ In patients with serum triglyceride levels greater than 1,000 mg/dL, the risk of acute pancreatitis is 5%, and at levels above 2,000 mg/dL, the risk is 10% to 20%.⁴

In our patient, the diagnosis of chylomicronemia was based on the appearance of the skin lesions, his history of pancreatitis and diabetes mellitus, and the results of the initial workup, which revealed severe hypertriglyceridemia and lipemia retinalis. The presence of eruptive xanthomas should raise suspicion for a grossly elevated triglyceride level. Therefore, in view of the increased risk of acute pancreatitis and pancreatic necrosis associated with chylomicronemic syndrome,² recognizing the cutaneous signs is mandatory.

A 30-year-old man presented with multiple asymptomatic skin-colored and yellowish papules and nodules over the elbows, wrists, feet, buttocks, hands, and forearms (Figure 1) that had appeared suddenly 2 weeks before.

His medical history included diabetes mellitus, Hansen disease diagnosed 6 years earlier and treated with a multibacillary regimen for 1 year, and pancreatitis diagnosed 6 months earlier.

Laboratory testing. When a serum sample was centrifuged at 1,500 rpm for 15 minutes, a large lipid layer formed at the top (Figure 2). Other results:

• Triglycerides 5,742 mg/dL (reference range < 160)
• Total cholesterol 293 mg/dL (< 240 mg/dL)
• Fasting blood glucose 473 mg/dL
• Low-density and high-density lipoprotein cholesterol levels normal
• Complete blood cell count within normal limits.

Other testing. The electrocardiogram was normal. Retinal examination showed evidence of lipemia retinalis. Histologic study of a biopsy specimen from one of the lesions showed foamy macrophages in the superficial dermis with lymphocytic infiltrate, confirming the diagnosis of eruptive xanthoma.

An urgent medical referral was sought. The patient was started on statins and fibrates, and the treatment resulted in remarkable improvement.

ERUPTIVE XANTHOMA

Cutaneous manifestations can be warning signs of systemic disease, and physicians should be aware of the dermatologic presentations of common medical conditions.

Eruptive xanthoma is a sign of severe hypertriglyceridemia and is almost always due to an acquired cause such as hyperchylomicro­nemia or an inherited lipoprotein disorder (eg, lipoprotein lipase deficiency in children, common familial hypertriglyceridemia type 5 in adults).¹

Chylomicronemia syndrome is characterized by triglyceride levels greater than 1,000 mg/dL in a patient with eruptive xanthoma, lipemia retinalis, or abdominal pain or pancreatitis. The syndrome has a prevalence of 1.7 out of 10,000 patients.² Treatment is a strict low-fat diet, with a minimal role for fibrates and nicotinic acid.

Eruptive xanthoma is seen at the time of presentation in 8.5% of patients with severe hypertriglyceridemia (serum level > 1,772 mg/dL).³ In patients with serum triglyceride levels greater than 1,000 mg/dL, the risk of acute pancreatitis is 5%, and at levels above 2,000 mg/dL, the risk is 10% to 20%.⁴

In our patient, the diagnosis of chylomicronemia was based on the appearance of the skin lesions, his history of pancreatitis and diabetes mellitus, and the results of the initial workup, which revealed severe hypertriglyceridemia and lipemia retinalis. The presence of eruptive xanthomas should raise suspicion for a grossly elevated triglyceride level. Therefore, in view of the increased risk of acute pancreatitis and pancreatic necrosis associated with chylomicronemic syndrome,² recognizing the cutaneous signs is mandatory.

Alpha-1 antitrypsin deficiency is a common but underrecognized genetic condition that increases the risk of chronic obstructive pulmonary disease (COPD) and liver disease. Primary care providers can play a critical role in detecting it and managing patients who have it.

RECOGNIZED CASES ARE THE TIP OF THE ICEBERG

First described in 1963,¹ alpha-1 antitrypsin deficiency is estimated to affect 100,000 Americans, fewer than 15,000 of whom have received a clinical diagnosis. As further evidence of its underrecognition,^2–7 many patients experience long delays between their first symptoms and the diagnosis. Early studies indicated that the average diagnostic delay was 7.2 years,4 and the latest studies, as recent as 2013, indicate a similar diagnostic delay.⁷

Furthermore, many patients see multiple healthcare providers before receiving the correct diagnosis. A 1994 survey by this author⁴ found that 43.7% of patients who had severe deficiency of alpha-1 antitrypsin saw at least three physicians before the correct diagnosis was made.

Why is the disease underrecognized?

Several reasons may account for underrecognition of this disease. Many clinicians—including, unfortunately, many pulmonologists—do not know much about it,^7,8 do not adhere to clinical guidelines,^9,10 or harbor the misperception that there is no therapy available and, therefore, no compelling reason to make a diagnosis.⁷

Regarding inadequate knowledge, in a study by Taliercio, Chatburn, and this author,⁸ internal medicine residents scored only 63% correct on a 10-question quiz on diagnostic features of alpha-1 antitrypsin deficiency. There was no evidence of a training effect—senior residents scored no higher than interns.

Similarly, when Greulich et al⁷ surveyed German and Italian internists, general practitioners, and pulmonologists, one-fourth to one-half of them (depending on specialty and country) stated that they knew either very little or nothing at all about alpha-1 antitrypsin deficiency. In addition, 7% to 8% agreed with the statement, “There is no treatment available for this disease.”⁷

Nonadoption of clinical guidelines has been widely recognized in medicine and is evident in the failure to implement various recommended practices,^9,10 such as low-stretch ventilation for acute respiratory distress syndrome and prophylaxis against deep vein thrombosis.

Finding the rest of the iceberg

Efforts to enhance compliance with guidelines on testing for alpha-1 antitrypsin deficiency have included using the electronic medical record to prompt physicians to test appropriate candidates.^11–13

Jain et al¹³ examined the effect of installing such a prompting system to remind physicians to test for alpha-1 antitrypsin deficiency in patients with airflow obstruction that does not reverse with a bronchodilator—a recognized indication for testing for this disease according to standards endorsed by the American Thoracic Society and European Respiratory Society.¹⁴ At baseline, only 4.7% of appropriate candidates were being tested; after a prompt was installed in the electronic medical record, the rate rose to 15.1%, still a minority of candidates.

Another strategy is to empower respiratory therapists who perform pulmonary function tests to invite patients to be tested if their pulmonary function tests show postbronchodilator airflow obstruction. Rahaghi et al¹⁵ showed that using this strategy, 20 (0.63%) of 3,152 patients who were found to have fixed airflow obstruction when they underwent pulmonary function testing were newly diagnosed with severe deficiency of alpha-1 antitrypsin. Other targeted detection studies in patients with COPD estimated the prevalence of alpha-1 antitrypsin deficiency at up to 12%.³

PHYSIOLOGY AND PATHOPHYSIOLOGY OF ALPHA-1 ANTITRYPSIN DEFICIENCY

Alpha-1 antitrypsin is a single-chain, 394-amino acid glycoprotein with three carbohydrate side chains found at asparagine residues along the primary structure.¹⁶

A major physiologic function of this molecule is to bind neutrophil elastase, which it does avidly. In a “mousetrap-like” mechanism,¹⁶ an active site on the alpha-1 antitrypsin molecule captures the neutrophil elastase and is cleaved, releasing steric energy in the molecule, catapulting the neutrophil elastase to the opposite side of the alpha-1 antitrypsin molecule, and inactivating it (Figure 1).

MM is normal, ZZ is not

Alpha-1 antitrypsin deficiency is inherited as an autosomal-codominant condition.¹⁷

The SERPINA1 gene, which codes for alpha-1 antitrypsin, is located on the long arm of the 14th chromosome, and more than 150 alleles of this gene have been identified to date. The normal allele is denoted M, and the allele most commonly associated with severe deficiency is denoted Z. People who are homozygous for the M allele (ie, normal) are called PI*MM (PI stands for “protease inhibitor”), and those who are homozygous for the Z allele are PI*ZZ. More than 90% of patients with severe alpha-1 antitrypsin deficiency are PI*ZZ.¹⁸

The Z allele has a single amino acid substitution (glutamic acid-to-lysine at position 342), which results in abnormal folding and formation of polymers of the Z molecule within hepatocytes.^19,20 These polymers are recognized on liver biopsy as periodic acid-Schiff diastase-resistant eosinophilic inclusion bodies on histologic staining (Figure 2).

With alpha-1 antitrypsin trapped as Z-molecule polymers in the liver, the amount in the bloodstream falls, and there is a consequent decrease in the amount available in the lung to oppose the proteolytic burden of neutrophil elastase, especially in people who smoke or work in dusty environments.²¹

Tan et al²² have shown that some of the polymerized Z protein can escape the liver and circulate in the blood and that alveolar macrophages may also produce Z polymers. These Z polymers are chemotactic for neutrophils,²³ so that their presence in the lung fuels the inflammatory cascade by recruiting more neutrophils to the lung, thereby increasing the proteolytic burden to the lung and increasing the risk of emphysema. Z monomers that do circulate can bind neutrophil elastase, but their binding avidity to neutrophil elastase is substantially lower than that of M-type alpha-1 antitrypsin.

CLINICAL MANIFESTATIONS

Alpha-1 antitrypsin deficiency of the PI*ZZ type is associated with two major clinical manifestations:

• Emphysema, resulting from the loss of proteolytic protection of the lung by alpha-1 antitrypsin (a toxic loss of function), and
• Liver diseases such as cirrhosis and chronic hepatitis, which result from abnormal accumulation of alpha-1 antitrypsin within hepatocytes (a toxic gain of function), and hepatoma.¹⁷

Other clinical manifestations of PI*ZZ alpha-1 antitrypsin deficiency include panniculitis and an association with cytoplasmic antineutrophil cytoplasmic antibody-positive vasculitis.¹⁷

Some uncertainty exists regarding the risk associated with the PI*MZ heterozygous state because there has been no systematic longitudinal study of people with this genotype. However, the weight of available experience suggests that PI*MZ individuals who have never smoked are not at increased risk of developing emphysema.²⁴

Findings from a national registry: PI*ZZ COPD resembles ‘usual’ COPD

Distinguishing patients with alpha-1 antitrypsin deficiency from those with “usual” COPD (ie, without alpha-1 antitrypsin deficiency) can be difficult, as shown in data from the National Heart, Lung, and Blood Institute’s   Alpha-1 Antitrypsin Deficiency Registry study.¹⁸ This multicenter, longitudinal, observational study contains the largest well-characterized cohort with severe deficiency of alpha-1 antitrypsin (PI*ZZ, PI*ZNull, etc), with 1,129 patients. 

Pulmonary function test results were consistent with emphysema in most of the patients in the registry. Mean postbronchodilator pulmonary function values (± standard error of the mean) were:

• Forced expiratory volume in 1 second (FEV₁) 46.7% of predicted (± 30%)
• Ratio of FEV₁ to forced vital capacity 42.9% (± 20.4% )
• Mean diffusing capacity for carbon monoxide 50.3% of predicted (± 22.5%).

Like many patients with usual COPD, 60% of the registry patients demonstrated a component of airway reactivity, with significant reversal of airflow obstruction over three spirometries after receiving a dose of an inhaled bronchodilator (characterized by a 12% and 200-mL postbronchodilator rise in FEV₁). Moreover, 78 patients had normal lung function.

Symptoms also resembled those in patients with usual emphysema, chronic bronchitis, or both. On enrollment in the registry, 83.9% of the patients had shortness of breath on exertion, 75.5% had wheezing with upper respiratory infections, 65.3% had wheezing without upper respiratory infection, 67.6% had recent debilitating chest illness, 42.4% had “usual” cough, and 49.6% had annual cough and phlegm episodes.

Imaging findings. Although the classic teaching is that emphysema due to alpha-1 antitrypsin deficiency produces lower-lobe hyperlucency on plain films, relying on this sign would lead to underrecognition, as 36% of PI*ZZ patients have apical-predominant emphysema on chest computed tomography,²⁴ which resembles the usual centriacinar emphysema pattern. Figure 3 shows axial computed tomographic scans through the apices and the bases of the lungs of a patient with alpha-1 antitrypsin deficiency.  

In view of these difficulties, guidelines from the American Thoracic Society and European Respiratory Society¹⁴ endorse testing for alpha-1 antitrypsin deficiency in all adults who have symptoms and fixed airflow obstruction (Table 1).

CONSEQUENCES OF ALPHA-1 ANTITRYPSIN DEFICIENCY

Two large screening studies^2,3,25,26 followed people who were identified at birth as having alpha-1 antitrypsin deficiency to examine the natural course of the disease.

The larger of the two studies²⁷ tested 200,000 Swedish newborns. Follow-up of this cohort to age 35 indicated that 35-year-old never-smoking PI*ZZ individuals have normal lung function and no excess emphysema on computed tomography compared with normal peers matched for age and sex.²⁷ In contrast, the few PI*ZZ ever-smokers demonstrated a lower level of transfer factor and significantly more emphysema on computed tomography than normal (PI*MM) never-smokers.

Faster decline in lung function

Data from the National Heart, Lung, and Blood Institute registry indicate that, on average, people with severe alpha-1 antitrypsin deficiency lose lung function faster than people without the disease.²⁸ Specifically, in never-smokers in the registry, the average rate of FEV₁ decline was 67 mL/year, and among ex-smokers, it was 54 mL/year. Both of these values exceed the general age-related rate of FEV₁ decline of approximately 20 to 25 mL/year in never-smoking, normal adults. Among current smokers in the registry with severe alpha-1 antitrypsin deficiency, the rate of FEV₁ decline was 109 mL/year.

Rates of FEV₁ decline over time vary among groups with differing degrees of airflow obstruction. For example, PI*ZZ patients with moderate COPD (stage II of the four-stage Global Initiative for Chronic Obstructive Lung Disease classification system) lose lung function faster than patients with either milder or more severe degrees of airflow obstruction.²⁹

As with COPD in general, exacerbations of COPD in people with severe deficiency of alpha-1 antitrypsin are associated with worsened clinical status. In one series,³⁰ 54% of 265 PI*ZZ patients experienced an exacerbation in the first year of follow-up, and 18% experienced at least three. Such exacerbations occurred in December and January in 32% of these individuals, likely due to a viral precipitant.

Increased mortality

Severe deficiency of alpha-1 antitrypsin is associated not only with severe morbidity but also death. In the national registry, the overall rate of death was 18.6% at 5 years of follow-up, or approximately 3% per year.²⁸

A low FEV₁ at entry was a bad sign. Patients entering the registry with FEV₁ values below 15% of predicted had a 36% mortality rate at 3 years, compared with 2.6% in those whose baseline FEV₁ exceeded 50% of predicted.

Underlying causes of death in registry participants included emphysema (accounting for 72% of deaths) and cirrhosis (10%),³¹ which were the only causes of death more frequent than in age- and sex-matched controls. In a series of never-smokers who had PI*ZZ alpha-1 antitrypsin deficiency,³² death was less frequently attributed to emphysema than in the national registry (46%) and more often attributed to cirrhosis (28%), indicating that never-smokers may more frequently escape the ravages of emphysema but experience a higher rate of developing cirrhosis later in life.³³

DIAGNOSING ALPHA-1 ANTITRYPSIN DEFICIENCY

Available blood tests for alpha-1 antitrypsin deficiency include:

The serum alpha-1 antitrypsin level, most often done by nephelometry. Normal serum levels generally range from 100 to 220 mg/dL.

Phenotyping, usually performed by isoelectric focusing, which can identify different band patterns associated with different alleles.

Genotyping involves determining which alpha-1 antitrypsin alleles are present, most often using polymerase chain reaction testing targeting the S and Z alleles and occasionally set up to detect less common alleles such as F and I.¹⁷

Gene sequencing is occasionally necessary to achieve an accurate, definitive  diagnosis.

Free, confidential testing is available

Clinical testing most often involves checking both a serum level and a phenotype or genotype. Such tests are often available in hospital laboratories and commercial laboratories, with testing also facilitated by the availability of free testing kits from several manufacturers of drugs for alpha-1 antitrypsin deficiency.

The Alpha-1 Foundation (www.alpha1.org)³⁴ also offers a free, home-based confidential testing kit through a research protocol at the Medical University of South Carolina (alphaone@musc.edu) called the Alpha-1 Coded Testing (ACT) study. Patients can receive a kit and lancet at home, submit the dried blood-spot specimen, and receive in the mail a confidential serum level and genotype.

The availability of such home-based confidential testing allows patients to seek testing without a physician’s order and makes it easier for facilitated allied health providers, such as respiratory therapists, to recommend testing in appropriate clinical circumstances.¹⁵

TREATMENT OF ALPHA-1 ANTITRYPSIN DEFICIENCY

The treatment of patients with severe deficiency of alpha-1 antitrypsin and emphysema generally resembles that of patients with usual COPD. Specifically, smoking cessation, bronchodilators, occasionally inhaled steroids, supplemental oxygen, preventive vaccinations, and pulmonary rehabilitation are indicated as per usual clinical assessment.

Lung volume reduction surgery, which is beneficial in appropriate subsets of COPD patients, is generally less effective in those with severe alpha-1 antitrypsin deficiency,³⁵ specifically because the magnitude of FEV₁ increase and the duration of such a rise are lower than in usual COPD patients.

Augmentation therapy

Specific therapy for alpha-1 antitrypsin deficiency currently involves weekly intravenous infusions of purified, pooled human-plasma-derived alpha-1 antitrypsin, so-called augmentation therapy. Four drugs have been approved for use in the United States:

• Prolastin-C (Grifols, Barcelona, Spain)
• Aralast NP (Baxalta, Bonneckborn, IL)
• Zemaira (CSL Behring, King of Prussia, PA)
• Glassia (Baxalta, Bonneckborn, IL, and Kamada, Ness Ziona, Israel).

All of these were approved for use in the United States on the basis of biochemical efficacy. Specifically, infusion of these drugs has been shown to raise serum levels above a protective threshold value (generally considered 57 mg/dL, the value below which the risk of developing emphysema increases beyond normal).

Randomized controlled trials^36,37 have addressed the efficacy of intravenous augmentation therapy, and although no single trial has been definitive, the weight of evidence shows that augmentation therapy can slow the progression of emphysema. For example, in a study by Dirksen et al,³⁷ augmentation therapy was associated with a slower progression of emphysema as assessed by the rate of loss of lung density on computed tomography.

On the basis of the available evidence, the American Thoracic Society and European Respiratory Society¹⁴ have recommended augmentation therapy in individuals with “established airflow obstruction from alpha-1 antitrypsin deficiency.”¹⁴ Their guidelines go on to say that the evidence that augmentation therapy is beneficial “is stronger for individuals with moderate airflow obstruction (eg, FEV₁ 35%–60% of predicted) than for those with severe airflow obstruction. Augmentation therapy is not currently recommended for individuals without emphysema.”

The guidelines recognize that although augmentation therapy does not satisfy the usual criteria for cost-effectiveness (< $50,000 per quality-adjusted life year) due to its high cost (approximately $100,000 per year if paid for out of pocket),³⁸ it is recommended for appropriate candidates because it is the only available specific therapy for severe deficiency of alpha-1 antitrypsin.

Novel therapies

In addition to current treatment approaches of augmentation therapy, a number of novel treatment strategies are being investigated, several of which hold much promise.

Gene therapy, using adeno-associated virus to transfect the normal human gene into individuals with severe deficiency of alpha-1 antitrypsin, has been undertaken and is currently under study. In addition, a variety of approaches to interdict production of abnormal Z protein from the liver are being examined, as well as inhaled hyaluronic acid to protect the lung.

Alpha-1 antitrypsin deficiency is a common but underrecognized genetic condition that increases the risk of chronic obstructive pulmonary disease (COPD) and liver disease. Primary care providers can play a critical role in detecting it and managing patients who have it.

RECOGNIZED CASES ARE THE TIP OF THE ICEBERG

First described in 1963,¹ alpha-1 antitrypsin deficiency is estimated to affect 100,000 Americans, fewer than 15,000 of whom have received a clinical diagnosis. As further evidence of its underrecognition,^2–7 many patients experience long delays between their first symptoms and the diagnosis. Early studies indicated that the average diagnostic delay was 7.2 years,4 and the latest studies, as recent as 2013, indicate a similar diagnostic delay.⁷

Furthermore, many patients see multiple healthcare providers before receiving the correct diagnosis. A 1994 survey by this author⁴ found that 43.7% of patients who had severe deficiency of alpha-1 antitrypsin saw at least three physicians before the correct diagnosis was made.

Why is the disease underrecognized?

Several reasons may account for underrecognition of this disease. Many clinicians—including, unfortunately, many pulmonologists—do not know much about it,^7,8 do not adhere to clinical guidelines,^9,10 or harbor the misperception that there is no therapy available and, therefore, no compelling reason to make a diagnosis.⁷

Regarding inadequate knowledge, in a study by Taliercio, Chatburn, and this author,⁸ internal medicine residents scored only 63% correct on a 10-question quiz on diagnostic features of alpha-1 antitrypsin deficiency. There was no evidence of a training effect—senior residents scored no higher than interns.

Similarly, when Greulich et al⁷ surveyed German and Italian internists, general practitioners, and pulmonologists, one-fourth to one-half of them (depending on specialty and country) stated that they knew either very little or nothing at all about alpha-1 antitrypsin deficiency. In addition, 7% to 8% agreed with the statement, “There is no treatment available for this disease.”⁷

Nonadoption of clinical guidelines has been widely recognized in medicine and is evident in the failure to implement various recommended practices,^9,10 such as low-stretch ventilation for acute respiratory distress syndrome and prophylaxis against deep vein thrombosis.

Finding the rest of the iceberg

Efforts to enhance compliance with guidelines on testing for alpha-1 antitrypsin deficiency have included using the electronic medical record to prompt physicians to test appropriate candidates.^11–13

Jain et al¹³ examined the effect of installing such a prompting system to remind physicians to test for alpha-1 antitrypsin deficiency in patients with airflow obstruction that does not reverse with a bronchodilator—a recognized indication for testing for this disease according to standards endorsed by the American Thoracic Society and European Respiratory Society.¹⁴ At baseline, only 4.7% of appropriate candidates were being tested; after a prompt was installed in the electronic medical record, the rate rose to 15.1%, still a minority of candidates.

Another strategy is to empower respiratory therapists who perform pulmonary function tests to invite patients to be tested if their pulmonary function tests show postbronchodilator airflow obstruction. Rahaghi et al¹⁵ showed that using this strategy, 20 (0.63%) of 3,152 patients who were found to have fixed airflow obstruction when they underwent pulmonary function testing were newly diagnosed with severe deficiency of alpha-1 antitrypsin. Other targeted detection studies in patients with COPD estimated the prevalence of alpha-1 antitrypsin deficiency at up to 12%.³

PHYSIOLOGY AND PATHOPHYSIOLOGY OF ALPHA-1 ANTITRYPSIN DEFICIENCY

Alpha-1 antitrypsin is a single-chain, 394-amino acid glycoprotein with three carbohydrate side chains found at asparagine residues along the primary structure.¹⁶

A major physiologic function of this molecule is to bind neutrophil elastase, which it does avidly. In a “mousetrap-like” mechanism,¹⁶ an active site on the alpha-1 antitrypsin molecule captures the neutrophil elastase and is cleaved, releasing steric energy in the molecule, catapulting the neutrophil elastase to the opposite side of the alpha-1 antitrypsin molecule, and inactivating it (Figure 1).

MM is normal, ZZ is not

Alpha-1 antitrypsin deficiency is inherited as an autosomal-codominant condition.¹⁷

The SERPINA1 gene, which codes for alpha-1 antitrypsin, is located on the long arm of the 14th chromosome, and more than 150 alleles of this gene have been identified to date. The normal allele is denoted M, and the allele most commonly associated with severe deficiency is denoted Z. People who are homozygous for the M allele (ie, normal) are called PI*MM (PI stands for “protease inhibitor”), and those who are homozygous for the Z allele are PI*ZZ. More than 90% of patients with severe alpha-1 antitrypsin deficiency are PI*ZZ.¹⁸

The Z allele has a single amino acid substitution (glutamic acid-to-lysine at position 342), which results in abnormal folding and formation of polymers of the Z molecule within hepatocytes.^19,20 These polymers are recognized on liver biopsy as periodic acid-Schiff diastase-resistant eosinophilic inclusion bodies on histologic staining (Figure 2).

With alpha-1 antitrypsin trapped as Z-molecule polymers in the liver, the amount in the bloodstream falls, and there is a consequent decrease in the amount available in the lung to oppose the proteolytic burden of neutrophil elastase, especially in people who smoke or work in dusty environments.²¹

Tan et al²² have shown that some of the polymerized Z protein can escape the liver and circulate in the blood and that alveolar macrophages may also produce Z polymers. These Z polymers are chemotactic for neutrophils,²³ so that their presence in the lung fuels the inflammatory cascade by recruiting more neutrophils to the lung, thereby increasing the proteolytic burden to the lung and increasing the risk of emphysema. Z monomers that do circulate can bind neutrophil elastase, but their binding avidity to neutrophil elastase is substantially lower than that of M-type alpha-1 antitrypsin.

CLINICAL MANIFESTATIONS

Alpha-1 antitrypsin deficiency of the PI*ZZ type is associated with two major clinical manifestations:

• Emphysema, resulting from the loss of proteolytic protection of the lung by alpha-1 antitrypsin (a toxic loss of function), and
• Liver diseases such as cirrhosis and chronic hepatitis, which result from abnormal accumulation of alpha-1 antitrypsin within hepatocytes (a toxic gain of function), and hepatoma.¹⁷

Other clinical manifestations of PI*ZZ alpha-1 antitrypsin deficiency include panniculitis and an association with cytoplasmic antineutrophil cytoplasmic antibody-positive vasculitis.¹⁷

Some uncertainty exists regarding the risk associated with the PI*MZ heterozygous state because there has been no systematic longitudinal study of people with this genotype. However, the weight of available experience suggests that PI*MZ individuals who have never smoked are not at increased risk of developing emphysema.²⁴

Findings from a national registry: PI*ZZ COPD resembles ‘usual’ COPD

Distinguishing patients with alpha-1 antitrypsin deficiency from those with “usual” COPD (ie, without alpha-1 antitrypsin deficiency) can be difficult, as shown in data from the National Heart, Lung, and Blood Institute’s   Alpha-1 Antitrypsin Deficiency Registry study.¹⁸ This multicenter, longitudinal, observational study contains the largest well-characterized cohort with severe deficiency of alpha-1 antitrypsin (PI*ZZ, PI*ZNull, etc), with 1,129 patients. 

Pulmonary function test results were consistent with emphysema in most of the patients in the registry. Mean postbronchodilator pulmonary function values (± standard error of the mean) were:

• Forced expiratory volume in 1 second (FEV₁) 46.7% of predicted (± 30%)
• Ratio of FEV₁ to forced vital capacity 42.9% (± 20.4% )
• Mean diffusing capacity for carbon monoxide 50.3% of predicted (± 22.5%).

Like many patients with usual COPD, 60% of the registry patients demonstrated a component of airway reactivity, with significant reversal of airflow obstruction over three spirometries after receiving a dose of an inhaled bronchodilator (characterized by a 12% and 200-mL postbronchodilator rise in FEV₁). Moreover, 78 patients had normal lung function.

Symptoms also resembled those in patients with usual emphysema, chronic bronchitis, or both. On enrollment in the registry, 83.9% of the patients had shortness of breath on exertion, 75.5% had wheezing with upper respiratory infections, 65.3% had wheezing without upper respiratory infection, 67.6% had recent debilitating chest illness, 42.4% had “usual” cough, and 49.6% had annual cough and phlegm episodes.

Imaging findings. Although the classic teaching is that emphysema due to alpha-1 antitrypsin deficiency produces lower-lobe hyperlucency on plain films, relying on this sign would lead to underrecognition, as 36% of PI*ZZ patients have apical-predominant emphysema on chest computed tomography,²⁴ which resembles the usual centriacinar emphysema pattern. Figure 3 shows axial computed tomographic scans through the apices and the bases of the lungs of a patient with alpha-1 antitrypsin deficiency.  

In view of these difficulties, guidelines from the American Thoracic Society and European Respiratory Society¹⁴ endorse testing for alpha-1 antitrypsin deficiency in all adults who have symptoms and fixed airflow obstruction (Table 1).

CONSEQUENCES OF ALPHA-1 ANTITRYPSIN DEFICIENCY

Two large screening studies^2,3,25,26 followed people who were identified at birth as having alpha-1 antitrypsin deficiency to examine the natural course of the disease.

The larger of the two studies²⁷ tested 200,000 Swedish newborns. Follow-up of this cohort to age 35 indicated that 35-year-old never-smoking PI*ZZ individuals have normal lung function and no excess emphysema on computed tomography compared with normal peers matched for age and sex.²⁷ In contrast, the few PI*ZZ ever-smokers demonstrated a lower level of transfer factor and significantly more emphysema on computed tomography than normal (PI*MM) never-smokers.

Faster decline in lung function

Data from the National Heart, Lung, and Blood Institute registry indicate that, on average, people with severe alpha-1 antitrypsin deficiency lose lung function faster than people without the disease.²⁸ Specifically, in never-smokers in the registry, the average rate of FEV₁ decline was 67 mL/year, and among ex-smokers, it was 54 mL/year. Both of these values exceed the general age-related rate of FEV₁ decline of approximately 20 to 25 mL/year in never-smoking, normal adults. Among current smokers in the registry with severe alpha-1 antitrypsin deficiency, the rate of FEV₁ decline was 109 mL/year.

Rates of FEV₁ decline over time vary among groups with differing degrees of airflow obstruction. For example, PI*ZZ patients with moderate COPD (stage II of the four-stage Global Initiative for Chronic Obstructive Lung Disease classification system) lose lung function faster than patients with either milder or more severe degrees of airflow obstruction.²⁹

As with COPD in general, exacerbations of COPD in people with severe deficiency of alpha-1 antitrypsin are associated with worsened clinical status. In one series,³⁰ 54% of 265 PI*ZZ patients experienced an exacerbation in the first year of follow-up, and 18% experienced at least three. Such exacerbations occurred in December and January in 32% of these individuals, likely due to a viral precipitant.

Increased mortality

Severe deficiency of alpha-1 antitrypsin is associated not only with severe morbidity but also death. In the national registry, the overall rate of death was 18.6% at 5 years of follow-up, or approximately 3% per year.²⁸

A low FEV₁ at entry was a bad sign. Patients entering the registry with FEV₁ values below 15% of predicted had a 36% mortality rate at 3 years, compared with 2.6% in those whose baseline FEV₁ exceeded 50% of predicted.

Underlying causes of death in registry participants included emphysema (accounting for 72% of deaths) and cirrhosis (10%),³¹ which were the only causes of death more frequent than in age- and sex-matched controls. In a series of never-smokers who had PI*ZZ alpha-1 antitrypsin deficiency,³² death was less frequently attributed to emphysema than in the national registry (46%) and more often attributed to cirrhosis (28%), indicating that never-smokers may more frequently escape the ravages of emphysema but experience a higher rate of developing cirrhosis later in life.³³

DIAGNOSING ALPHA-1 ANTITRYPSIN DEFICIENCY

Available blood tests for alpha-1 antitrypsin deficiency include:

The serum alpha-1 antitrypsin level, most often done by nephelometry. Normal serum levels generally range from 100 to 220 mg/dL.

Phenotyping, usually performed by isoelectric focusing, which can identify different band patterns associated with different alleles.

Genotyping involves determining which alpha-1 antitrypsin alleles are present, most often using polymerase chain reaction testing targeting the S and Z alleles and occasionally set up to detect less common alleles such as F and I.¹⁷

Gene sequencing is occasionally necessary to achieve an accurate, definitive  diagnosis.

Free, confidential testing is available

Clinical testing most often involves checking both a serum level and a phenotype or genotype. Such tests are often available in hospital laboratories and commercial laboratories, with testing also facilitated by the availability of free testing kits from several manufacturers of drugs for alpha-1 antitrypsin deficiency.

The Alpha-1 Foundation (www.alpha1.org)³⁴ also offers a free, home-based confidential testing kit through a research protocol at the Medical University of South Carolina (alphaone@musc.edu) called the Alpha-1 Coded Testing (ACT) study. Patients can receive a kit and lancet at home, submit the dried blood-spot specimen, and receive in the mail a confidential serum level and genotype.

The availability of such home-based confidential testing allows patients to seek testing without a physician’s order and makes it easier for facilitated allied health providers, such as respiratory therapists, to recommend testing in appropriate clinical circumstances.¹⁵

TREATMENT OF ALPHA-1 ANTITRYPSIN DEFICIENCY

The treatment of patients with severe deficiency of alpha-1 antitrypsin and emphysema generally resembles that of patients with usual COPD. Specifically, smoking cessation, bronchodilators, occasionally inhaled steroids, supplemental oxygen, preventive vaccinations, and pulmonary rehabilitation are indicated as per usual clinical assessment.

Lung volume reduction surgery, which is beneficial in appropriate subsets of COPD patients, is generally less effective in those with severe alpha-1 antitrypsin deficiency,³⁵ specifically because the magnitude of FEV₁ increase and the duration of such a rise are lower than in usual COPD patients.

Augmentation therapy

Specific therapy for alpha-1 antitrypsin deficiency currently involves weekly intravenous infusions of purified, pooled human-plasma-derived alpha-1 antitrypsin, so-called augmentation therapy. Four drugs have been approved for use in the United States:

• Prolastin-C (Grifols, Barcelona, Spain)
• Aralast NP (Baxalta, Bonneckborn, IL)
• Zemaira (CSL Behring, King of Prussia, PA)
• Glassia (Baxalta, Bonneckborn, IL, and Kamada, Ness Ziona, Israel).

All of these were approved for use in the United States on the basis of biochemical efficacy. Specifically, infusion of these drugs has been shown to raise serum levels above a protective threshold value (generally considered 57 mg/dL, the value below which the risk of developing emphysema increases beyond normal).

Randomized controlled trials^36,37 have addressed the efficacy of intravenous augmentation therapy, and although no single trial has been definitive, the weight of evidence shows that augmentation therapy can slow the progression of emphysema. For example, in a study by Dirksen et al,³⁷ augmentation therapy was associated with a slower progression of emphysema as assessed by the rate of loss of lung density on computed tomography.

On the basis of the available evidence, the American Thoracic Society and European Respiratory Society¹⁴ have recommended augmentation therapy in individuals with “established airflow obstruction from alpha-1 antitrypsin deficiency.”¹⁴ Their guidelines go on to say that the evidence that augmentation therapy is beneficial “is stronger for individuals with moderate airflow obstruction (eg, FEV₁ 35%–60% of predicted) than for those with severe airflow obstruction. Augmentation therapy is not currently recommended for individuals without emphysema.”

The guidelines recognize that although augmentation therapy does not satisfy the usual criteria for cost-effectiveness (< $50,000 per quality-adjusted life year) due to its high cost (approximately $100,000 per year if paid for out of pocket),³⁸ it is recommended for appropriate candidates because it is the only available specific therapy for severe deficiency of alpha-1 antitrypsin.

Novel therapies

In addition to current treatment approaches of augmentation therapy, a number of novel treatment strategies are being investigated, several of which hold much promise.

Gene therapy, using adeno-associated virus to transfect the normal human gene into individuals with severe deficiency of alpha-1 antitrypsin, has been undertaken and is currently under study. In addition, a variety of approaches to interdict production of abnormal Z protein from the liver are being examined, as well as inhaled hyaluronic acid to protect the lung.

Alpha-1 antitrypsin deficiency is a common but underrecognized genetic condition that increases the risk of chronic obstructive pulmonary disease (COPD) and liver disease. Primary care providers can play a critical role in detecting it and managing patients who have it.

RECOGNIZED CASES ARE THE TIP OF THE ICEBERG

First described in 1963,¹ alpha-1 antitrypsin deficiency is estimated to affect 100,000 Americans, fewer than 15,000 of whom have received a clinical diagnosis. As further evidence of its underrecognition,^2–7 many patients experience long delays between their first symptoms and the diagnosis. Early studies indicated that the average diagnostic delay was 7.2 years,4 and the latest studies, as recent as 2013, indicate a similar diagnostic delay.⁷

Furthermore, many patients see multiple healthcare providers before receiving the correct diagnosis. A 1994 survey by this author⁴ found that 43.7% of patients who had severe deficiency of alpha-1 antitrypsin saw at least three physicians before the correct diagnosis was made.

Why is the disease underrecognized?

Several reasons may account for underrecognition of this disease. Many clinicians—including, unfortunately, many pulmonologists—do not know much about it,^7,8 do not adhere to clinical guidelines,^9,10 or harbor the misperception that there is no therapy available and, therefore, no compelling reason to make a diagnosis.⁷

Regarding inadequate knowledge, in a study by Taliercio, Chatburn, and this author,⁸ internal medicine residents scored only 63% correct on a 10-question quiz on diagnostic features of alpha-1 antitrypsin deficiency. There was no evidence of a training effect—senior residents scored no higher than interns.

Similarly, when Greulich et al⁷ surveyed German and Italian internists, general practitioners, and pulmonologists, one-fourth to one-half of them (depending on specialty and country) stated that they knew either very little or nothing at all about alpha-1 antitrypsin deficiency. In addition, 7% to 8% agreed with the statement, “There is no treatment available for this disease.”⁷

Nonadoption of clinical guidelines has been widely recognized in medicine and is evident in the failure to implement various recommended practices,^9,10 such as low-stretch ventilation for acute respiratory distress syndrome and prophylaxis against deep vein thrombosis.

Finding the rest of the iceberg

Efforts to enhance compliance with guidelines on testing for alpha-1 antitrypsin deficiency have included using the electronic medical record to prompt physicians to test appropriate candidates.^11–13

Jain et al¹³ examined the effect of installing such a prompting system to remind physicians to test for alpha-1 antitrypsin deficiency in patients with airflow obstruction that does not reverse with a bronchodilator—a recognized indication for testing for this disease according to standards endorsed by the American Thoracic Society and European Respiratory Society.¹⁴ At baseline, only 4.7% of appropriate candidates were being tested; after a prompt was installed in the electronic medical record, the rate rose to 15.1%, still a minority of candidates.

Another strategy is to empower respiratory therapists who perform pulmonary function tests to invite patients to be tested if their pulmonary function tests show postbronchodilator airflow obstruction. Rahaghi et al¹⁵ showed that using this strategy, 20 (0.63%) of 3,152 patients who were found to have fixed airflow obstruction when they underwent pulmonary function testing were newly diagnosed with severe deficiency of alpha-1 antitrypsin. Other targeted detection studies in patients with COPD estimated the prevalence of alpha-1 antitrypsin deficiency at up to 12%.³

PHYSIOLOGY AND PATHOPHYSIOLOGY OF ALPHA-1 ANTITRYPSIN DEFICIENCY

Alpha-1 antitrypsin is a single-chain, 394-amino acid glycoprotein with three carbohydrate side chains found at asparagine residues along the primary structure.¹⁶

A major physiologic function of this molecule is to bind neutrophil elastase, which it does avidly. In a “mousetrap-like” mechanism,¹⁶ an active site on the alpha-1 antitrypsin molecule captures the neutrophil elastase and is cleaved, releasing steric energy in the molecule, catapulting the neutrophil elastase to the opposite side of the alpha-1 antitrypsin molecule, and inactivating it (Figure 1).

MM is normal, ZZ is not

Alpha-1 antitrypsin deficiency is inherited as an autosomal-codominant condition.¹⁷

The SERPINA1 gene, which codes for alpha-1 antitrypsin, is located on the long arm of the 14th chromosome, and more than 150 alleles of this gene have been identified to date. The normal allele is denoted M, and the allele most commonly associated with severe deficiency is denoted Z. People who are homozygous for the M allele (ie, normal) are called PI*MM (PI stands for “protease inhibitor”), and those who are homozygous for the Z allele are PI*ZZ. More than 90% of patients with severe alpha-1 antitrypsin deficiency are PI*ZZ.¹⁸

The Z allele has a single amino acid substitution (glutamic acid-to-lysine at position 342), which results in abnormal folding and formation of polymers of the Z molecule within hepatocytes.^19,20 These polymers are recognized on liver biopsy as periodic acid-Schiff diastase-resistant eosinophilic inclusion bodies on histologic staining (Figure 2).

With alpha-1 antitrypsin trapped as Z-molecule polymers in the liver, the amount in the bloodstream falls, and there is a consequent decrease in the amount available in the lung to oppose the proteolytic burden of neutrophil elastase, especially in people who smoke or work in dusty environments.²¹

Tan et al²² have shown that some of the polymerized Z protein can escape the liver and circulate in the blood and that alveolar macrophages may also produce Z polymers. These Z polymers are chemotactic for neutrophils,²³ so that their presence in the lung fuels the inflammatory cascade by recruiting more neutrophils to the lung, thereby increasing the proteolytic burden to the lung and increasing the risk of emphysema. Z monomers that do circulate can bind neutrophil elastase, but their binding avidity to neutrophil elastase is substantially lower than that of M-type alpha-1 antitrypsin.

CLINICAL MANIFESTATIONS

Alpha-1 antitrypsin deficiency of the PI*ZZ type is associated with two major clinical manifestations:

• Emphysema, resulting from the loss of proteolytic protection of the lung by alpha-1 antitrypsin (a toxic loss of function), and
• Liver diseases such as cirrhosis and chronic hepatitis, which result from abnormal accumulation of alpha-1 antitrypsin within hepatocytes (a toxic gain of function), and hepatoma.¹⁷

Other clinical manifestations of PI*ZZ alpha-1 antitrypsin deficiency include panniculitis and an association with cytoplasmic antineutrophil cytoplasmic antibody-positive vasculitis.¹⁷

Some uncertainty exists regarding the risk associated with the PI*MZ heterozygous state because there has been no systematic longitudinal study of people with this genotype. However, the weight of available experience suggests that PI*MZ individuals who have never smoked are not at increased risk of developing emphysema.²⁴

Findings from a national registry: PI*ZZ COPD resembles ‘usual’ COPD

Distinguishing patients with alpha-1 antitrypsin deficiency from those with “usual” COPD (ie, without alpha-1 antitrypsin deficiency) can be difficult, as shown in data from the National Heart, Lung, and Blood Institute’s   Alpha-1 Antitrypsin Deficiency Registry study.¹⁸ This multicenter, longitudinal, observational study contains the largest well-characterized cohort with severe deficiency of alpha-1 antitrypsin (PI*ZZ, PI*ZNull, etc), with 1,129 patients. 

Pulmonary function test results were consistent with emphysema in most of the patients in the registry. Mean postbronchodilator pulmonary function values (± standard error of the mean) were:

• Forced expiratory volume in 1 second (FEV₁) 46.7% of predicted (± 30%)
• Ratio of FEV₁ to forced vital capacity 42.9% (± 20.4% )
• Mean diffusing capacity for carbon monoxide 50.3% of predicted (± 22.5%).

Like many patients with usual COPD, 60% of the registry patients demonstrated a component of airway reactivity, with significant reversal of airflow obstruction over three spirometries after receiving a dose of an inhaled bronchodilator (characterized by a 12% and 200-mL postbronchodilator rise in FEV₁). Moreover, 78 patients had normal lung function.

Symptoms also resembled those in patients with usual emphysema, chronic bronchitis, or both. On enrollment in the registry, 83.9% of the patients had shortness of breath on exertion, 75.5% had wheezing with upper respiratory infections, 65.3% had wheezing without upper respiratory infection, 67.6% had recent debilitating chest illness, 42.4% had “usual” cough, and 49.6% had annual cough and phlegm episodes.

Imaging findings. Although the classic teaching is that emphysema due to alpha-1 antitrypsin deficiency produces lower-lobe hyperlucency on plain films, relying on this sign would lead to underrecognition, as 36% of PI*ZZ patients have apical-predominant emphysema on chest computed tomography,²⁴ which resembles the usual centriacinar emphysema pattern. Figure 3 shows axial computed tomographic scans through the apices and the bases of the lungs of a patient with alpha-1 antitrypsin deficiency.  

In view of these difficulties, guidelines from the American Thoracic Society and European Respiratory Society¹⁴ endorse testing for alpha-1 antitrypsin deficiency in all adults who have symptoms and fixed airflow obstruction (Table 1).

CONSEQUENCES OF ALPHA-1 ANTITRYPSIN DEFICIENCY

Two large screening studies^2,3,25,26 followed people who were identified at birth as having alpha-1 antitrypsin deficiency to examine the natural course of the disease.

The larger of the two studies²⁷ tested 200,000 Swedish newborns. Follow-up of this cohort to age 35 indicated that 35-year-old never-smoking PI*ZZ individuals have normal lung function and no excess emphysema on computed tomography compared with normal peers matched for age and sex.²⁷ In contrast, the few PI*ZZ ever-smokers demonstrated a lower level of transfer factor and significantly more emphysema on computed tomography than normal (PI*MM) never-smokers.

Faster decline in lung function

Data from the National Heart, Lung, and Blood Institute registry indicate that, on average, people with severe alpha-1 antitrypsin deficiency lose lung function faster than people without the disease.²⁸ Specifically, in never-smokers in the registry, the average rate of FEV₁ decline was 67 mL/year, and among ex-smokers, it was 54 mL/year. Both of these values exceed the general age-related rate of FEV₁ decline of approximately 20 to 25 mL/year in never-smoking, normal adults. Among current smokers in the registry with severe alpha-1 antitrypsin deficiency, the rate of FEV₁ decline was 109 mL/year.

Rates of FEV₁ decline over time vary among groups with differing degrees of airflow obstruction. For example, PI*ZZ patients with moderate COPD (stage II of the four-stage Global Initiative for Chronic Obstructive Lung Disease classification system) lose lung function faster than patients with either milder or more severe degrees of airflow obstruction.²⁹

As with COPD in general, exacerbations of COPD in people with severe deficiency of alpha-1 antitrypsin are associated with worsened clinical status. In one series,³⁰ 54% of 265 PI*ZZ patients experienced an exacerbation in the first year of follow-up, and 18% experienced at least three. Such exacerbations occurred in December and January in 32% of these individuals, likely due to a viral precipitant.

Increased mortality

Severe deficiency of alpha-1 antitrypsin is associated not only with severe morbidity but also death. In the national registry, the overall rate of death was 18.6% at 5 years of follow-up, or approximately 3% per year.²⁸

A low FEV₁ at entry was a bad sign. Patients entering the registry with FEV₁ values below 15% of predicted had a 36% mortality rate at 3 years, compared with 2.6% in those whose baseline FEV₁ exceeded 50% of predicted.

Underlying causes of death in registry participants included emphysema (accounting for 72% of deaths) and cirrhosis (10%),³¹ which were the only causes of death more frequent than in age- and sex-matched controls. In a series of never-smokers who had PI*ZZ alpha-1 antitrypsin deficiency,³² death was less frequently attributed to emphysema than in the national registry (46%) and more often attributed to cirrhosis (28%), indicating that never-smokers may more frequently escape the ravages of emphysema but experience a higher rate of developing cirrhosis later in life.³³

DIAGNOSING ALPHA-1 ANTITRYPSIN DEFICIENCY

Available blood tests for alpha-1 antitrypsin deficiency include:

The serum alpha-1 antitrypsin level, most often done by nephelometry. Normal serum levels generally range from 100 to 220 mg/dL.

Phenotyping, usually performed by isoelectric focusing, which can identify different band patterns associated with different alleles.

Genotyping involves determining which alpha-1 antitrypsin alleles are present, most often using polymerase chain reaction testing targeting the S and Z alleles and occasionally set up to detect less common alleles such as F and I.¹⁷

Gene sequencing is occasionally necessary to achieve an accurate, definitive  diagnosis.

Free, confidential testing is available

Clinical testing most often involves checking both a serum level and a phenotype or genotype. Such tests are often available in hospital laboratories and commercial laboratories, with testing also facilitated by the availability of free testing kits from several manufacturers of drugs for alpha-1 antitrypsin deficiency.

The Alpha-1 Foundation (www.alpha1.org)³⁴ also offers a free, home-based confidential testing kit through a research protocol at the Medical University of South Carolina (alphaone@musc.edu) called the Alpha-1 Coded Testing (ACT) study. Patients can receive a kit and lancet at home, submit the dried blood-spot specimen, and receive in the mail a confidential serum level and genotype.

The availability of such home-based confidential testing allows patients to seek testing without a physician’s order and makes it easier for facilitated allied health providers, such as respiratory therapists, to recommend testing in appropriate clinical circumstances.¹⁵

TREATMENT OF ALPHA-1 ANTITRYPSIN DEFICIENCY

The treatment of patients with severe deficiency of alpha-1 antitrypsin and emphysema generally resembles that of patients with usual COPD. Specifically, smoking cessation, bronchodilators, occasionally inhaled steroids, supplemental oxygen, preventive vaccinations, and pulmonary rehabilitation are indicated as per usual clinical assessment.

Lung volume reduction surgery, which is beneficial in appropriate subsets of COPD patients, is generally less effective in those with severe alpha-1 antitrypsin deficiency,³⁵ specifically because the magnitude of FEV₁ increase and the duration of such a rise are lower than in usual COPD patients.

Augmentation therapy

Specific therapy for alpha-1 antitrypsin deficiency currently involves weekly intravenous infusions of purified, pooled human-plasma-derived alpha-1 antitrypsin, so-called augmentation therapy. Four drugs have been approved for use in the United States:

• Prolastin-C (Grifols, Barcelona, Spain)
• Aralast NP (Baxalta, Bonneckborn, IL)
• Zemaira (CSL Behring, King of Prussia, PA)
• Glassia (Baxalta, Bonneckborn, IL, and Kamada, Ness Ziona, Israel).

All of these were approved for use in the United States on the basis of biochemical efficacy. Specifically, infusion of these drugs has been shown to raise serum levels above a protective threshold value (generally considered 57 mg/dL, the value below which the risk of developing emphysema increases beyond normal).

Randomized controlled trials^36,37 have addressed the efficacy of intravenous augmentation therapy, and although no single trial has been definitive, the weight of evidence shows that augmentation therapy can slow the progression of emphysema. For example, in a study by Dirksen et al,³⁷ augmentation therapy was associated with a slower progression of emphysema as assessed by the rate of loss of lung density on computed tomography.

On the basis of the available evidence, the American Thoracic Society and European Respiratory Society¹⁴ have recommended augmentation therapy in individuals with “established airflow obstruction from alpha-1 antitrypsin deficiency.”¹⁴ Their guidelines go on to say that the evidence that augmentation therapy is beneficial “is stronger for individuals with moderate airflow obstruction (eg, FEV₁ 35%–60% of predicted) than for those with severe airflow obstruction. Augmentation therapy is not currently recommended for individuals without emphysema.”

The guidelines recognize that although augmentation therapy does not satisfy the usual criteria for cost-effectiveness (< $50,000 per quality-adjusted life year) due to its high cost (approximately $100,000 per year if paid for out of pocket),³⁸ it is recommended for appropriate candidates because it is the only available specific therapy for severe deficiency of alpha-1 antitrypsin.

Novel therapies

In addition to current treatment approaches of augmentation therapy, a number of novel treatment strategies are being investigated, several of which hold much promise.

Gene therapy, using adeno-associated virus to transfect the normal human gene into individuals with severe deficiency of alpha-1 antitrypsin, has been undertaken and is currently under study. In addition, a variety of approaches to interdict production of abnormal Z protein from the liver are being examined, as well as inhaled hyaluronic acid to protect the lung.

Inheriting the wrong genes and eating the wrong food (ie, gluten) are necessary for celiac disease to develop, but are not enough by themselves. Something else must be contributing, and evidence is pointing to the mix of bacteria that make our guts their home, collectively called the microbiome.

See related article

Celiac disease is a highly prevalent, chronic, immune-mediated form of enteropathy.¹ It affects 0.5% to 1% of the population, and although it is mostly seen in people of northern European descent, those in other populations can develop the disease as well. Historically, celiac disease was classified as an infant condition. However, it now commonly presents later in life (between ages 10 and 40) and often with extraintestinal manifestations.²

In this issue of Cleveland Clinic Journal of Medicine, Kochhar et al provide a comprehensive updated review of celiac disease.³

GENES AND GLUTEN ARE NECESSARY BUT NOT SUFFICIENT

Although genetic factors and exposure to gluten in the diet are proven to be necessary for celiac disease to develop, they are not sufficient. Evidence of this is in the numbers; although one-third of the general population carries the HLA susceptibility genes (specifically HLA-DQ2 and DQ8),⁴ only 2% to 5% of people with these genes develop clinically evident celiac disease.

Additional environmental factors must be contributing to disease development, but these other factors are poorly understood. Some of the possible culprits that might influence the risk of disease occurrence and the timing of its onset include⁵:

• The amount and quality of gluten ingested—the higher the concentration of gluten, the higher the risk, and different grains have gluten varieties with more or less immunogenic capabilities, ie, T-cell activation properties
• The pattern of infant feeding—the risk may be lower with breastfeeding than with formula
• The age at which gluten is introduced into the diet—the risk may be higher if gluten is introduced earlier.⁶

More recently, studies of the pathogenesis of celiac disease and gene-environmental interactions have expanded beyond host predisposition and dietary factors.

OUR BODIES, OUR MICROBIOMES: A SYMBIOTIC RELATIONSHIP

The role of the human microbiome in autoimmune disease is now being elucidated.⁷ Remarkably, the microorganisms living in our bodies outnumber our body cells by a factor of 10, and their genomes vastly exceed our own protein-coding genome capabilities by a factor of 100.

The gut microbiome is now considered a true bioreactor with enzymatic and immunologic capabilities beyond (and complementary to) those of its host. The commensal microbiome of the host intestine provides benefits that can be broken down into three broad categories:

• Nutritional—producing essential amino acids and vitamins
• Metabolic—degrading complex polysaccharides from dietary fibers
• Immunologic—shaping the host immune system while cooperating with it against pathogenic microorganisms.

The immunologic function is highly relevant. We have coevolved with our bacteria in a mutually beneficial, symbiotic relationship in which we maintain an active state of low inflammation so that a constant bacterial and dietary antigenic load can be tolerated.

Evidence points to dysbiosis as a factor leading to celiac disease and other autoimmune disorders

Is there a core human microbiome shared by all individuals? And what is the impact of altering the relative microbial composition (dysbiosis) in physiologic and disease states? To find out, the National Institutes of Health launched the Human Microbiome Project⁸ in 2008. Important tools in this work include novel culture-independent approaches (high-throughput DNA sequencing and whole-microbiome “shotgun” sequencing with metagenomic analysis) and computational analytical tools.⁹

An accumulating body of evidence is now available from animal models and human studies correlating states of intestinal dysbiosis (disruption in homeostatic community composition) with various disease processes. These have ranged from inflammatory bowel disease to systemic autoimmune disorders such as psoriasis, inflammatory arthropathies, and demyelinating central nervous system diseases.^10–14

RESEARCH INTO THE MICROBIOME IN CELIAC DISEASE

Celiac disease has also served as a unique model for studying this biologic relationship, and the microbiome has been postulated to have a role in its pathogenesis.¹⁵ Multiple clinical studies demonstrate that a state of intestinal dysbiosis is indeed associated with celiac disease.

Specifically, decreases in the abundance of Firmicutes spp and increases in Proteobacteria spp have been detected in both children and adults with active celiac disease.^16,17 Intriguingly, overrepresentation of Proteobacteria was also correlated with disease activity. Other studies have reported decreases in the proportion of reportedly protective, anti-inflammatory bacteria such as Bifidobacterium and increases in the proportion of Bacteroides and Escherichia coli in patients with active disease.^18,19 Altered diversity and altered metabolic function, ie, decreased concentration of protective short-chain fatty acids of the microbiota, have also been reported in patients with celiac disease.^19,20

To move beyond correlative studies and mechanistically address the possibility of causation, multiple groups have used a gnotobiotic approach, ie, maintaining animals under germ-free conditions and incorporating microbes of interest. This approach is highly relevant in studying whether the bacterial community composition is capable of modulating loss of tolerance to gluten in genetically susceptible hosts. A few notable examples have been published.

In germ-free rats, long-term feeding of gliadin, but not albumin, from birth until 2 months of age induced moderate small-intestinal damage.²¹ Similarly, germ-free nonobese diabetic-DQ8 mice developed more severe gluten-induced disease than mice with normal intestinal bacteria.²²

In small studies, people with celiac disease had fewer Firmicutes and Bifidobacteria and more Proteobacteria, Bacteroides, and E coli

These findings suggest that the normal gut microbiome may have intrinsic beneficial properties capable of reducing the inflammatory effects associated with gluten ingestion. Notably, the specific composition of the intestinal microbiome can define the fate of gluten-induced pathology. Mice colonized with commensal microbiota are indeed protected from gluten-induced pathology, while mice colonized with Proteobacteria spp develop a moderate degree of gluten-induced disease. When Escherichia coli derived from patients with celiac disease is added to commensal colonization, the celiac disease-like phenotype develops.²³

Taken together, these studies support the hypothesis that the intestinal microbiome may be another environmental factor involved in the development of celiac disease.

QUESTIONS AND CHALLENGES REMAIN

The results of clinical studies are not necessarily consistent at the taxonomy level. The fields of metagenomics, which investigates all genes and their enzymatic function in a given community, and metabolomics, which identifies bacterial end-products, characterizing their functional capabilities, are still in their infancy and will be required to further investigate functionality of the altered microbiome in celiac disease.

Second, the directionality—the causality or consequences of this dysbiosis—and timing—the moment at which changes occur, ie, after introducing gluten or at the time when symptoms appear—remain elusive, and prospective studies in humans will be essential.

Finally, more mechanistic studies in animal models are needed to dissect the host immune response to dietary gluten and perturbation of intestinal community composition. This may lead to the possibility of future interventions in the form of prebiotics, probiotics, or specific metabolites, complementary to gluten avoidance.

In the meantime, increasing disease awareness and rapid diagnosis and treatment continue to be of utmost importance to address the clinical consequences of celiac disease in both children and adults.

Inheriting the wrong genes and eating the wrong food (ie, gluten) are necessary for celiac disease to develop, but are not enough by themselves. Something else must be contributing, and evidence is pointing to the mix of bacteria that make our guts their home, collectively called the microbiome.

See related article

Celiac disease is a highly prevalent, chronic, immune-mediated form of enteropathy.¹ It affects 0.5% to 1% of the population, and although it is mostly seen in people of northern European descent, those in other populations can develop the disease as well. Historically, celiac disease was classified as an infant condition. However, it now commonly presents later in life (between ages 10 and 40) and often with extraintestinal manifestations.²

In this issue of Cleveland Clinic Journal of Medicine, Kochhar et al provide a comprehensive updated review of celiac disease.³

GENES AND GLUTEN ARE NECESSARY BUT NOT SUFFICIENT

Although genetic factors and exposure to gluten in the diet are proven to be necessary for celiac disease to develop, they are not sufficient. Evidence of this is in the numbers; although one-third of the general population carries the HLA susceptibility genes (specifically HLA-DQ2 and DQ8),⁴ only 2% to 5% of people with these genes develop clinically evident celiac disease.

Additional environmental factors must be contributing to disease development, but these other factors are poorly understood. Some of the possible culprits that might influence the risk of disease occurrence and the timing of its onset include⁵:

• The amount and quality of gluten ingested—the higher the concentration of gluten, the higher the risk, and different grains have gluten varieties with more or less immunogenic capabilities, ie, T-cell activation properties
• The pattern of infant feeding—the risk may be lower with breastfeeding than with formula
• The age at which gluten is introduced into the diet—the risk may be higher if gluten is introduced earlier.⁶

More recently, studies of the pathogenesis of celiac disease and gene-environmental interactions have expanded beyond host predisposition and dietary factors.

OUR BODIES, OUR MICROBIOMES: A SYMBIOTIC RELATIONSHIP

The role of the human microbiome in autoimmune disease is now being elucidated.⁷ Remarkably, the microorganisms living in our bodies outnumber our body cells by a factor of 10, and their genomes vastly exceed our own protein-coding genome capabilities by a factor of 100.

The gut microbiome is now considered a true bioreactor with enzymatic and immunologic capabilities beyond (and complementary to) those of its host. The commensal microbiome of the host intestine provides benefits that can be broken down into three broad categories:

• Nutritional—producing essential amino acids and vitamins
• Metabolic—degrading complex polysaccharides from dietary fibers
• Immunologic—shaping the host immune system while cooperating with it against pathogenic microorganisms.

The immunologic function is highly relevant. We have coevolved with our bacteria in a mutually beneficial, symbiotic relationship in which we maintain an active state of low inflammation so that a constant bacterial and dietary antigenic load can be tolerated.

Evidence points to dysbiosis as a factor leading to celiac disease and other autoimmune disorders

Is there a core human microbiome shared by all individuals? And what is the impact of altering the relative microbial composition (dysbiosis) in physiologic and disease states? To find out, the National Institutes of Health launched the Human Microbiome Project⁸ in 2008. Important tools in this work include novel culture-independent approaches (high-throughput DNA sequencing and whole-microbiome “shotgun” sequencing with metagenomic analysis) and computational analytical tools.⁹

An accumulating body of evidence is now available from animal models and human studies correlating states of intestinal dysbiosis (disruption in homeostatic community composition) with various disease processes. These have ranged from inflammatory bowel disease to systemic autoimmune disorders such as psoriasis, inflammatory arthropathies, and demyelinating central nervous system diseases.^10–14

RESEARCH INTO THE MICROBIOME IN CELIAC DISEASE

Celiac disease has also served as a unique model for studying this biologic relationship, and the microbiome has been postulated to have a role in its pathogenesis.¹⁵ Multiple clinical studies demonstrate that a state of intestinal dysbiosis is indeed associated with celiac disease.

Specifically, decreases in the abundance of Firmicutes spp and increases in Proteobacteria spp have been detected in both children and adults with active celiac disease.^16,17 Intriguingly, overrepresentation of Proteobacteria was also correlated with disease activity. Other studies have reported decreases in the proportion of reportedly protective, anti-inflammatory bacteria such as Bifidobacterium and increases in the proportion of Bacteroides and Escherichia coli in patients with active disease.^18,19 Altered diversity and altered metabolic function, ie, decreased concentration of protective short-chain fatty acids of the microbiota, have also been reported in patients with celiac disease.^19,20

To move beyond correlative studies and mechanistically address the possibility of causation, multiple groups have used a gnotobiotic approach, ie, maintaining animals under germ-free conditions and incorporating microbes of interest. This approach is highly relevant in studying whether the bacterial community composition is capable of modulating loss of tolerance to gluten in genetically susceptible hosts. A few notable examples have been published.

In germ-free rats, long-term feeding of gliadin, but not albumin, from birth until 2 months of age induced moderate small-intestinal damage.²¹ Similarly, germ-free nonobese diabetic-DQ8 mice developed more severe gluten-induced disease than mice with normal intestinal bacteria.²²

In small studies, people with celiac disease had fewer Firmicutes and Bifidobacteria and more Proteobacteria, Bacteroides, and E coli

These findings suggest that the normal gut microbiome may have intrinsic beneficial properties capable of reducing the inflammatory effects associated with gluten ingestion. Notably, the specific composition of the intestinal microbiome can define the fate of gluten-induced pathology. Mice colonized with commensal microbiota are indeed protected from gluten-induced pathology, while mice colonized with Proteobacteria spp develop a moderate degree of gluten-induced disease. When Escherichia coli derived from patients with celiac disease is added to commensal colonization, the celiac disease-like phenotype develops.²³

Taken together, these studies support the hypothesis that the intestinal microbiome may be another environmental factor involved in the development of celiac disease.

QUESTIONS AND CHALLENGES REMAIN

The results of clinical studies are not necessarily consistent at the taxonomy level. The fields of metagenomics, which investigates all genes and their enzymatic function in a given community, and metabolomics, which identifies bacterial end-products, characterizing their functional capabilities, are still in their infancy and will be required to further investigate functionality of the altered microbiome in celiac disease.

Second, the directionality—the causality or consequences of this dysbiosis—and timing—the moment at which changes occur, ie, after introducing gluten or at the time when symptoms appear—remain elusive, and prospective studies in humans will be essential.

Finally, more mechanistic studies in animal models are needed to dissect the host immune response to dietary gluten and perturbation of intestinal community composition. This may lead to the possibility of future interventions in the form of prebiotics, probiotics, or specific metabolites, complementary to gluten avoidance.

In the meantime, increasing disease awareness and rapid diagnosis and treatment continue to be of utmost importance to address the clinical consequences of celiac disease in both children and adults.

Inheriting the wrong genes and eating the wrong food (ie, gluten) are necessary for celiac disease to develop, but are not enough by themselves. Something else must be contributing, and evidence is pointing to the mix of bacteria that make our guts their home, collectively called the microbiome.

See related article

Celiac disease is a highly prevalent, chronic, immune-mediated form of enteropathy.¹ It affects 0.5% to 1% of the population, and although it is mostly seen in people of northern European descent, those in other populations can develop the disease as well. Historically, celiac disease was classified as an infant condition. However, it now commonly presents later in life (between ages 10 and 40) and often with extraintestinal manifestations.²

In this issue of Cleveland Clinic Journal of Medicine, Kochhar et al provide a comprehensive updated review of celiac disease.³

GENES AND GLUTEN ARE NECESSARY BUT NOT SUFFICIENT

Although genetic factors and exposure to gluten in the diet are proven to be necessary for celiac disease to develop, they are not sufficient. Evidence of this is in the numbers; although one-third of the general population carries the HLA susceptibility genes (specifically HLA-DQ2 and DQ8),⁴ only 2% to 5% of people with these genes develop clinically evident celiac disease.

Additional environmental factors must be contributing to disease development, but these other factors are poorly understood. Some of the possible culprits that might influence the risk of disease occurrence and the timing of its onset include⁵:

• The amount and quality of gluten ingested—the higher the concentration of gluten, the higher the risk, and different grains have gluten varieties with more or less immunogenic capabilities, ie, T-cell activation properties
• The pattern of infant feeding—the risk may be lower with breastfeeding than with formula
• The age at which gluten is introduced into the diet—the risk may be higher if gluten is introduced earlier.⁶

More recently, studies of the pathogenesis of celiac disease and gene-environmental interactions have expanded beyond host predisposition and dietary factors.

OUR BODIES, OUR MICROBIOMES: A SYMBIOTIC RELATIONSHIP

The role of the human microbiome in autoimmune disease is now being elucidated.⁷ Remarkably, the microorganisms living in our bodies outnumber our body cells by a factor of 10, and their genomes vastly exceed our own protein-coding genome capabilities by a factor of 100.

The gut microbiome is now considered a true bioreactor with enzymatic and immunologic capabilities beyond (and complementary to) those of its host. The commensal microbiome of the host intestine provides benefits that can be broken down into three broad categories:

• Nutritional—producing essential amino acids and vitamins
• Metabolic—degrading complex polysaccharides from dietary fibers
• Immunologic—shaping the host immune system while cooperating with it against pathogenic microorganisms.

The immunologic function is highly relevant. We have coevolved with our bacteria in a mutually beneficial, symbiotic relationship in which we maintain an active state of low inflammation so that a constant bacterial and dietary antigenic load can be tolerated.

Evidence points to dysbiosis as a factor leading to celiac disease and other autoimmune disorders

Is there a core human microbiome shared by all individuals? And what is the impact of altering the relative microbial composition (dysbiosis) in physiologic and disease states? To find out, the National Institutes of Health launched the Human Microbiome Project⁸ in 2008. Important tools in this work include novel culture-independent approaches (high-throughput DNA sequencing and whole-microbiome “shotgun” sequencing with metagenomic analysis) and computational analytical tools.⁹

An accumulating body of evidence is now available from animal models and human studies correlating states of intestinal dysbiosis (disruption in homeostatic community composition) with various disease processes. These have ranged from inflammatory bowel disease to systemic autoimmune disorders such as psoriasis, inflammatory arthropathies, and demyelinating central nervous system diseases.^10–14

RESEARCH INTO THE MICROBIOME IN CELIAC DISEASE

Celiac disease has also served as a unique model for studying this biologic relationship, and the microbiome has been postulated to have a role in its pathogenesis.¹⁵ Multiple clinical studies demonstrate that a state of intestinal dysbiosis is indeed associated with celiac disease.

Specifically, decreases in the abundance of Firmicutes spp and increases in Proteobacteria spp have been detected in both children and adults with active celiac disease.^16,17 Intriguingly, overrepresentation of Proteobacteria was also correlated with disease activity. Other studies have reported decreases in the proportion of reportedly protective, anti-inflammatory bacteria such as Bifidobacterium and increases in the proportion of Bacteroides and Escherichia coli in patients with active disease.^18,19 Altered diversity and altered metabolic function, ie, decreased concentration of protective short-chain fatty acids of the microbiota, have also been reported in patients with celiac disease.^19,20

To move beyond correlative studies and mechanistically address the possibility of causation, multiple groups have used a gnotobiotic approach, ie, maintaining animals under germ-free conditions and incorporating microbes of interest. This approach is highly relevant in studying whether the bacterial community composition is capable of modulating loss of tolerance to gluten in genetically susceptible hosts. A few notable examples have been published.

In germ-free rats, long-term feeding of gliadin, but not albumin, from birth until 2 months of age induced moderate small-intestinal damage.²¹ Similarly, germ-free nonobese diabetic-DQ8 mice developed more severe gluten-induced disease than mice with normal intestinal bacteria.²²

In small studies, people with celiac disease had fewer Firmicutes and Bifidobacteria and more Proteobacteria, Bacteroides, and E coli

These findings suggest that the normal gut microbiome may have intrinsic beneficial properties capable of reducing the inflammatory effects associated with gluten ingestion. Notably, the specific composition of the intestinal microbiome can define the fate of gluten-induced pathology. Mice colonized with commensal microbiota are indeed protected from gluten-induced pathology, while mice colonized with Proteobacteria spp develop a moderate degree of gluten-induced disease. When Escherichia coli derived from patients with celiac disease is added to commensal colonization, the celiac disease-like phenotype develops.²³

Taken together, these studies support the hypothesis that the intestinal microbiome may be another environmental factor involved in the development of celiac disease.

QUESTIONS AND CHALLENGES REMAIN

The results of clinical studies are not necessarily consistent at the taxonomy level. The fields of metagenomics, which investigates all genes and their enzymatic function in a given community, and metabolomics, which identifies bacterial end-products, characterizing their functional capabilities, are still in their infancy and will be required to further investigate functionality of the altered microbiome in celiac disease.

Second, the directionality—the causality or consequences of this dysbiosis—and timing—the moment at which changes occur, ie, after introducing gluten or at the time when symptoms appear—remain elusive, and prospective studies in humans will be essential.

Finally, more mechanistic studies in animal models are needed to dissect the host immune response to dietary gluten and perturbation of intestinal community composition. This may lead to the possibility of future interventions in the form of prebiotics, probiotics, or specific metabolites, complementary to gluten avoidance.

In the meantime, increasing disease awareness and rapid diagnosis and treatment continue to be of utmost importance to address the clinical consequences of celiac disease in both children and adults.

Celiac disease is an autoimmune disorder that occurs in genetically predisposed individuals in response to ingestion of gluten. Its prevalence is about 0.7% of the US population.¹

See related editorial

The gold standard for diagnosis is duodenal biopsy, in which the histologic features may include varying gradations of flattening of intestinal villi, crypt hyperplasia, and infiltration of the lamina propria by lymphocytes. Many patients have no symptoms at the time of diagnosis, but presenting symptoms can include diarrhea along with features of malabsorption,² and, in about 25% of patients (mainly adults), a bullous cutaneous disorder called dermatitis herpetiformis.^3,4 The pathogenesis of celiac disease and that of dermatitis herpetiformis are similar in that in both, ingestion of gluten induces an inflammatory reaction leading to the clinical manifestations.

The mainstay of treatment of celiac disease remains avoidance of gluten in the diet.

GENETIC PREDISPOSITION AND DIETARY TRIGGER

The pathogenesis of celiac disease has been well studied in both humans and animals. The disease is thought to develop by an interplay of genetic and autoimmune factors and the ingestion of gluten (ie, an environmental factor).

Celiac disease occurs in genetically predisposed individuals, ie, those who carry the HLA alleles DQ2 (DQA1*05, DQB1*02), DQ8 (DQA1*03, DQB1*0302), or both.⁵

Ingestion of gluten is necessary for the disease to develop. Gluten, the protein component of wheat, barley, and rye, contains proteins called prolamins, which vary among the different types of grain. In wheat, the prolamin is gliadin, which is alcohol-soluble. In barley the prolamin is hordein, and in rye it is secalin.⁴ The prolamin content in gluten makes it resistant to degradation by gastric, pancreatic, and intestinal brush border proteases.⁶ Gluten crosses the epithelial barrier and promotes an inflammatory reaction by both the innate and adaptive immune systems that can ultimately result in flattening of villi and crypt hyperplasia (Figure 1).⁷

Tissue transglutaminase also plays a central role in the pathogenesis, as it further deaminates gliadin and increases its immunogenicity by causing it to bind to receptors on antigen-presenting cells with stronger affinity. Furthermore, gliadin-tissue transglutaminase complexes formed by protein cross-linkages generate an autoantibody response (predominantly immunoglobulin A [IgA] type) that can exacerbate the inflammatory process.^8,9

Certain viral infections during childhood, such as rotavirus and adenovirus infection, can increase the risk of celiac disease.^10–13 Although earlier studies reported that breast-feeding seemed to have a protective effect,¹⁴ as did introducing grains in the diet in the 4th to 6th months of life as opposed to earlier or later,¹⁵ more recent studies have not confirmed these benefits.^16,17

CLINICAL FEATURES

Most adults diagnosed with celiac disease are in their 30s, 40s, or 50s, and most are women.

Diarrhea remains a common presenting symptom, although the percentage of patients with celiac disease who present with diarrhea has decreased over time.^18,19

Abdominal pain and weight loss are also common.²⁰

Pallor or decreased exercise tolerance can develop due to anemia from iron malabsorption, and some patients have easy bruising due to vitamin K malabsorption.

Gynecologic and obstetric complications associated with celiac disease include delayed menarche, amenorrhea, spontaneous abortion, intrauterine growth retardation, preterm delivery, and low-birth-weight babies.^21,22 Patients who follow a gluten-free diet tend to have a lower incidence of intrauterine growth retardation, preterm delivery, and low-birth-weight babies compared with untreated patients.^21,22

Osteoporosis and osteopenia due to malabsorption of vitamin D are common and are seen in two-thirds of patients presenting with celiac disease.²³ A meta-analysis and position statement from Canada concluded that dual-energy x-ray absorptiometry should be done at the time of diagnosis of celiac disease if the patient is at risk of osteoporosis.²⁴ If the scan is abnormal, it should be repeated 1 to 2 years after initiation of a gluten-free diet and vitamin D supplementation to ensure that the osteopenia has improved.²⁴

OTHER DISEASE ASSOCIATIONS

Celiac disease is associated with various other autoimmune diseases (Table 1), including Hashimoto thyroiditis,²⁵ type 1 diabetes mellitus,²⁶ primary biliary cirrhosis,²⁷ primary sclerosing cholangitis,²⁸ and Addison disease.²⁹

Dermatitis herpetiformis

Dermatitis herpetiformis is one of the most common cutaneous manifestations of celiac disease. It presents between ages 10 and 50, and unlike celiac disease, it is more common in males.³⁰

Photo courtesy of Alok Vij, Department of Dermatology, Cleveland Clinic.

The characteristic lesions are pruritic, grouped erythematous papules surmounted by vesicles distributed symmetrically over the extensor surfaces of the upper and lower extremities, elbows, knees, scalp, nuchal area, and buttocks³¹ (Figures 2 and 3). In addition, some patients also present with vesicles, erythematous macules, and erosions in the oral mucosa³² or purpura on the palms and soles.^33–35

Photo courtesy of Alok Vij, MD, Department of Dermatology, Cleveland Clinic.

The pathogenesis of dermatitis herpetiformis in the skin is related to the pathogenesis of celiac disease in the gut. Like celiac disease, dermatitis herpetiformis is more common in genetically predisposed individuals carrying either the HLA-DQ2 or the HLA-DQ8 haplotype. In the skin, there is an analogue of tissue transglutaminase called epidermal transglutaminase, which helps in maintaining the integrity of cornified epithelium.³⁶ In patients with celiac disease, along with formation of IgA antibodies to tissue transglutaminase, there is also formation of IgA antibodies to epidermal transglutaminase. IgA antibodies are deposit- ed in the tips of dermal papillae and along the basement membrane.^37–39 These deposits then initiate an inflammatory response that is predominantly neutrophilic and results in formation of vesicles and bullae in the skin.⁴⁰ Also supporting the linkage between celiac disease and dermatitis herpetiformis, if patients adhere to a gluten-free diet, the deposits of immune complexes in the skin disappear.⁴¹

CELIAC DISEASE-ASSOCIATED MALIGNANCY

Patients with celiac disease have a higher risk of developing enteric malignancies, particularly intestinal T-cell lymphoma, and they have smaller increased risk of colon, oropharyngeal, esophageal, pancreatic, and hepatobiliary cancer.^42–45 For all of these cancers, the risk is higher than in the general public in the first year after celiac disease is diagnosed, but after the first year, the risk is increased only for small-bowel and hepatobiliary malignancies.⁴⁶

T-cell lymphoma

T-cell lymphoma is a rare but serious complication that has a poor prognosis.⁴⁷ Its prevalence has been increasing with time and is currently estimated to be around 0.01 to 0.02 per 100,000 people in the population as a whole.^48,49 The risk of developing lymphoma is 2.5 times higher in people with celiac disease than in the general population.⁵⁰ T-cell lymphoma is seen more commonly in patients with refractory celiac disease and DQ2 homozygosity.⁵¹

This disease is difficult to detect clinically, but sometimes it presents as an acute exacer­bation of celiac disease symptoms despite strict adherence to a gluten-free diet. Associated alarm symptoms include fever, night sweats, and laboratory abnormalities such as low albumin and high lactate dehydrogenase levels.

Strict adherence to a gluten-free diet remains the only way to prevent intestinal T-cell lymphoma.⁵²

Other malignancies

Some earlier studies reported an increased risk of thyroid cancer and malignant melanoma, but two newer studies have refuted this finding.^53,54 Conversely, celiac disease appears to have a protective effect against breast, ovarian, and endometrial cancers.⁵⁵

DIAGNOSIS: SEROLOGY, BIOPSY, GENETIC TESTING

Serologic tests

Patients strongly suspected of having celiac disease should be screened for IgA antibodies to tissue transglutaminase while on a gluten-containing diet, according to recommendations of the American College of Gastroenterology (Figure 4).⁵⁶ The sensitivity and specificity of this test are around 95%. If the patient has an IgA deficiency, screening should be done by checking the level of IgG antibodies to tissue transglutaminase.

Biopsy for confirmation

If testing for IgA to tissue transglutaminase is positive, upper endoscopy with biopsy is needed. Ideally, one to two samples should be taken from the duodenal bulb and at least four samples from the rest of the duodenum, preferably from two different locations.⁵⁶

Celiac disease has a broad spectrum of pathologic expressions, from mild distortion of crypt architecture to total villous atrophy and infiltration of lamina propria by lymphocytes⁵⁷ (Figures 5 and 6). Because these changes can be seen in a variety of diarrheal diseases, their reversal after adherence to a gluten-free diet is part of the current diagnostic criteria for the diagnosis of celiac disease.⁵⁶

Genetic testing

Photomicrograph courtesy of Homer Wiland MD, Department of Pathology, Cleveland Clinic.

Although the combination of positive serologic tests and pathologic changes confirms the diagnosis of celiac disease, in some cases one type of test is positive and the other is negative. In this situation, genetic testing for HLA-DQ2 and HLA-DQ8 can help rule out the diagnosis, as a negative genetic test rules out celiac disease in more than 99% of cases.⁵⁸

Genetic testing is also useful in patients who are already adhering to a gluten-free diet at the time of presentation to the clinic and who have had no testing done for celiac disease in the past. Here again, a negative test for both HLA-DQ2 and HLA-DQ8 makes a diagnosis of celiac disease highly unlikely.

If the test is positive, further testing needs to be done, as a positive genetic test cannot differentiate celiac disease from nonceliac gluten sensitivity. In this case, a gluten challenge needs to be done, ideally for 8 weeks, but for at least 2 weeks if the patient cannot tolerate gluten-containing food for a longer period of time. The gluten challenge is to be followed by testing for antibodies to tissue transglutaminase or obtaining duodenal biopsies to confirm the presence or absence of celiac disease.

Standard laboratory tests

Standard laboratory tests do not help much in diagnosing celiac disease, but they should include a complete blood chemistry along with a complete metabolic panel. Usually, serum albumin levels are normal.

Due to malabsorption of iron, patients may have iron deficiency anemia,⁵⁹ but anemia can also be due to a deficiency of folate or vitamin B₁₂. In patients undergoing endoscopic evaluation of iron deficiency anemia of unknown cause, celiac disease was discovered in approximately 15%.⁶⁰ Therefore, some experts believe that any patient presenting with unexplained iron deficiency anemia should be screened for celiac disease.

Because of malabsorption of vitamin D, levels of vitamin D can be low.

Elevations in levels of aminotransferases are also fairly common and usually resolve after the start of a gluten-free diet. If they persist despite adherence to a gluten-free diet, then an alternate cause of liver disease should be sought.⁶¹

Diagnosis of dermatitis herpetiformis

When trying to diagnose dermatitis herpetiformis, antibodies against epidermal transglutaminase can also be checked if testing for antibody against tissue transglutaminase is negative. A significant number of patients with biopsy-confirmed dermatitis herpetiformis are positive for epidermal transglutaminase antibodies but not for tissue transglutaminase antibodies.⁶²

The confirmatory test for dermatitis herpetiformis remains skin biopsy. Ideally, the sample should be taken while the patient is on a gluten-containing diet and from an area of normal-appearing skin around the lesions.⁶³ On histopathologic study, neutrophilic infiltrates are seen in dermal papillae and a perivascular lymphocytic infiltrate can also be seen in the superficial zones.⁶⁴ This presentation can also be seen in other bullous disorders, however. To differentiate dermatitis herpetiformis from other disorders, direct immunofluorescence is needed, which will detect granular IgA deposits in the dermal papillae or along the basement membrane, a finding pathognomic of dermatitis herpetiformis.⁶³

A GLUTEN-FREE DIET IS THE MAINSTAY OF TREATMENT

The mainstay of treatment is lifelong adherence to a gluten-free diet. Most patients report improvement in abdominal pain within days of starting this diet and improvement of diarrhea within 4 weeks.⁶⁵

The maximum amount of gluten that can be tolerated is debatable. A study established that intake of less than 10 mg a day is associated with fewer histologic abnormalities,⁶⁶ and an earlier study noted that intake of less than 50 mg a day was clinically well tolerated.⁶⁷ But patients differ in their tolerance for gluten, and it is hard to predict what the threshold of tolerance for gluten will be for a particular individual. Thus, it is better to avoid gluten completely.

Gluten-free if it is inherently gluten-free. If the food has a gluten-containing grain, then it should be processed to remove the gluten, and the resultant food product should not contain more than 20 parts per million of gluten. Gluten-free products that have gluten-containing grain that has been processed usually have a label indicating the gluten content in the food in parts per million.

Patients who understand the need to adhere to a gluten-free diet and the implications of not adhering to it are generally more compliant. Thus, patients need to be strongly educated that they need to adhere to a gluten-free diet and that nonadherence can cause further damage to the gut and can pose a higher risk of malignancy. Even though patients are usually concerned about the cost of gluten-free food and worry about adherence to the diet, these factors do not generally limit diet adherence.⁶⁸ All patients diagnosed with celiac disease should meet with a registered dietitian to discuss diet options based on their food preferences and to better address all their concerns.

With increasing awareness of celiac disease and with increasing numbers of patients being diagnosed with it, the food industry has recognized the need to produce gluten-free items. There are now plenty of food products available for these patients, who no longer have to forgo cakes, cookies, and other such items. Table 2 lists some common foods that patients with celiac disease can consume.

Nutritional supplements for some

If anemia is due purely to iron deficiency, it may resolve after starting a gluten-free diet, and no additional supplementation may be needed. However, if it is due to a combination of iron plus folate or vitamin B₁₂ deficiency, then folate, vitamin B₁₂, or both should be given.

In addition, if the patient is found to have a deficiency of vitamin D, then a vitamin D supplement should be given.⁶⁹ At the time of diagnosis, all patients with celiac disease should be screened for deficiencies of vitamins A, B₁₂, D, E, and K, as well as copper, zinc, folic acid, and iron.

Follow-up at 3 to 6 months

A follow-up visit should be scheduled for 3 to 6 months after the diagnosis and after that on an annual basis, and many of the abnormal laboratory tests will need to be repeated.

If intestinal or extraintestinal symptoms or nutrient deficiencies persist, then the patient’s adherence to the gluten-free diet needs to be checked. Adherence to a gluten-free diet can be assessed by checking for serologic markers of celiac disease. A decrease in baseline values can be seen within a few months of starting the diet.⁷⁰ Failure of serologic markers to decrease by the end of 1 year of a gluten-free diet usually indicates gluten contamination.⁷¹ If adherence is confirmed (ie, if baseline values fall) but symptoms persist, then further workup needs to be done to find the cause of refractory disease.

Skin lesions should also respond to a gluten-free diet

The first and foremost therapy for the skin lesions in dermatitis herpetiformis is the same as that for the intestinal manifestations in celiac disease, ie, adherence to a gluten-free diet. Soon after patients begin a gluten-free diet, the itching around the skin lesions goes away, and over time, most patients have complete resolution of the skin manifestations.

Dapsone is also frequently used to treat dermatitis herpetiformis if there is an incomplete response to a gluten-free diet or as an adjunct to diet to treat the pruritus. Patients often have a good response to dapsone.⁷²

The recommended starting dosage is 100 to 200 mg a day, and a response is usually seen within a few days. If the symptoms do not improve, the dose can be increased. Once the lesions resolve, the dose can be tapered and patients may not require any further medication. In some cases, patients may need to be chronically maintained on the lowest dose possible, due to the side effects of the drug.³

Dapsone is associated with significant adverse effects. Methemoglobinemia is the most common and is seen particularly in dosages exceeding 200 mg a day. Hemolytic anemia, another common adverse effect, is seen with dosages of more than 100 mg a day. Patients with a deficiency of glucose-6-phosphate dehydrogenase (G6PD) are at increased risk of hemolysis, and screening for G6PD deficiency is usually done before starting dapsone. Other rare adverse effects of dapsone include agranulocytosis, peripheral neuropathy, psychosis,⁷³ pancreatitis, cholestatic jaundice, bullous and exfoliative dermatitis, Stevens-Johnson syndrome, toxic epidermal necrolysis, nephrotic syndrome, and renal papillary necrosis.

Besides testing for G6PD deficiency, a complete blood cell count, a reticulocyte count, a hepatic function panel, renal function tests, and urinalysis should be done before starting dapsone therapy and repeated while on therapy. The complete blood cell count and reticulocyte count should be checked weekly for the first month, twice a month for the next 2 months, and then once every 3 months. Liver and renal function tests are to be done once every 3 months.⁷⁴

NOVEL THERAPIES BEING TESTED

Research is under way for other treatments for celiac disease besides a gluten-free diet.

Larazotide (Alba Therapeutics, Baltimore, MD) is being tested in a randomized, placebo-controlled trial. Early results indicate that it is effective in controlling both gastrointestinal and nongastrointestinal symptoms of celiac disease, but it still has to undergo phase 3 clinical trials.

Sorghum is a grain commonly used in Asia and Africa. The gluten in sorghum is different from that in wheat and is not immunogenic. In a small case series in patients with known celiac disease, sorghum did not induce diarrhea or change in levels of antibodies to tissue transglutaminase.⁷⁵

Nonimmunogenic wheat that does not contain the immunogenic gluten is being developed.

Oral enzyme supplements called glutenases are being developed. Glutenases can cleave gluten, particularly the proline and glutamine residues that make gluten resistant to degradation by gastric, pancreatic, and intestinal brush border proteases. A phase 2 trial of one of these oral enzyme supplements showed that it appeared to attenuate mucosal injury in patients with biopsy-proven celiac disease.⁷⁶

These novel therapies look promising, but for now the best treatment is lifelong adherence to the gluten-free diet.

NONRESPONSIVE AND REFRACTORY CELIAC DISEASE

Celiac disease is considered nonresponsive if its symptoms or laboratory abnormalities persist after the patient is on a gluten-free diet for 6 to 12 months. It is considered refractory if symptoms persist or recur along with villous atrophy despite adherence to the diet for more than 12 months in the absence of other causes of the symptoms. Refractory celiac disease can be further classified either as type 1 if there are typical intraepithelial lymphocytes, or as type 2 if there are atypical intraepithelial lymphocytes.

Celiac disease is nonresponsive in about 10% to 19% of cases,⁷⁶ and it is refractory in 1% to 2%.⁷⁷

Managing nonresponsive celiac disease

The first step in managing a patient with nonresponsive celiac disease is to confirm the diagnosis by reviewing the serologic tests and the biopsy samples from the time of diagnosis. If celiac disease is confirmed, then one should re-evaluate for gluten ingestion, the most common cause of nonresponsiveness.⁷⁸ If strict adherence is confirmed, then check for other causes of symptoms such as lactose or fructose intolerance. If no other cause is found, then repeat the duodenal biopsies with flow cytometry to look for CD3 and CD8 expression in T cells in the small-bowel mucosa.⁷⁹ Presence or absence of villous atrophy can point to possible other causes of malabsorption including pancreatic insufficiency, small intestinal bowel overgrowth, and microscopic colitis.

Managing refractory celiac disease

Traditionally, corticosteroids have been shown to be beneficial in alleviating symptoms in patients with refractory celiac disease but do not improve the histologic findings.⁸⁰ Because of the adverse effects associated with long-term corticosteroid use, azathioprine has been successfully used to maintain remission of the disease after induction with corticosteroids in patients with type 1 refractory celiac disease.⁸¹

Cladribine, a chemotherapeutic agent used to treat hairy cell leukemia, has shown some benefit in treating type 2 refractory celiac disease.⁸²

In type 2 refractory celiac disease, use of an immunomodulator agent carries an increased risk of transformation to lymphoma.

Because of the lack of a satisfactory response to the agents available so far to treat refractory celiac disease, more treatment options acting at the molecular level are being explored.

NONCELIAC GLUTEN SENSITIVITY DISORDER

Nonceliac gluten sensitivity disorder is an evolving concept. The clinical presentation of this disorder is similar to celiac disease in that patients may have diarrhea or other extra­intestinal symptoms when on a regular diet and have resolution of symptoms on a gluten-free diet. But unlike celiac disease, there is no serologic or histologic evidence of celiac disease even when patients are on a regular diet.

One of every 17 patients who presents with clinical features suggestive of celiac disease is found to have nonceliac gluten sensitivity disorder, not celiac disease.⁸³ In contrast to celiac disease, in which the adaptive immune system is thought to contribute to the disease process, in nonceliac gluten sensitivity disorder the innate immune system is believed to play the dominant role,⁸⁴ but the exact pathogenesis of the disease is still unclear.

The diagnosis of nonceliac gluten sensitivity disorder is one of exclusion. Celiac disease needs to be ruled out by serologic testing and by duodenal biopsy while the patient is on a regular diet, and then a trial of a gluten-free diet needs to be done to confirm resolution of symptoms before the diagnosis of nonceliac gluten sensitivity disorder can be established.

As with celiac disease, the treatment involves adhering to a gluten-free diet, but it is still not known if patients need to stay on it for the rest of their life, or if they will be able to tolerate gluten-containing products after a few years.

Celiac disease is an autoimmune disorder that occurs in genetically predisposed individuals in response to ingestion of gluten. Its prevalence is about 0.7% of the US population.¹

See related editorial

The gold standard for diagnosis is duodenal biopsy, in which the histologic features may include varying gradations of flattening of intestinal villi, crypt hyperplasia, and infiltration of the lamina propria by lymphocytes. Many patients have no symptoms at the time of diagnosis, but presenting symptoms can include diarrhea along with features of malabsorption,² and, in about 25% of patients (mainly adults), a bullous cutaneous disorder called dermatitis herpetiformis.^3,4 The pathogenesis of celiac disease and that of dermatitis herpetiformis are similar in that in both, ingestion of gluten induces an inflammatory reaction leading to the clinical manifestations.

The mainstay of treatment of celiac disease remains avoidance of gluten in the diet.

GENETIC PREDISPOSITION AND DIETARY TRIGGER

The pathogenesis of celiac disease has been well studied in both humans and animals. The disease is thought to develop by an interplay of genetic and autoimmune factors and the ingestion of gluten (ie, an environmental factor).

Celiac disease occurs in genetically predisposed individuals, ie, those who carry the HLA alleles DQ2 (DQA1*05, DQB1*02), DQ8 (DQA1*03, DQB1*0302), or both.⁵

Ingestion of gluten is necessary for the disease to develop. Gluten, the protein component of wheat, barley, and rye, contains proteins called prolamins, which vary among the different types of grain. In wheat, the prolamin is gliadin, which is alcohol-soluble. In barley the prolamin is hordein, and in rye it is secalin.⁴ The prolamin content in gluten makes it resistant to degradation by gastric, pancreatic, and intestinal brush border proteases.⁶ Gluten crosses the epithelial barrier and promotes an inflammatory reaction by both the innate and adaptive immune systems that can ultimately result in flattening of villi and crypt hyperplasia (Figure 1).⁷

Tissue transglutaminase also plays a central role in the pathogenesis, as it further deaminates gliadin and increases its immunogenicity by causing it to bind to receptors on antigen-presenting cells with stronger affinity. Furthermore, gliadin-tissue transglutaminase complexes formed by protein cross-linkages generate an autoantibody response (predominantly immunoglobulin A [IgA] type) that can exacerbate the inflammatory process.^8,9

Certain viral infections during childhood, such as rotavirus and adenovirus infection, can increase the risk of celiac disease.^10–13 Although earlier studies reported that breast-feeding seemed to have a protective effect,¹⁴ as did introducing grains in the diet in the 4th to 6th months of life as opposed to earlier or later,¹⁵ more recent studies have not confirmed these benefits.^16,17

CLINICAL FEATURES

Most adults diagnosed with celiac disease are in their 30s, 40s, or 50s, and most are women.

Diarrhea remains a common presenting symptom, although the percentage of patients with celiac disease who present with diarrhea has decreased over time.^18,19

Abdominal pain and weight loss are also common.²⁰

Pallor or decreased exercise tolerance can develop due to anemia from iron malabsorption, and some patients have easy bruising due to vitamin K malabsorption.

Gynecologic and obstetric complications associated with celiac disease include delayed menarche, amenorrhea, spontaneous abortion, intrauterine growth retardation, preterm delivery, and low-birth-weight babies.^21,22 Patients who follow a gluten-free diet tend to have a lower incidence of intrauterine growth retardation, preterm delivery, and low-birth-weight babies compared with untreated patients.^21,22

Osteoporosis and osteopenia due to malabsorption of vitamin D are common and are seen in two-thirds of patients presenting with celiac disease.²³ A meta-analysis and position statement from Canada concluded that dual-energy x-ray absorptiometry should be done at the time of diagnosis of celiac disease if the patient is at risk of osteoporosis.²⁴ If the scan is abnormal, it should be repeated 1 to 2 years after initiation of a gluten-free diet and vitamin D supplementation to ensure that the osteopenia has improved.²⁴

OTHER DISEASE ASSOCIATIONS

Celiac disease is associated with various other autoimmune diseases (Table 1), including Hashimoto thyroiditis,²⁵ type 1 diabetes mellitus,²⁶ primary biliary cirrhosis,²⁷ primary sclerosing cholangitis,²⁸ and Addison disease.²⁹

Dermatitis herpetiformis

Dermatitis herpetiformis is one of the most common cutaneous manifestations of celiac disease. It presents between ages 10 and 50, and unlike celiac disease, it is more common in males.³⁰

Photo courtesy of Alok Vij, Department of Dermatology, Cleveland Clinic.

The characteristic lesions are pruritic, grouped erythematous papules surmounted by vesicles distributed symmetrically over the extensor surfaces of the upper and lower extremities, elbows, knees, scalp, nuchal area, and buttocks³¹ (Figures 2 and 3). In addition, some patients also present with vesicles, erythematous macules, and erosions in the oral mucosa³² or purpura on the palms and soles.^33–35

Photo courtesy of Alok Vij, MD, Department of Dermatology, Cleveland Clinic.

The pathogenesis of dermatitis herpetiformis in the skin is related to the pathogenesis of celiac disease in the gut. Like celiac disease, dermatitis herpetiformis is more common in genetically predisposed individuals carrying either the HLA-DQ2 or the HLA-DQ8 haplotype. In the skin, there is an analogue of tissue transglutaminase called epidermal transglutaminase, which helps in maintaining the integrity of cornified epithelium.³⁶ In patients with celiac disease, along with formation of IgA antibodies to tissue transglutaminase, there is also formation of IgA antibodies to epidermal transglutaminase. IgA antibodies are deposit- ed in the tips of dermal papillae and along the basement membrane.^37–39 These deposits then initiate an inflammatory response that is predominantly neutrophilic and results in formation of vesicles and bullae in the skin.⁴⁰ Also supporting the linkage between celiac disease and dermatitis herpetiformis, if patients adhere to a gluten-free diet, the deposits of immune complexes in the skin disappear.⁴¹

CELIAC DISEASE-ASSOCIATED MALIGNANCY

Patients with celiac disease have a higher risk of developing enteric malignancies, particularly intestinal T-cell lymphoma, and they have smaller increased risk of colon, oropharyngeal, esophageal, pancreatic, and hepatobiliary cancer.^42–45 For all of these cancers, the risk is higher than in the general public in the first year after celiac disease is diagnosed, but after the first year, the risk is increased only for small-bowel and hepatobiliary malignancies.⁴⁶

T-cell lymphoma

T-cell lymphoma is a rare but serious complication that has a poor prognosis.⁴⁷ Its prevalence has been increasing with time and is currently estimated to be around 0.01 to 0.02 per 100,000 people in the population as a whole.^48,49 The risk of developing lymphoma is 2.5 times higher in people with celiac disease than in the general population.⁵⁰ T-cell lymphoma is seen more commonly in patients with refractory celiac disease and DQ2 homozygosity.⁵¹

This disease is difficult to detect clinically, but sometimes it presents as an acute exacer­bation of celiac disease symptoms despite strict adherence to a gluten-free diet. Associated alarm symptoms include fever, night sweats, and laboratory abnormalities such as low albumin and high lactate dehydrogenase levels.

Strict adherence to a gluten-free diet remains the only way to prevent intestinal T-cell lymphoma.⁵²

Other malignancies

Some earlier studies reported an increased risk of thyroid cancer and malignant melanoma, but two newer studies have refuted this finding.^53,54 Conversely, celiac disease appears to have a protective effect against breast, ovarian, and endometrial cancers.⁵⁵

DIAGNOSIS: SEROLOGY, BIOPSY, GENETIC TESTING

Serologic tests

Patients strongly suspected of having celiac disease should be screened for IgA antibodies to tissue transglutaminase while on a gluten-containing diet, according to recommendations of the American College of Gastroenterology (Figure 4).⁵⁶ The sensitivity and specificity of this test are around 95%. If the patient has an IgA deficiency, screening should be done by checking the level of IgG antibodies to tissue transglutaminase.

Biopsy for confirmation

If testing for IgA to tissue transglutaminase is positive, upper endoscopy with biopsy is needed. Ideally, one to two samples should be taken from the duodenal bulb and at least four samples from the rest of the duodenum, preferably from two different locations.⁵⁶

Celiac disease has a broad spectrum of pathologic expressions, from mild distortion of crypt architecture to total villous atrophy and infiltration of lamina propria by lymphocytes⁵⁷ (Figures 5 and 6). Because these changes can be seen in a variety of diarrheal diseases, their reversal after adherence to a gluten-free diet is part of the current diagnostic criteria for the diagnosis of celiac disease.⁵⁶

Genetic testing

Photomicrograph courtesy of Homer Wiland MD, Department of Pathology, Cleveland Clinic.

Although the combination of positive serologic tests and pathologic changes confirms the diagnosis of celiac disease, in some cases one type of test is positive and the other is negative. In this situation, genetic testing for HLA-DQ2 and HLA-DQ8 can help rule out the diagnosis, as a negative genetic test rules out celiac disease in more than 99% of cases.⁵⁸

Genetic testing is also useful in patients who are already adhering to a gluten-free diet at the time of presentation to the clinic and who have had no testing done for celiac disease in the past. Here again, a negative test for both HLA-DQ2 and HLA-DQ8 makes a diagnosis of celiac disease highly unlikely.

If the test is positive, further testing needs to be done, as a positive genetic test cannot differentiate celiac disease from nonceliac gluten sensitivity. In this case, a gluten challenge needs to be done, ideally for 8 weeks, but for at least 2 weeks if the patient cannot tolerate gluten-containing food for a longer period of time. The gluten challenge is to be followed by testing for antibodies to tissue transglutaminase or obtaining duodenal biopsies to confirm the presence or absence of celiac disease.

Standard laboratory tests

Standard laboratory tests do not help much in diagnosing celiac disease, but they should include a complete blood chemistry along with a complete metabolic panel. Usually, serum albumin levels are normal.

Due to malabsorption of iron, patients may have iron deficiency anemia,⁵⁹ but anemia can also be due to a deficiency of folate or vitamin B₁₂. In patients undergoing endoscopic evaluation of iron deficiency anemia of unknown cause, celiac disease was discovered in approximately 15%.⁶⁰ Therefore, some experts believe that any patient presenting with unexplained iron deficiency anemia should be screened for celiac disease.

Because of malabsorption of vitamin D, levels of vitamin D can be low.

Elevations in levels of aminotransferases are also fairly common and usually resolve after the start of a gluten-free diet. If they persist despite adherence to a gluten-free diet, then an alternate cause of liver disease should be sought.⁶¹

Diagnosis of dermatitis herpetiformis

When trying to diagnose dermatitis herpetiformis, antibodies against epidermal transglutaminase can also be checked if testing for antibody against tissue transglutaminase is negative. A significant number of patients with biopsy-confirmed dermatitis herpetiformis are positive for epidermal transglutaminase antibodies but not for tissue transglutaminase antibodies.⁶²

The confirmatory test for dermatitis herpetiformis remains skin biopsy. Ideally, the sample should be taken while the patient is on a gluten-containing diet and from an area of normal-appearing skin around the lesions.⁶³ On histopathologic study, neutrophilic infiltrates are seen in dermal papillae and a perivascular lymphocytic infiltrate can also be seen in the superficial zones.⁶⁴ This presentation can also be seen in other bullous disorders, however. To differentiate dermatitis herpetiformis from other disorders, direct immunofluorescence is needed, which will detect granular IgA deposits in the dermal papillae or along the basement membrane, a finding pathognomic of dermatitis herpetiformis.⁶³

A GLUTEN-FREE DIET IS THE MAINSTAY OF TREATMENT

The mainstay of treatment is lifelong adherence to a gluten-free diet. Most patients report improvement in abdominal pain within days of starting this diet and improvement of diarrhea within 4 weeks.⁶⁵

The maximum amount of gluten that can be tolerated is debatable. A study established that intake of less than 10 mg a day is associated with fewer histologic abnormalities,⁶⁶ and an earlier study noted that intake of less than 50 mg a day was clinically well tolerated.⁶⁷ But patients differ in their tolerance for gluten, and it is hard to predict what the threshold of tolerance for gluten will be for a particular individual. Thus, it is better to avoid gluten completely.

Gluten-free if it is inherently gluten-free. If the food has a gluten-containing grain, then it should be processed to remove the gluten, and the resultant food product should not contain more than 20 parts per million of gluten. Gluten-free products that have gluten-containing grain that has been processed usually have a label indicating the gluten content in the food in parts per million.

Patients who understand the need to adhere to a gluten-free diet and the implications of not adhering to it are generally more compliant. Thus, patients need to be strongly educated that they need to adhere to a gluten-free diet and that nonadherence can cause further damage to the gut and can pose a higher risk of malignancy. Even though patients are usually concerned about the cost of gluten-free food and worry about adherence to the diet, these factors do not generally limit diet adherence.⁶⁸ All patients diagnosed with celiac disease should meet with a registered dietitian to discuss diet options based on their food preferences and to better address all their concerns.

With increasing awareness of celiac disease and with increasing numbers of patients being diagnosed with it, the food industry has recognized the need to produce gluten-free items. There are now plenty of food products available for these patients, who no longer have to forgo cakes, cookies, and other such items. Table 2 lists some common foods that patients with celiac disease can consume.

Nutritional supplements for some

If anemia is due purely to iron deficiency, it may resolve after starting a gluten-free diet, and no additional supplementation may be needed. However, if it is due to a combination of iron plus folate or vitamin B₁₂ deficiency, then folate, vitamin B₁₂, or both should be given.

In addition, if the patient is found to have a deficiency of vitamin D, then a vitamin D supplement should be given.⁶⁹ At the time of diagnosis, all patients with celiac disease should be screened for deficiencies of vitamins A, B₁₂, D, E, and K, as well as copper, zinc, folic acid, and iron.

Follow-up at 3 to 6 months

A follow-up visit should be scheduled for 3 to 6 months after the diagnosis and after that on an annual basis, and many of the abnormal laboratory tests will need to be repeated.

If intestinal or extraintestinal symptoms or nutrient deficiencies persist, then the patient’s adherence to the gluten-free diet needs to be checked. Adherence to a gluten-free diet can be assessed by checking for serologic markers of celiac disease. A decrease in baseline values can be seen within a few months of starting the diet.⁷⁰ Failure of serologic markers to decrease by the end of 1 year of a gluten-free diet usually indicates gluten contamination.⁷¹ If adherence is confirmed (ie, if baseline values fall) but symptoms persist, then further workup needs to be done to find the cause of refractory disease.

Skin lesions should also respond to a gluten-free diet

The first and foremost therapy for the skin lesions in dermatitis herpetiformis is the same as that for the intestinal manifestations in celiac disease, ie, adherence to a gluten-free diet. Soon after patients begin a gluten-free diet, the itching around the skin lesions goes away, and over time, most patients have complete resolution of the skin manifestations.

Dapsone is also frequently used to treat dermatitis herpetiformis if there is an incomplete response to a gluten-free diet or as an adjunct to diet to treat the pruritus. Patients often have a good response to dapsone.⁷²

The recommended starting dosage is 100 to 200 mg a day, and a response is usually seen within a few days. If the symptoms do not improve, the dose can be increased. Once the lesions resolve, the dose can be tapered and patients may not require any further medication. In some cases, patients may need to be chronically maintained on the lowest dose possible, due to the side effects of the drug.³

Dapsone is associated with significant adverse effects. Methemoglobinemia is the most common and is seen particularly in dosages exceeding 200 mg a day. Hemolytic anemia, another common adverse effect, is seen with dosages of more than 100 mg a day. Patients with a deficiency of glucose-6-phosphate dehydrogenase (G6PD) are at increased risk of hemolysis, and screening for G6PD deficiency is usually done before starting dapsone. Other rare adverse effects of dapsone include agranulocytosis, peripheral neuropathy, psychosis,⁷³ pancreatitis, cholestatic jaundice, bullous and exfoliative dermatitis, Stevens-Johnson syndrome, toxic epidermal necrolysis, nephrotic syndrome, and renal papillary necrosis.

Besides testing for G6PD deficiency, a complete blood cell count, a reticulocyte count, a hepatic function panel, renal function tests, and urinalysis should be done before starting dapsone therapy and repeated while on therapy. The complete blood cell count and reticulocyte count should be checked weekly for the first month, twice a month for the next 2 months, and then once every 3 months. Liver and renal function tests are to be done once every 3 months.⁷⁴

NOVEL THERAPIES BEING TESTED

Research is under way for other treatments for celiac disease besides a gluten-free diet.

Larazotide (Alba Therapeutics, Baltimore, MD) is being tested in a randomized, placebo-controlled trial. Early results indicate that it is effective in controlling both gastrointestinal and nongastrointestinal symptoms of celiac disease, but it still has to undergo phase 3 clinical trials.

Sorghum is a grain commonly used in Asia and Africa. The gluten in sorghum is different from that in wheat and is not immunogenic. In a small case series in patients with known celiac disease, sorghum did not induce diarrhea or change in levels of antibodies to tissue transglutaminase.⁷⁵

Nonimmunogenic wheat that does not contain the immunogenic gluten is being developed.

Oral enzyme supplements called glutenases are being developed. Glutenases can cleave gluten, particularly the proline and glutamine residues that make gluten resistant to degradation by gastric, pancreatic, and intestinal brush border proteases. A phase 2 trial of one of these oral enzyme supplements showed that it appeared to attenuate mucosal injury in patients with biopsy-proven celiac disease.⁷⁶

These novel therapies look promising, but for now the best treatment is lifelong adherence to the gluten-free diet.

NONRESPONSIVE AND REFRACTORY CELIAC DISEASE

Celiac disease is considered nonresponsive if its symptoms or laboratory abnormalities persist after the patient is on a gluten-free diet for 6 to 12 months. It is considered refractory if symptoms persist or recur along with villous atrophy despite adherence to the diet for more than 12 months in the absence of other causes of the symptoms. Refractory celiac disease can be further classified either as type 1 if there are typical intraepithelial lymphocytes, or as type 2 if there are atypical intraepithelial lymphocytes.

Celiac disease is nonresponsive in about 10% to 19% of cases,⁷⁶ and it is refractory in 1% to 2%.⁷⁷

Managing nonresponsive celiac disease

The first step in managing a patient with nonresponsive celiac disease is to confirm the diagnosis by reviewing the serologic tests and the biopsy samples from the time of diagnosis. If celiac disease is confirmed, then one should re-evaluate for gluten ingestion, the most common cause of nonresponsiveness.⁷⁸ If strict adherence is confirmed, then check for other causes of symptoms such as lactose or fructose intolerance. If no other cause is found, then repeat the duodenal biopsies with flow cytometry to look for CD3 and CD8 expression in T cells in the small-bowel mucosa.⁷⁹ Presence or absence of villous atrophy can point to possible other causes of malabsorption including pancreatic insufficiency, small intestinal bowel overgrowth, and microscopic colitis.

Managing refractory celiac disease

Traditionally, corticosteroids have been shown to be beneficial in alleviating symptoms in patients with refractory celiac disease but do not improve the histologic findings.⁸⁰ Because of the adverse effects associated with long-term corticosteroid use, azathioprine has been successfully used to maintain remission of the disease after induction with corticosteroids in patients with type 1 refractory celiac disease.⁸¹

Cladribine, a chemotherapeutic agent used to treat hairy cell leukemia, has shown some benefit in treating type 2 refractory celiac disease.⁸²

In type 2 refractory celiac disease, use of an immunomodulator agent carries an increased risk of transformation to lymphoma.

Because of the lack of a satisfactory response to the agents available so far to treat refractory celiac disease, more treatment options acting at the molecular level are being explored.

NONCELIAC GLUTEN SENSITIVITY DISORDER

Nonceliac gluten sensitivity disorder is an evolving concept. The clinical presentation of this disorder is similar to celiac disease in that patients may have diarrhea or other extra­intestinal symptoms when on a regular diet and have resolution of symptoms on a gluten-free diet. But unlike celiac disease, there is no serologic or histologic evidence of celiac disease even when patients are on a regular diet.

One of every 17 patients who presents with clinical features suggestive of celiac disease is found to have nonceliac gluten sensitivity disorder, not celiac disease.⁸³ In contrast to celiac disease, in which the adaptive immune system is thought to contribute to the disease process, in nonceliac gluten sensitivity disorder the innate immune system is believed to play the dominant role,⁸⁴ but the exact pathogenesis of the disease is still unclear.

The diagnosis of nonceliac gluten sensitivity disorder is one of exclusion. Celiac disease needs to be ruled out by serologic testing and by duodenal biopsy while the patient is on a regular diet, and then a trial of a gluten-free diet needs to be done to confirm resolution of symptoms before the diagnosis of nonceliac gluten sensitivity disorder can be established.

As with celiac disease, the treatment involves adhering to a gluten-free diet, but it is still not known if patients need to stay on it for the rest of their life, or if they will be able to tolerate gluten-containing products after a few years.

Celiac disease is an autoimmune disorder that occurs in genetically predisposed individuals in response to ingestion of gluten. Its prevalence is about 0.7% of the US population.¹

See related editorial

The gold standard for diagnosis is duodenal biopsy, in which the histologic features may include varying gradations of flattening of intestinal villi, crypt hyperplasia, and infiltration of the lamina propria by lymphocytes. Many patients have no symptoms at the time of diagnosis, but presenting symptoms can include diarrhea along with features of malabsorption,² and, in about 25% of patients (mainly adults), a bullous cutaneous disorder called dermatitis herpetiformis.^3,4 The pathogenesis of celiac disease and that of dermatitis herpetiformis are similar in that in both, ingestion of gluten induces an inflammatory reaction leading to the clinical manifestations.

The mainstay of treatment of celiac disease remains avoidance of gluten in the diet.

GENETIC PREDISPOSITION AND DIETARY TRIGGER

The pathogenesis of celiac disease has been well studied in both humans and animals. The disease is thought to develop by an interplay of genetic and autoimmune factors and the ingestion of gluten (ie, an environmental factor).

Celiac disease occurs in genetically predisposed individuals, ie, those who carry the HLA alleles DQ2 (DQA1*05, DQB1*02), DQ8 (DQA1*03, DQB1*0302), or both.⁵

Ingestion of gluten is necessary for the disease to develop. Gluten, the protein component of wheat, barley, and rye, contains proteins called prolamins, which vary among the different types of grain. In wheat, the prolamin is gliadin, which is alcohol-soluble. In barley the prolamin is hordein, and in rye it is secalin.⁴ The prolamin content in gluten makes it resistant to degradation by gastric, pancreatic, and intestinal brush border proteases.⁶ Gluten crosses the epithelial barrier and promotes an inflammatory reaction by both the innate and adaptive immune systems that can ultimately result in flattening of villi and crypt hyperplasia (Figure 1).⁷

Tissue transglutaminase also plays a central role in the pathogenesis, as it further deaminates gliadin and increases its immunogenicity by causing it to bind to receptors on antigen-presenting cells with stronger affinity. Furthermore, gliadin-tissue transglutaminase complexes formed by protein cross-linkages generate an autoantibody response (predominantly immunoglobulin A [IgA] type) that can exacerbate the inflammatory process.^8,9

Certain viral infections during childhood, such as rotavirus and adenovirus infection, can increase the risk of celiac disease.^10–13 Although earlier studies reported that breast-feeding seemed to have a protective effect,¹⁴ as did introducing grains in the diet in the 4th to 6th months of life as opposed to earlier or later,¹⁵ more recent studies have not confirmed these benefits.^16,17

CLINICAL FEATURES

Most adults diagnosed with celiac disease are in their 30s, 40s, or 50s, and most are women.

Diarrhea remains a common presenting symptom, although the percentage of patients with celiac disease who present with diarrhea has decreased over time.^18,19

Abdominal pain and weight loss are also common.²⁰

Pallor or decreased exercise tolerance can develop due to anemia from iron malabsorption, and some patients have easy bruising due to vitamin K malabsorption.

Gynecologic and obstetric complications associated with celiac disease include delayed menarche, amenorrhea, spontaneous abortion, intrauterine growth retardation, preterm delivery, and low-birth-weight babies.^21,22 Patients who follow a gluten-free diet tend to have a lower incidence of intrauterine growth retardation, preterm delivery, and low-birth-weight babies compared with untreated patients.^21,22

Osteoporosis and osteopenia due to malabsorption of vitamin D are common and are seen in two-thirds of patients presenting with celiac disease.²³ A meta-analysis and position statement from Canada concluded that dual-energy x-ray absorptiometry should be done at the time of diagnosis of celiac disease if the patient is at risk of osteoporosis.²⁴ If the scan is abnormal, it should be repeated 1 to 2 years after initiation of a gluten-free diet and vitamin D supplementation to ensure that the osteopenia has improved.²⁴

OTHER DISEASE ASSOCIATIONS

Celiac disease is associated with various other autoimmune diseases (Table 1), including Hashimoto thyroiditis,²⁵ type 1 diabetes mellitus,²⁶ primary biliary cirrhosis,²⁷ primary sclerosing cholangitis,²⁸ and Addison disease.²⁹

Dermatitis herpetiformis

Dermatitis herpetiformis is one of the most common cutaneous manifestations of celiac disease. It presents between ages 10 and 50, and unlike celiac disease, it is more common in males.³⁰

Photo courtesy of Alok Vij, Department of Dermatology, Cleveland Clinic.

The characteristic lesions are pruritic, grouped erythematous papules surmounted by vesicles distributed symmetrically over the extensor surfaces of the upper and lower extremities, elbows, knees, scalp, nuchal area, and buttocks³¹ (Figures 2 and 3). In addition, some patients also present with vesicles, erythematous macules, and erosions in the oral mucosa³² or purpura on the palms and soles.^33–35

Photo courtesy of Alok Vij, MD, Department of Dermatology, Cleveland Clinic.

The pathogenesis of dermatitis herpetiformis in the skin is related to the pathogenesis of celiac disease in the gut. Like celiac disease, dermatitis herpetiformis is more common in genetically predisposed individuals carrying either the HLA-DQ2 or the HLA-DQ8 haplotype. In the skin, there is an analogue of tissue transglutaminase called epidermal transglutaminase, which helps in maintaining the integrity of cornified epithelium.³⁶ In patients with celiac disease, along with formation of IgA antibodies to tissue transglutaminase, there is also formation of IgA antibodies to epidermal transglutaminase. IgA antibodies are deposit- ed in the tips of dermal papillae and along the basement membrane.^37–39 These deposits then initiate an inflammatory response that is predominantly neutrophilic and results in formation of vesicles and bullae in the skin.⁴⁰ Also supporting the linkage between celiac disease and dermatitis herpetiformis, if patients adhere to a gluten-free diet, the deposits of immune complexes in the skin disappear.⁴¹

CELIAC DISEASE-ASSOCIATED MALIGNANCY

Patients with celiac disease have a higher risk of developing enteric malignancies, particularly intestinal T-cell lymphoma, and they have smaller increased risk of colon, oropharyngeal, esophageal, pancreatic, and hepatobiliary cancer.^42–45 For all of these cancers, the risk is higher than in the general public in the first year after celiac disease is diagnosed, but after the first year, the risk is increased only for small-bowel and hepatobiliary malignancies.⁴⁶

T-cell lymphoma

T-cell lymphoma is a rare but serious complication that has a poor prognosis.⁴⁷ Its prevalence has been increasing with time and is currently estimated to be around 0.01 to 0.02 per 100,000 people in the population as a whole.^48,49 The risk of developing lymphoma is 2.5 times higher in people with celiac disease than in the general population.⁵⁰ T-cell lymphoma is seen more commonly in patients with refractory celiac disease and DQ2 homozygosity.⁵¹

This disease is difficult to detect clinically, but sometimes it presents as an acute exacer­bation of celiac disease symptoms despite strict adherence to a gluten-free diet. Associated alarm symptoms include fever, night sweats, and laboratory abnormalities such as low albumin and high lactate dehydrogenase levels.

Strict adherence to a gluten-free diet remains the only way to prevent intestinal T-cell lymphoma.⁵²

Other malignancies

Some earlier studies reported an increased risk of thyroid cancer and malignant melanoma, but two newer studies have refuted this finding.^53,54 Conversely, celiac disease appears to have a protective effect against breast, ovarian, and endometrial cancers.⁵⁵

DIAGNOSIS: SEROLOGY, BIOPSY, GENETIC TESTING

Serologic tests

Patients strongly suspected of having celiac disease should be screened for IgA antibodies to tissue transglutaminase while on a gluten-containing diet, according to recommendations of the American College of Gastroenterology (Figure 4).⁵⁶ The sensitivity and specificity of this test are around 95%. If the patient has an IgA deficiency, screening should be done by checking the level of IgG antibodies to tissue transglutaminase.

Biopsy for confirmation

If testing for IgA to tissue transglutaminase is positive, upper endoscopy with biopsy is needed. Ideally, one to two samples should be taken from the duodenal bulb and at least four samples from the rest of the duodenum, preferably from two different locations.⁵⁶

Celiac disease has a broad spectrum of pathologic expressions, from mild distortion of crypt architecture to total villous atrophy and infiltration of lamina propria by lymphocytes⁵⁷ (Figures 5 and 6). Because these changes can be seen in a variety of diarrheal diseases, their reversal after adherence to a gluten-free diet is part of the current diagnostic criteria for the diagnosis of celiac disease.⁵⁶

Genetic testing

Photomicrograph courtesy of Homer Wiland MD, Department of Pathology, Cleveland Clinic.

Although the combination of positive serologic tests and pathologic changes confirms the diagnosis of celiac disease, in some cases one type of test is positive and the other is negative. In this situation, genetic testing for HLA-DQ2 and HLA-DQ8 can help rule out the diagnosis, as a negative genetic test rules out celiac disease in more than 99% of cases.⁵⁸

Genetic testing is also useful in patients who are already adhering to a gluten-free diet at the time of presentation to the clinic and who have had no testing done for celiac disease in the past. Here again, a negative test for both HLA-DQ2 and HLA-DQ8 makes a diagnosis of celiac disease highly unlikely.

If the test is positive, further testing needs to be done, as a positive genetic test cannot differentiate celiac disease from nonceliac gluten sensitivity. In this case, a gluten challenge needs to be done, ideally for 8 weeks, but for at least 2 weeks if the patient cannot tolerate gluten-containing food for a longer period of time. The gluten challenge is to be followed by testing for antibodies to tissue transglutaminase or obtaining duodenal biopsies to confirm the presence or absence of celiac disease.

Standard laboratory tests

Standard laboratory tests do not help much in diagnosing celiac disease, but they should include a complete blood chemistry along with a complete metabolic panel. Usually, serum albumin levels are normal.

Due to malabsorption of iron, patients may have iron deficiency anemia,⁵⁹ but anemia can also be due to a deficiency of folate or vitamin B₁₂. In patients undergoing endoscopic evaluation of iron deficiency anemia of unknown cause, celiac disease was discovered in approximately 15%.⁶⁰ Therefore, some experts believe that any patient presenting with unexplained iron deficiency anemia should be screened for celiac disease.

Because of malabsorption of vitamin D, levels of vitamin D can be low.

Elevations in levels of aminotransferases are also fairly common and usually resolve after the start of a gluten-free diet. If they persist despite adherence to a gluten-free diet, then an alternate cause of liver disease should be sought.⁶¹

Diagnosis of dermatitis herpetiformis

When trying to diagnose dermatitis herpetiformis, antibodies against epidermal transglutaminase can also be checked if testing for antibody against tissue transglutaminase is negative. A significant number of patients with biopsy-confirmed dermatitis herpetiformis are positive for epidermal transglutaminase antibodies but not for tissue transglutaminase antibodies.⁶²

The confirmatory test for dermatitis herpetiformis remains skin biopsy. Ideally, the sample should be taken while the patient is on a gluten-containing diet and from an area of normal-appearing skin around the lesions.⁶³ On histopathologic study, neutrophilic infiltrates are seen in dermal papillae and a perivascular lymphocytic infiltrate can also be seen in the superficial zones.⁶⁴ This presentation can also be seen in other bullous disorders, however. To differentiate dermatitis herpetiformis from other disorders, direct immunofluorescence is needed, which will detect granular IgA deposits in the dermal papillae or along the basement membrane, a finding pathognomic of dermatitis herpetiformis.⁶³

A GLUTEN-FREE DIET IS THE MAINSTAY OF TREATMENT

The mainstay of treatment is lifelong adherence to a gluten-free diet. Most patients report improvement in abdominal pain within days of starting this diet and improvement of diarrhea within 4 weeks.⁶⁵

The maximum amount of gluten that can be tolerated is debatable. A study established that intake of less than 10 mg a day is associated with fewer histologic abnormalities,⁶⁶ and an earlier study noted that intake of less than 50 mg a day was clinically well tolerated.⁶⁷ But patients differ in their tolerance for gluten, and it is hard to predict what the threshold of tolerance for gluten will be for a particular individual. Thus, it is better to avoid gluten completely.

Gluten-free if it is inherently gluten-free. If the food has a gluten-containing grain, then it should be processed to remove the gluten, and the resultant food product should not contain more than 20 parts per million of gluten. Gluten-free products that have gluten-containing grain that has been processed usually have a label indicating the gluten content in the food in parts per million.

Patients who understand the need to adhere to a gluten-free diet and the implications of not adhering to it are generally more compliant. Thus, patients need to be strongly educated that they need to adhere to a gluten-free diet and that nonadherence can cause further damage to the gut and can pose a higher risk of malignancy. Even though patients are usually concerned about the cost of gluten-free food and worry about adherence to the diet, these factors do not generally limit diet adherence.⁶⁸ All patients diagnosed with celiac disease should meet with a registered dietitian to discuss diet options based on their food preferences and to better address all their concerns.

With increasing awareness of celiac disease and with increasing numbers of patients being diagnosed with it, the food industry has recognized the need to produce gluten-free items. There are now plenty of food products available for these patients, who no longer have to forgo cakes, cookies, and other such items. Table 2 lists some common foods that patients with celiac disease can consume.

Nutritional supplements for some

If anemia is due purely to iron deficiency, it may resolve after starting a gluten-free diet, and no additional supplementation may be needed. However, if it is due to a combination of iron plus folate or vitamin B₁₂ deficiency, then folate, vitamin B₁₂, or both should be given.

In addition, if the patient is found to have a deficiency of vitamin D, then a vitamin D supplement should be given.⁶⁹ At the time of diagnosis, all patients with celiac disease should be screened for deficiencies of vitamins A, B₁₂, D, E, and K, as well as copper, zinc, folic acid, and iron.

Follow-up at 3 to 6 months

A follow-up visit should be scheduled for 3 to 6 months after the diagnosis and after that on an annual basis, and many of the abnormal laboratory tests will need to be repeated.

If intestinal or extraintestinal symptoms or nutrient deficiencies persist, then the patient’s adherence to the gluten-free diet needs to be checked. Adherence to a gluten-free diet can be assessed by checking for serologic markers of celiac disease. A decrease in baseline values can be seen within a few months of starting the diet.⁷⁰ Failure of serologic markers to decrease by the end of 1 year of a gluten-free diet usually indicates gluten contamination.⁷¹ If adherence is confirmed (ie, if baseline values fall) but symptoms persist, then further workup needs to be done to find the cause of refractory disease.

Skin lesions should also respond to a gluten-free diet

The first and foremost therapy for the skin lesions in dermatitis herpetiformis is the same as that for the intestinal manifestations in celiac disease, ie, adherence to a gluten-free diet. Soon after patients begin a gluten-free diet, the itching around the skin lesions goes away, and over time, most patients have complete resolution of the skin manifestations.

Dapsone is also frequently used to treat dermatitis herpetiformis if there is an incomplete response to a gluten-free diet or as an adjunct to diet to treat the pruritus. Patients often have a good response to dapsone.⁷²

The recommended starting dosage is 100 to 200 mg a day, and a response is usually seen within a few days. If the symptoms do not improve, the dose can be increased. Once the lesions resolve, the dose can be tapered and patients may not require any further medication. In some cases, patients may need to be chronically maintained on the lowest dose possible, due to the side effects of the drug.³

Dapsone is associated with significant adverse effects. Methemoglobinemia is the most common and is seen particularly in dosages exceeding 200 mg a day. Hemolytic anemia, another common adverse effect, is seen with dosages of more than 100 mg a day. Patients with a deficiency of glucose-6-phosphate dehydrogenase (G6PD) are at increased risk of hemolysis, and screening for G6PD deficiency is usually done before starting dapsone. Other rare adverse effects of dapsone include agranulocytosis, peripheral neuropathy, psychosis,⁷³ pancreatitis, cholestatic jaundice, bullous and exfoliative dermatitis, Stevens-Johnson syndrome, toxic epidermal necrolysis, nephrotic syndrome, and renal papillary necrosis.

Besides testing for G6PD deficiency, a complete blood cell count, a reticulocyte count, a hepatic function panel, renal function tests, and urinalysis should be done before starting dapsone therapy and repeated while on therapy. The complete blood cell count and reticulocyte count should be checked weekly for the first month, twice a month for the next 2 months, and then once every 3 months. Liver and renal function tests are to be done once every 3 months.⁷⁴

NOVEL THERAPIES BEING TESTED

Research is under way for other treatments for celiac disease besides a gluten-free diet.

Larazotide (Alba Therapeutics, Baltimore, MD) is being tested in a randomized, placebo-controlled trial. Early results indicate that it is effective in controlling both gastrointestinal and nongastrointestinal symptoms of celiac disease, but it still has to undergo phase 3 clinical trials.

Sorghum is a grain commonly used in Asia and Africa. The gluten in sorghum is different from that in wheat and is not immunogenic. In a small case series in patients with known celiac disease, sorghum did not induce diarrhea or change in levels of antibodies to tissue transglutaminase.⁷⁵

Nonimmunogenic wheat that does not contain the immunogenic gluten is being developed.

Oral enzyme supplements called glutenases are being developed. Glutenases can cleave gluten, particularly the proline and glutamine residues that make gluten resistant to degradation by gastric, pancreatic, and intestinal brush border proteases. A phase 2 trial of one of these oral enzyme supplements showed that it appeared to attenuate mucosal injury in patients with biopsy-proven celiac disease.⁷⁶

These novel therapies look promising, but for now the best treatment is lifelong adherence to the gluten-free diet.

NONRESPONSIVE AND REFRACTORY CELIAC DISEASE

Celiac disease is considered nonresponsive if its symptoms or laboratory abnormalities persist after the patient is on a gluten-free diet for 6 to 12 months. It is considered refractory if symptoms persist or recur along with villous atrophy despite adherence to the diet for more than 12 months in the absence of other causes of the symptoms. Refractory celiac disease can be further classified either as type 1 if there are typical intraepithelial lymphocytes, or as type 2 if there are atypical intraepithelial lymphocytes.

Celiac disease is nonresponsive in about 10% to 19% of cases,⁷⁶ and it is refractory in 1% to 2%.⁷⁷

Managing nonresponsive celiac disease

The first step in managing a patient with nonresponsive celiac disease is to confirm the diagnosis by reviewing the serologic tests and the biopsy samples from the time of diagnosis. If celiac disease is confirmed, then one should re-evaluate for gluten ingestion, the most common cause of nonresponsiveness.⁷⁸ If strict adherence is confirmed, then check for other causes of symptoms such as lactose or fructose intolerance. If no other cause is found, then repeat the duodenal biopsies with flow cytometry to look for CD3 and CD8 expression in T cells in the small-bowel mucosa.⁷⁹ Presence or absence of villous atrophy can point to possible other causes of malabsorption including pancreatic insufficiency, small intestinal bowel overgrowth, and microscopic colitis.

Managing refractory celiac disease

Traditionally, corticosteroids have been shown to be beneficial in alleviating symptoms in patients with refractory celiac disease but do not improve the histologic findings.⁸⁰ Because of the adverse effects associated with long-term corticosteroid use, azathioprine has been successfully used to maintain remission of the disease after induction with corticosteroids in patients with type 1 refractory celiac disease.⁸¹

Cladribine, a chemotherapeutic agent used to treat hairy cell leukemia, has shown some benefit in treating type 2 refractory celiac disease.⁸²

In type 2 refractory celiac disease, use of an immunomodulator agent carries an increased risk of transformation to lymphoma.

Because of the lack of a satisfactory response to the agents available so far to treat refractory celiac disease, more treatment options acting at the molecular level are being explored.

NONCELIAC GLUTEN SENSITIVITY DISORDER

Nonceliac gluten sensitivity disorder is an evolving concept. The clinical presentation of this disorder is similar to celiac disease in that patients may have diarrhea or other extra­intestinal symptoms when on a regular diet and have resolution of symptoms on a gluten-free diet. But unlike celiac disease, there is no serologic or histologic evidence of celiac disease even when patients are on a regular diet.

One of every 17 patients who presents with clinical features suggestive of celiac disease is found to have nonceliac gluten sensitivity disorder, not celiac disease.⁸³ In contrast to celiac disease, in which the adaptive immune system is thought to contribute to the disease process, in nonceliac gluten sensitivity disorder the innate immune system is believed to play the dominant role,⁸⁴ but the exact pathogenesis of the disease is still unclear.

The diagnosis of nonceliac gluten sensitivity disorder is one of exclusion. Celiac disease needs to be ruled out by serologic testing and by duodenal biopsy while the patient is on a regular diet, and then a trial of a gluten-free diet needs to be done to confirm resolution of symptoms before the diagnosis of nonceliac gluten sensitivity disorder can be established.

As with celiac disease, the treatment involves adhering to a gluten-free diet, but it is still not known if patients need to stay on it for the rest of their life, or if they will be able to tolerate gluten-containing products after a few years.

While most cases of breast cancer are sporadic (ie, not inherited), up to 10% are attributable to single-gene hereditary cancer syndromes.^1–4 People with these syndromes have a lifetime risk of breast cancer much higher than in the general population, and the cancers often occur at a much earlier age.

With genetic testing becoming more common, primary care physicians need to be familiar with the known syndromes, associated risks, and evidence-based recommendations for management. Here, we review the management of cancer risk in the most common hereditary breast cancer syndromes, ie:

• Hereditary breast and ovarian cancer syndrome⁵
• Hereditary diffuse gastric cancer
• Cowden syndrome (PTEN hamartoma tumor syndrome)
• Peutz-Jeghers syndrome
• Li-Fraumeni syndrome.

IT TAKES A TEAM, BUT PRIMARY CARE PHYSICIANS ARE CENTRAL

Women who have a hereditary predisposition to breast cancer face complex and emotional decisions about the best ways to manage and reduce their risks. Their management includes close clinical surveillance, chemoprevention, and surgical risk reduction.^1,4

Referral to multiple subspecialists is an important component of these patients’ preventive care. They may need referrals to a cancer genetic counselor, a high-risk breast clinic, a gynecologic oncologist, and counseling services. They may also require referrals to gastroenterologists, colorectal surgeons, endocrinologists, and endocrine surgeons, depending on the syndrome identified.

Find a genetic counselor at www.nsgc.orgConsultation with a certified genetic counselor is critical for patients harboring mutations associated with cancer risk. The National Society of Genetic Counselors maintains a directory of genetic counselors by location and practice specialty at www.nsgc.org. The counselor’s evaluation will provide patients with a detailed explanation of the cancer risks and management guidelines for their particular condition, along with offering diagnostic genetic testing if appropriate. Women with germline mutations who plan to have children should be informed about preimplantation genetic diagnosis and about fertility specialists who can perform this service if they are interested in pursuing it.⁶

Screening and management guidelines for hereditary breast cancer syndromes are evolving. While subspecialists may be involved in enhanced surveillance and preventive care, the primary care physician is the central player, with both a broader perspective and knowledge of the patient’s competing medical issues, risks, and preferences.

In addition to breast cancer, the risk of other malignancies is also higher, with the pattern varying by syndrome (Table 1).^7–20 The management of these additional risks is beyond the scope of this review; however, primary care physicians need to be familiar with these risks to provide adequate referrals.

WHO IS AT INCREASED RISK OF BREAST CANCER?

In considering recommendations to reduce the risk of breast cancer, it is useful to think of a patient as being at either high risk or average risk.

The risk of breast cancer in women in the general population is about 12%, and most cases of breast cancer occur in patients who have no known risk factors for it. “High risk” of breast cancer generally means having more than a 20% lifetime risk (ie, before age 70) of developing the condition.

Even without a hereditary cancer syndrome, a combination of reproductive, environmental, personal, and family history factors can confer a 20% lifetime risk. But for women with hereditary syndromes, the risk far exceeds 20% regardless of such risk factors. It is likely that interactions with reproductive, environmental, and personal risk factors likely affect the individual risk of a woman with a known genetic mutation, and evidence is emerging with regard to further risk stratification.

In an earlier article in this journal, Smith and colleagues²¹ reviewed how to recognize heritable breast cancer syndromes. In general, referral for genetic counseling should be considered for patients and their families who have:

• Early-onset breast cancers (before age 50)
• Bilateral breast cancers at any age
• Ovarian cancers at any age
• “Triple-negative” breast cancers (ie, estrogen receptor-negative, progesterone receptor-negative, and human epidermal growth factor receptor 2-nonamplified (HER2-negative)
• Male breast cancer at any age
• Cancers affecting multiple individuals and in multiple generations.
• Breast, ovarian, pancreatic or prostate cancer in families with Ashkenazi Jewish ancestry

HEREDITARY BREAST CANCER SYNDROMES

Hereditary breast and ovarian cancer syndrome

The most common of these syndromes is hereditary breast and ovarian cancer syndrome, caused by germline mutations in the tumor-suppressor genes BRCA1 or BRCA2.⁷ The estimated prevalence of BRCA1 mutations is 1 in 250 to 300, and the prevalence of BRCA2 mutations is 1 in 800.^1,4 However, in families of Ashkenazi Jewish ancestry, the population frequency of either a BRCA1 or BRCA2 mutation is approximately 1 in 40.^1,4,6

Women with BRCA1 or BRCA2 mutations have a lifetime risk of breast cancer of up to 87%

Women with BRCA1 or BRCA2 mutations have a lifetime risk of breast cancer of up to 87%, or 5 to 7 times higher than in the general population, with the risk rising steeply beginning at age 30.^1,5,8 In addition, the lifetime risk of ovarian cancer is nearly 59% in BRCA1 mutation carriers and 17% in BRCA2 mutation carriers.²²

A meta-analysis found that BRCA1 mutation carriers diagnosed with cancer in one breast have a 5-year risk of developing cancer in the other breast of 15%, and BRCA2 mutation carriers have a risk of 9%.²³ Overall, the risk of contralateral breast cancer is about 3% per year.^3,4,24

BRCA1 mutations are strongly associated with triple-negative breast cancers.^1,3,4

Hereditary diffuse gastric cancer

Hereditary diffuse gastric cancer is an autosomal-dominant syndrome associated with mutations in the CDH1 gene, although up to 75% of patients with this syndrome do not have an identifiable CDH1 mutation.^9,25,26 In cases in which there is no identifiable CDH1 mutation, the diagnosis is made on the basis of the patient’s medical and family history.

Hereditary diffuse gastric cancer is associated with an increased risk of the lobular subtype of breast cancer as well as diffuse gastric cancer. The cumulative lifetime risk of breast cancer in women with CDH1 mutations is 39% to 52%,^6,9–11,25 and their lifetime risk of diffuse gastric cancer is 83%.⁹ The combined risk of breast cancer and gastric cancer in women with this syndrome is 90% by age 80.⁹

Cowden syndrome (PTEN hamartoma tumor syndrome)

Cowden syndrome (PTEN hamartoma tumor syndrome) is caused by mutations in PTEN, another tumor-suppressor gene.¹¹ The primary clinical concerns are melanoma and breast, endometrial, thyroid (follicular or papillary), colon, and renal cell cancers. Women with a PTEN mutation have a twofold greater risk of developing any type of cancer than men with a PTEN mutation.¹² The cumulative lifetime risk of invasive breast cancer in women with this syndrome is 70% to 85%.^11–13

Peutz-Jeghers syndrome

Peutz-Jeghers syndrome is an autosomal dominant polyposis disorder caused, in most patients, by a mutation in the serine/threonine kinase tumor-suppressor gene STK11.¹⁴

Patients with Peutz-Jeghers syndrome have higher risks of gastrointestinal, breast, gynecologic (uterine, ovarian, and cervical), pancreatic, and lung cancers. In women, the lifetime risk of breast cancer is 44% to 50% by age 70, regardless of the type of mutation.^6,14,15 Breast cancers associated with Peutz-Jeghers syndrome are usually ductal, and the mean age at diagnosis is 37 years.¹⁶

Li-Fraumeni syndrome

Li-Fraumeni syndrome is an autosomal-dominant disorder caused by germline mutations in the TP53 gene, which codes for a transcription factor associated with cell proliferation and apoptosis.²⁷

Think about other highly penetrant genetic mutations in a young breast cancer patient with no mutation in BRCA1 or BRCA2These mutations confer a lifetime cancer risk of 93% in women (mainly breast cancer) and 68% in men.^1,27 Other cancers associated with TP53 mutations include sarcomas, brain cancer, leukemia, and adrenocortical tumors. Germline TP53 mutations are responsible for approximately 1% of all breast cancers.^1,4

Breast cancers can occur at a young age in patients with a TP53 mutation. Women with TP53 mutations are 18 times more likely to develop breast cancer before age 45 compared with the general population.⁴

It is important to consider a TP53 mutation in premenopausal women or women less than 30 years of age with breast cancer who have no mutations in BRCA1 and BRCA2.¹

MANAGING PATIENTS WITH GENETIC PREDISPOSITION TO BREAST CANCER

Management for patients at high risk fall into three broad categories: clinical surveillance, chemoprevention, and surgical risk reduction. The utility and benefit of each depend to a large degree on the patient’s specific mutation, family history, and comorbidities. Decisions must be shared with the patient.

CLOSE CLINICAL SURVEILLANCE

Consensus guidelines for cancer screening in the syndromes described here are available from the National Comprehensive Cancer Network at www.nccn.org and are summarized in Table 2.^26,28 While the guidelines are broadly applicable to all women with these conditions, some individualization is required based on personal and family medical history.

In general, screening begins at the ages listed in Table 2 or 10 years earlier than the age at which cancer developed in the first affected relative, whichever is earlier. However, screening decisions are shared with the patient and are sometimes affected by significant out-of-pocket costs for the patient and anxiety resulting from the test or subsequent test findings, which must all be considered.

Breast self-awareness and clinical breast examination

Although controversial in the general population, breast self-examination is recommended for patients carrying mutations that increase risk.⁶

Discuss breast self-awareness with all women patients at age 18

A discussion about breast self-awareness is recommended for all women at the age of 18. It should include the signs and symptoms of breast cancer, what feels “normal” to the patient, and what is known about modifiable risk factors for breast cancer. The patient should also be told to report any changes in her personal or family history.

Clinical breast examinations should be done every 6 months, as some cancers are found clinically, particularly in young women with dense tissue, and confirmed by diagnostic imaging and targeted ultrasonography.

Radiographic surveillance

Mammography and magnetic resonance imaging (MRI) are also important components of a breast cancer surveillance regimen in women at high risk. Adherence to a well-formulated plan of clinical and radiographic examinations increases early detection in patients who have a hereditary predisposition to breast cancer.

MRI is more sensitive than mammography and reduces the likelihood of finding advanced cancers by up to 70% compared with mammography in women at high risk of breast cancer.^29–31 The sensitivity of breast MRI alone ranges from 71% to 100%, and the sensitivity increases to 89% to 100% when combined with mammography. In contrast, the sensitivity of mammography alone is 25% to 59%.²⁹ MRI has also been shown to be cost-effective when added to mammography and physical examination in women at high risk.^5,32

Adding MRI to the breast cancer screening regimen has been under discussion and has been endorsed by the American Cancer Society in formal recommendations set forth in 2007 for patients with known hereditary cancer syndromes, in untested first-degree relatives of identified genetic mutation carriers, or in women who have an estimated lifetime risk of breast cancer of 20% or more, as determined by models largely dependent on family history.³³

But MRI has a downside—it is less specific than mammography.^29,33 Its lower specificity (77% to 90% vs 95% with mammography alone) leads to additional radiographic studies and tissue samplings for the “suspicious” lesions discovered. From 3% to 15% of screening breast MRIs result in a biopsy, and the proportion of biopsies that reveal cancer is 13% to 40%.³³ Furthermore, by itself, MRI has not been shown to reduce mortality in any high-risk group.

Mammography remains useful in conjunction with MRI due to its ability to detect breast calcifications, which may be the earliest sign of breast cancer, and ability to detect changes in breast architecture. A typical screening program (Table 2) should incorporate both modalities, commonly offset by 6 months (eg, mammography at baseline, then MRI 6 months later, then mammography again 6 months after that, and so on) to increase the detection of interval cancer development.

Chemoprevention

Chemoprevention means taking medications to reduce the risk. Certain selective estrogen receptor modulators and aromatase inhibitors decrease the risk of invasive breast cancer in healthy women at high risk. These drugs include tamoxifen, which can be used before menopause, and raloxifene, anastrozole, and exemestane, which must be used only after menopause.

Because data are limited, we cannot make any generalized recommendations about chemoprevention in patients with hereditary breast cancer syndromes. Decisions about chemoprevention should take into account the patient’s personal and family histories. Often, a medical oncologist or medical breast specialist can help by discussing the risks and benefits for the individual patient.

Tamoxifen has been the most studied, mainly in BRCA mutation carriers.^6,34–37 As in the general population, tamoxifen reduces the incidence of estrogen receptor-positive breast cancers by 50%.^36–38 It has not been shown to significantly reduce breast cancer risk in premenopausal women with BRCA1 mutations,³⁷ most likely because most cancers that occur in this group are estrogen receptor-negative. In patients with a history of breast cancer, however, tamoxifen has been shown to reduce the risk of developing contralateral breast cancer by 45% to 60% in both BRCA1 and BRCA2 mutation carriers.^6,35

There is also little evidence that giving a chemopreventive agent after bilateral salpingo-oophorectomy reduces the risk further in premenopausal BRCA mutation carriers.³⁵ These patients often receive hormonal therapy with estrogen, which currently would preclude the use of tamoxifen. Tamoxifen in postmenopausal women is associated with a small increased risk of venous thromboembolic disease and endometrial cancer.³⁸

Oral contraceptives reduce the risk of ovarian cancer by up to 50% in BRCA1 mutation carriers and up to 60% in BRCA2 mutation carriers.⁶ However, data conflict on their effect on the risk of breast cancer in BRCA1 and BRCA2 mutation carriers.³⁹

Decisions about chemoprevention with agents other than tamoxifen and in syndromes other than hereditary breast and ovarian cancer syndrome must take into consideration the existing lack of data in this area.

SURGICAL PROPHYLAXIS

Surgical prophylactic options for patients at genetic risk of breast cancer are bilateral mastectomy and bilateral salpingo-oophorectomy.

Prophylactic mastectomy

Bilateral risk-reducing mastectomy reduces the risk of breast cancer by at least 90%^24,39,40 and greatly reduces the need for complex surveillance. Patients are often followed annually clinically, with single-view mammography if they have tissue flap reconstruction.

Nipple-sparing and skin-sparing mastectomies, which facilitate reconstruction and cosmetic outcomes, are an option in the risk-reduction setting and have been shown thus far to be safe.^41–43 In patients with breast cancer, the overall breast cancer recurrence rates with nipple-sparing mastectomy are similar to those of traditional mastectomy and breast conservation treatment.⁴¹

In patients at very high risk of breast cancer, risk-reducing operations also reduce the risk of ultimately needing chemotherapy and radiation to treat breast cancer, as the risk of developing breast cancer is significantly lowered.

The timing of risk-reducing mastectomy depends largely on personal and family medical history and personal choice. Bilateral mastectomy at age 25 results in the greatest survival gain for patients with hereditary breast and ovarian cancer syndrome.⁵ Such precise data are not available for other hereditary cancer syndromes, but it is reasonable to consider bilateral mastectomy as an option for any woman with a highly penetrant genetic mutation that predisposes her to breast cancer. Special consideration in the timing of risk-reducing mastectomy must be given to women with Li-Fraumeni syndrome, as this condition is often associated with an earlier age at breast cancer diagnosis (before age 30).¹

Family planning, sexuality, self-image, and the anxiety associated with both cancer risk and surveillance are all factors women consider when deciding whether and when to undergo mastectomy. A survey of 12 high-risk women who elected prophylactic mastectomy elicited feelings of some regret in 3 of them, while all expressed a sense of relief and reduced anxiety related to both cancer risk and screenings.²⁴ Another group of 14 women surveyed after the surgery reported initial distress related to physical appearance, self-image, and intimacy but also reported a significant decrease in anxiety related to breast cancer risk and were largely satisfied with their decision.⁴⁴

Prophylactic salpingo-oophorectomy

In patients who have pathogenic mutations in BRCA1 or 2, prophylactic salpingo-oophorectomy before age 40 decreases the risk of ovarian cancer by up to 96% and breast cancer by 50%.^1,37,45 This operation, in fact, is the only intervention that has been shown to reduce the mortality rate in patients with a hereditary predisposition to cancer.⁴⁶

We recommend that women with hereditary breast and ovarian cancer syndrome strongly consider prophylactic salpingo-oophorectomy by age 40 or when childbearing is complete for the greatest reduction in risk.^1,5 In 2006, Domchek et al⁴⁶ reported an overall decrease in the mortality rate in BRCA1/2-positive patients who underwent this surgery, but not in breast cancer-specific or ovarian cancer-specific mortality.

On the other hand, removing the ovaries before menopause places women at risk of serious complications associated with premature loss of gonadal hormones, including cardiovascular disease, decreased bone density, reduced sexual satisfaction, dyspareunia, hot flashes, and night sweats.⁴⁷ Therefore, it is generally reserved for women who are also at risk of ovarian cancer.

Hormonal therapy, ie, estrogen therapy for patients who choose complete hysterectomy, and estrogen-progesterone therapy for patients who choose to keep their uterus, reduces menopausal symptoms and symptoms of sexual dissatisfaction and has not thus far been shown to increase breast cancer risk.^1,34 However, this information is from nonrandomized studies, which are inherently limited.

It is important to address and modify risk factors for heart disease and osteoporosis in women with premature surgical menopause, as they may be particularly vulnerable to these conditions.

HEREDITARY BREAST CANCER IN MEN

Fewer than 1% of cases of breast cancer arise in men, and fewer than 1% of cases of cancer in men are breast cancer.

Male breast cancer is more likely than female breast cancer to be estrogen receptor- and progesterone receptor-positive. In an analysis of the Surveillance, Epidemiology, and End Results registry between 1973 and 2005, triple-negative breast cancer was found in 23% of female patients but only 7.6% of male patients.²

Male breast cancer is most common in families with BRCA2, and to a lesser degree, BRCA1 mutations. Other genetic disorders including Li-Fraumeni syndrome, hereditary nonpolyposis colorectal cancer, and Klinefelter syndrome also increase the risk of male breast cancer. A genetic predisposition for breast cancer is present in approximately 10% of male breast cancer patients.² Any man with breast cancer, therefore, should be referred for genetic counseling.

In men, a BRCA2 mutation confers a lifetime risk of breast cancer of 5% to 10%.² This is similar to the lifetime risk of breast cancer for the average woman but it is still significant, as the lifetime risk of breast cancer for the average man is 0.1%.^1,2

Five-year survival rates in male breast cancer range from only 36% to 66%, most likely because it is usually diagnosed in later stages, as men are not routinely screened for breast cancer. In men with known hereditary susceptibility, National Comprehensive Cancer Network guidelines recommend that they be educated about and begin breast self-examination at the age of 35 and be clinically examined every 12 months starting at age 35.⁴⁸ There are limited data to support breast imaging in men. High-risk surveillance with MRI screening in this group is not recommended. Prostate cancer screening is recommended for men with BRCA2 mutations starting at age 40, and should be considered for men with BRCA1 mutations starting at age 40.

No specific guidelines exist for pancreatic cancer and melanoma, but screening may be individualized based on cancers observed in the family.

While most cases of breast cancer are sporadic (ie, not inherited), up to 10% are attributable to single-gene hereditary cancer syndromes.^1–4 People with these syndromes have a lifetime risk of breast cancer much higher than in the general population, and the cancers often occur at a much earlier age.

With genetic testing becoming more common, primary care physicians need to be familiar with the known syndromes, associated risks, and evidence-based recommendations for management. Here, we review the management of cancer risk in the most common hereditary breast cancer syndromes, ie:

• Hereditary breast and ovarian cancer syndrome⁵
• Hereditary diffuse gastric cancer
• Cowden syndrome (PTEN hamartoma tumor syndrome)
• Peutz-Jeghers syndrome
• Li-Fraumeni syndrome.

IT TAKES A TEAM, BUT PRIMARY CARE PHYSICIANS ARE CENTRAL

Women who have a hereditary predisposition to breast cancer face complex and emotional decisions about the best ways to manage and reduce their risks. Their management includes close clinical surveillance, chemoprevention, and surgical risk reduction.^1,4

Referral to multiple subspecialists is an important component of these patients’ preventive care. They may need referrals to a cancer genetic counselor, a high-risk breast clinic, a gynecologic oncologist, and counseling services. They may also require referrals to gastroenterologists, colorectal surgeons, endocrinologists, and endocrine surgeons, depending on the syndrome identified.

Find a genetic counselor at www.nsgc.orgConsultation with a certified genetic counselor is critical for patients harboring mutations associated with cancer risk. The National Society of Genetic Counselors maintains a directory of genetic counselors by location and practice specialty at www.nsgc.org. The counselor’s evaluation will provide patients with a detailed explanation of the cancer risks and management guidelines for their particular condition, along with offering diagnostic genetic testing if appropriate. Women with germline mutations who plan to have children should be informed about preimplantation genetic diagnosis and about fertility specialists who can perform this service if they are interested in pursuing it.⁶

Screening and management guidelines for hereditary breast cancer syndromes are evolving. While subspecialists may be involved in enhanced surveillance and preventive care, the primary care physician is the central player, with both a broader perspective and knowledge of the patient’s competing medical issues, risks, and preferences.

In addition to breast cancer, the risk of other malignancies is also higher, with the pattern varying by syndrome (Table 1).^7–20 The management of these additional risks is beyond the scope of this review; however, primary care physicians need to be familiar with these risks to provide adequate referrals.

WHO IS AT INCREASED RISK OF BREAST CANCER?

In considering recommendations to reduce the risk of breast cancer, it is useful to think of a patient as being at either high risk or average risk.

The risk of breast cancer in women in the general population is about 12%, and most cases of breast cancer occur in patients who have no known risk factors for it. “High risk” of breast cancer generally means having more than a 20% lifetime risk (ie, before age 70) of developing the condition.

Even without a hereditary cancer syndrome, a combination of reproductive, environmental, personal, and family history factors can confer a 20% lifetime risk. But for women with hereditary syndromes, the risk far exceeds 20% regardless of such risk factors. It is likely that interactions with reproductive, environmental, and personal risk factors likely affect the individual risk of a woman with a known genetic mutation, and evidence is emerging with regard to further risk stratification.

In an earlier article in this journal, Smith and colleagues²¹ reviewed how to recognize heritable breast cancer syndromes. In general, referral for genetic counseling should be considered for patients and their families who have:

• Early-onset breast cancers (before age 50)
• Bilateral breast cancers at any age
• Ovarian cancers at any age
• “Triple-negative” breast cancers (ie, estrogen receptor-negative, progesterone receptor-negative, and human epidermal growth factor receptor 2-nonamplified (HER2-negative)
• Male breast cancer at any age
• Cancers affecting multiple individuals and in multiple generations.
• Breast, ovarian, pancreatic or prostate cancer in families with Ashkenazi Jewish ancestry

HEREDITARY BREAST CANCER SYNDROMES

Hereditary breast and ovarian cancer syndrome

The most common of these syndromes is hereditary breast and ovarian cancer syndrome, caused by germline mutations in the tumor-suppressor genes BRCA1 or BRCA2.⁷ The estimated prevalence of BRCA1 mutations is 1 in 250 to 300, and the prevalence of BRCA2 mutations is 1 in 800.^1,4 However, in families of Ashkenazi Jewish ancestry, the population frequency of either a BRCA1 or BRCA2 mutation is approximately 1 in 40.^1,4,6

Women with BRCA1 or BRCA2 mutations have a lifetime risk of breast cancer of up to 87%

Women with BRCA1 or BRCA2 mutations have a lifetime risk of breast cancer of up to 87%, or 5 to 7 times higher than in the general population, with the risk rising steeply beginning at age 30.^1,5,8 In addition, the lifetime risk of ovarian cancer is nearly 59% in BRCA1 mutation carriers and 17% in BRCA2 mutation carriers.²²

A meta-analysis found that BRCA1 mutation carriers diagnosed with cancer in one breast have a 5-year risk of developing cancer in the other breast of 15%, and BRCA2 mutation carriers have a risk of 9%.²³ Overall, the risk of contralateral breast cancer is about 3% per year.^3,4,24

BRCA1 mutations are strongly associated with triple-negative breast cancers.^1,3,4

Hereditary diffuse gastric cancer

Hereditary diffuse gastric cancer is an autosomal-dominant syndrome associated with mutations in the CDH1 gene, although up to 75% of patients with this syndrome do not have an identifiable CDH1 mutation.^9,25,26 In cases in which there is no identifiable CDH1 mutation, the diagnosis is made on the basis of the patient’s medical and family history.

Hereditary diffuse gastric cancer is associated with an increased risk of the lobular subtype of breast cancer as well as diffuse gastric cancer. The cumulative lifetime risk of breast cancer in women with CDH1 mutations is 39% to 52%,^6,9–11,25 and their lifetime risk of diffuse gastric cancer is 83%.⁹ The combined risk of breast cancer and gastric cancer in women with this syndrome is 90% by age 80.⁹

Cowden syndrome (PTEN hamartoma tumor syndrome)

Cowden syndrome (PTEN hamartoma tumor syndrome) is caused by mutations in PTEN, another tumor-suppressor gene.¹¹ The primary clinical concerns are melanoma and breast, endometrial, thyroid (follicular or papillary), colon, and renal cell cancers. Women with a PTEN mutation have a twofold greater risk of developing any type of cancer than men with a PTEN mutation.¹² The cumulative lifetime risk of invasive breast cancer in women with this syndrome is 70% to 85%.^11–13

Peutz-Jeghers syndrome

Peutz-Jeghers syndrome is an autosomal dominant polyposis disorder caused, in most patients, by a mutation in the serine/threonine kinase tumor-suppressor gene STK11.¹⁴

Patients with Peutz-Jeghers syndrome have higher risks of gastrointestinal, breast, gynecologic (uterine, ovarian, and cervical), pancreatic, and lung cancers. In women, the lifetime risk of breast cancer is 44% to 50% by age 70, regardless of the type of mutation.^6,14,15 Breast cancers associated with Peutz-Jeghers syndrome are usually ductal, and the mean age at diagnosis is 37 years.¹⁶

Li-Fraumeni syndrome

Li-Fraumeni syndrome is an autosomal-dominant disorder caused by germline mutations in the TP53 gene, which codes for a transcription factor associated with cell proliferation and apoptosis.²⁷

Think about other highly penetrant genetic mutations in a young breast cancer patient with no mutation in BRCA1 or BRCA2These mutations confer a lifetime cancer risk of 93% in women (mainly breast cancer) and 68% in men.^1,27 Other cancers associated with TP53 mutations include sarcomas, brain cancer, leukemia, and adrenocortical tumors. Germline TP53 mutations are responsible for approximately 1% of all breast cancers.^1,4

Breast cancers can occur at a young age in patients with a TP53 mutation. Women with TP53 mutations are 18 times more likely to develop breast cancer before age 45 compared with the general population.⁴

It is important to consider a TP53 mutation in premenopausal women or women less than 30 years of age with breast cancer who have no mutations in BRCA1 and BRCA2.¹

MANAGING PATIENTS WITH GENETIC PREDISPOSITION TO BREAST CANCER

Management for patients at high risk fall into three broad categories: clinical surveillance, chemoprevention, and surgical risk reduction. The utility and benefit of each depend to a large degree on the patient’s specific mutation, family history, and comorbidities. Decisions must be shared with the patient.

CLOSE CLINICAL SURVEILLANCE

Consensus guidelines for cancer screening in the syndromes described here are available from the National Comprehensive Cancer Network at www.nccn.org and are summarized in Table 2.^26,28 While the guidelines are broadly applicable to all women with these conditions, some individualization is required based on personal and family medical history.

In general, screening begins at the ages listed in Table 2 or 10 years earlier than the age at which cancer developed in the first affected relative, whichever is earlier. However, screening decisions are shared with the patient and are sometimes affected by significant out-of-pocket costs for the patient and anxiety resulting from the test or subsequent test findings, which must all be considered.

Breast self-awareness and clinical breast examination

Although controversial in the general population, breast self-examination is recommended for patients carrying mutations that increase risk.⁶

Discuss breast self-awareness with all women patients at age 18

A discussion about breast self-awareness is recommended for all women at the age of 18. It should include the signs and symptoms of breast cancer, what feels “normal” to the patient, and what is known about modifiable risk factors for breast cancer. The patient should also be told to report any changes in her personal or family history.

Clinical breast examinations should be done every 6 months, as some cancers are found clinically, particularly in young women with dense tissue, and confirmed by diagnostic imaging and targeted ultrasonography.

Radiographic surveillance

Mammography and magnetic resonance imaging (MRI) are also important components of a breast cancer surveillance regimen in women at high risk. Adherence to a well-formulated plan of clinical and radiographic examinations increases early detection in patients who have a hereditary predisposition to breast cancer.

MRI is more sensitive than mammography and reduces the likelihood of finding advanced cancers by up to 70% compared with mammography in women at high risk of breast cancer.^29–31 The sensitivity of breast MRI alone ranges from 71% to 100%, and the sensitivity increases to 89% to 100% when combined with mammography. In contrast, the sensitivity of mammography alone is 25% to 59%.²⁹ MRI has also been shown to be cost-effective when added to mammography and physical examination in women at high risk.^5,32

Adding MRI to the breast cancer screening regimen has been under discussion and has been endorsed by the American Cancer Society in formal recommendations set forth in 2007 for patients with known hereditary cancer syndromes, in untested first-degree relatives of identified genetic mutation carriers, or in women who have an estimated lifetime risk of breast cancer of 20% or more, as determined by models largely dependent on family history.³³

But MRI has a downside—it is less specific than mammography.^29,33 Its lower specificity (77% to 90% vs 95% with mammography alone) leads to additional radiographic studies and tissue samplings for the “suspicious” lesions discovered. From 3% to 15% of screening breast MRIs result in a biopsy, and the proportion of biopsies that reveal cancer is 13% to 40%.³³ Furthermore, by itself, MRI has not been shown to reduce mortality in any high-risk group.

Mammography remains useful in conjunction with MRI due to its ability to detect breast calcifications, which may be the earliest sign of breast cancer, and ability to detect changes in breast architecture. A typical screening program (Table 2) should incorporate both modalities, commonly offset by 6 months (eg, mammography at baseline, then MRI 6 months later, then mammography again 6 months after that, and so on) to increase the detection of interval cancer development.

Chemoprevention

Chemoprevention means taking medications to reduce the risk. Certain selective estrogen receptor modulators and aromatase inhibitors decrease the risk of invasive breast cancer in healthy women at high risk. These drugs include tamoxifen, which can be used before menopause, and raloxifene, anastrozole, and exemestane, which must be used only after menopause.

Because data are limited, we cannot make any generalized recommendations about chemoprevention in patients with hereditary breast cancer syndromes. Decisions about chemoprevention should take into account the patient’s personal and family histories. Often, a medical oncologist or medical breast specialist can help by discussing the risks and benefits for the individual patient.

Tamoxifen has been the most studied, mainly in BRCA mutation carriers.^6,34–37 As in the general population, tamoxifen reduces the incidence of estrogen receptor-positive breast cancers by 50%.^36–38 It has not been shown to significantly reduce breast cancer risk in premenopausal women with BRCA1 mutations,³⁷ most likely because most cancers that occur in this group are estrogen receptor-negative. In patients with a history of breast cancer, however, tamoxifen has been shown to reduce the risk of developing contralateral breast cancer by 45% to 60% in both BRCA1 and BRCA2 mutation carriers.^6,35

There is also little evidence that giving a chemopreventive agent after bilateral salpingo-oophorectomy reduces the risk further in premenopausal BRCA mutation carriers.³⁵ These patients often receive hormonal therapy with estrogen, which currently would preclude the use of tamoxifen. Tamoxifen in postmenopausal women is associated with a small increased risk of venous thromboembolic disease and endometrial cancer.³⁸

Oral contraceptives reduce the risk of ovarian cancer by up to 50% in BRCA1 mutation carriers and up to 60% in BRCA2 mutation carriers.⁶ However, data conflict on their effect on the risk of breast cancer in BRCA1 and BRCA2 mutation carriers.³⁹

Decisions about chemoprevention with agents other than tamoxifen and in syndromes other than hereditary breast and ovarian cancer syndrome must take into consideration the existing lack of data in this area.

SURGICAL PROPHYLAXIS

Surgical prophylactic options for patients at genetic risk of breast cancer are bilateral mastectomy and bilateral salpingo-oophorectomy.

Prophylactic mastectomy

Bilateral risk-reducing mastectomy reduces the risk of breast cancer by at least 90%^24,39,40 and greatly reduces the need for complex surveillance. Patients are often followed annually clinically, with single-view mammography if they have tissue flap reconstruction.

Nipple-sparing and skin-sparing mastectomies, which facilitate reconstruction and cosmetic outcomes, are an option in the risk-reduction setting and have been shown thus far to be safe.^41–43 In patients with breast cancer, the overall breast cancer recurrence rates with nipple-sparing mastectomy are similar to those of traditional mastectomy and breast conservation treatment.⁴¹

In patients at very high risk of breast cancer, risk-reducing operations also reduce the risk of ultimately needing chemotherapy and radiation to treat breast cancer, as the risk of developing breast cancer is significantly lowered.

The timing of risk-reducing mastectomy depends largely on personal and family medical history and personal choice. Bilateral mastectomy at age 25 results in the greatest survival gain for patients with hereditary breast and ovarian cancer syndrome.⁵ Such precise data are not available for other hereditary cancer syndromes, but it is reasonable to consider bilateral mastectomy as an option for any woman with a highly penetrant genetic mutation that predisposes her to breast cancer. Special consideration in the timing of risk-reducing mastectomy must be given to women with Li-Fraumeni syndrome, as this condition is often associated with an earlier age at breast cancer diagnosis (before age 30).¹

Family planning, sexuality, self-image, and the anxiety associated with both cancer risk and surveillance are all factors women consider when deciding whether and when to undergo mastectomy. A survey of 12 high-risk women who elected prophylactic mastectomy elicited feelings of some regret in 3 of them, while all expressed a sense of relief and reduced anxiety related to both cancer risk and screenings.²⁴ Another group of 14 women surveyed after the surgery reported initial distress related to physical appearance, self-image, and intimacy but also reported a significant decrease in anxiety related to breast cancer risk and were largely satisfied with their decision.⁴⁴

Prophylactic salpingo-oophorectomy

In patients who have pathogenic mutations in BRCA1 or 2, prophylactic salpingo-oophorectomy before age 40 decreases the risk of ovarian cancer by up to 96% and breast cancer by 50%.^1,37,45 This operation, in fact, is the only intervention that has been shown to reduce the mortality rate in patients with a hereditary predisposition to cancer.⁴⁶

We recommend that women with hereditary breast and ovarian cancer syndrome strongly consider prophylactic salpingo-oophorectomy by age 40 or when childbearing is complete for the greatest reduction in risk.^1,5 In 2006, Domchek et al⁴⁶ reported an overall decrease in the mortality rate in BRCA1/2-positive patients who underwent this surgery, but not in breast cancer-specific or ovarian cancer-specific mortality.

On the other hand, removing the ovaries before menopause places women at risk of serious complications associated with premature loss of gonadal hormones, including cardiovascular disease, decreased bone density, reduced sexual satisfaction, dyspareunia, hot flashes, and night sweats.⁴⁷ Therefore, it is generally reserved for women who are also at risk of ovarian cancer.

Hormonal therapy, ie, estrogen therapy for patients who choose complete hysterectomy, and estrogen-progesterone therapy for patients who choose to keep their uterus, reduces menopausal symptoms and symptoms of sexual dissatisfaction and has not thus far been shown to increase breast cancer risk.^1,34 However, this information is from nonrandomized studies, which are inherently limited.

It is important to address and modify risk factors for heart disease and osteoporosis in women with premature surgical menopause, as they may be particularly vulnerable to these conditions.

HEREDITARY BREAST CANCER IN MEN

Fewer than 1% of cases of breast cancer arise in men, and fewer than 1% of cases of cancer in men are breast cancer.

Male breast cancer is more likely than female breast cancer to be estrogen receptor- and progesterone receptor-positive. In an analysis of the Surveillance, Epidemiology, and End Results registry between 1973 and 2005, triple-negative breast cancer was found in 23% of female patients but only 7.6% of male patients.²

Male breast cancer is most common in families with BRCA2, and to a lesser degree, BRCA1 mutations. Other genetic disorders including Li-Fraumeni syndrome, hereditary nonpolyposis colorectal cancer, and Klinefelter syndrome also increase the risk of male breast cancer. A genetic predisposition for breast cancer is present in approximately 10% of male breast cancer patients.² Any man with breast cancer, therefore, should be referred for genetic counseling.

In men, a BRCA2 mutation confers a lifetime risk of breast cancer of 5% to 10%.² This is similar to the lifetime risk of breast cancer for the average woman but it is still significant, as the lifetime risk of breast cancer for the average man is 0.1%.^1,2

Five-year survival rates in male breast cancer range from only 36% to 66%, most likely because it is usually diagnosed in later stages, as men are not routinely screened for breast cancer. In men with known hereditary susceptibility, National Comprehensive Cancer Network guidelines recommend that they be educated about and begin breast self-examination at the age of 35 and be clinically examined every 12 months starting at age 35.⁴⁸ There are limited data to support breast imaging in men. High-risk surveillance with MRI screening in this group is not recommended. Prostate cancer screening is recommended for men with BRCA2 mutations starting at age 40, and should be considered for men with BRCA1 mutations starting at age 40.

No specific guidelines exist for pancreatic cancer and melanoma, but screening may be individualized based on cancers observed in the family.

While most cases of breast cancer are sporadic (ie, not inherited), up to 10% are attributable to single-gene hereditary cancer syndromes.^1–4 People with these syndromes have a lifetime risk of breast cancer much higher than in the general population, and the cancers often occur at a much earlier age.

With genetic testing becoming more common, primary care physicians need to be familiar with the known syndromes, associated risks, and evidence-based recommendations for management. Here, we review the management of cancer risk in the most common hereditary breast cancer syndromes, ie:

• Hereditary breast and ovarian cancer syndrome⁵
• Hereditary diffuse gastric cancer
• Cowden syndrome (PTEN hamartoma tumor syndrome)
• Peutz-Jeghers syndrome
• Li-Fraumeni syndrome.

IT TAKES A TEAM, BUT PRIMARY CARE PHYSICIANS ARE CENTRAL

Women who have a hereditary predisposition to breast cancer face complex and emotional decisions about the best ways to manage and reduce their risks. Their management includes close clinical surveillance, chemoprevention, and surgical risk reduction.^1,4

Referral to multiple subspecialists is an important component of these patients’ preventive care. They may need referrals to a cancer genetic counselor, a high-risk breast clinic, a gynecologic oncologist, and counseling services. They may also require referrals to gastroenterologists, colorectal surgeons, endocrinologists, and endocrine surgeons, depending on the syndrome identified.

Find a genetic counselor at www.nsgc.orgConsultation with a certified genetic counselor is critical for patients harboring mutations associated with cancer risk. The National Society of Genetic Counselors maintains a directory of genetic counselors by location and practice specialty at www.nsgc.org. The counselor’s evaluation will provide patients with a detailed explanation of the cancer risks and management guidelines for their particular condition, along with offering diagnostic genetic testing if appropriate. Women with germline mutations who plan to have children should be informed about preimplantation genetic diagnosis and about fertility specialists who can perform this service if they are interested in pursuing it.⁶

Screening and management guidelines for hereditary breast cancer syndromes are evolving. While subspecialists may be involved in enhanced surveillance and preventive care, the primary care physician is the central player, with both a broader perspective and knowledge of the patient’s competing medical issues, risks, and preferences.

In addition to breast cancer, the risk of other malignancies is also higher, with the pattern varying by syndrome (Table 1).^7–20 The management of these additional risks is beyond the scope of this review; however, primary care physicians need to be familiar with these risks to provide adequate referrals.

WHO IS AT INCREASED RISK OF BREAST CANCER?

In considering recommendations to reduce the risk of breast cancer, it is useful to think of a patient as being at either high risk or average risk.

The risk of breast cancer in women in the general population is about 12%, and most cases of breast cancer occur in patients who have no known risk factors for it. “High risk” of breast cancer generally means having more than a 20% lifetime risk (ie, before age 70) of developing the condition.

Even without a hereditary cancer syndrome, a combination of reproductive, environmental, personal, and family history factors can confer a 20% lifetime risk. But for women with hereditary syndromes, the risk far exceeds 20% regardless of such risk factors. It is likely that interactions with reproductive, environmental, and personal risk factors likely affect the individual risk of a woman with a known genetic mutation, and evidence is emerging with regard to further risk stratification.

In an earlier article in this journal, Smith and colleagues²¹ reviewed how to recognize heritable breast cancer syndromes. In general, referral for genetic counseling should be considered for patients and their families who have:

• Early-onset breast cancers (before age 50)
• Bilateral breast cancers at any age
• Ovarian cancers at any age
• “Triple-negative” breast cancers (ie, estrogen receptor-negative, progesterone receptor-negative, and human epidermal growth factor receptor 2-nonamplified (HER2-negative)
• Male breast cancer at any age
• Cancers affecting multiple individuals and in multiple generations.
• Breast, ovarian, pancreatic or prostate cancer in families with Ashkenazi Jewish ancestry

HEREDITARY BREAST CANCER SYNDROMES

Hereditary breast and ovarian cancer syndrome

The most common of these syndromes is hereditary breast and ovarian cancer syndrome, caused by germline mutations in the tumor-suppressor genes BRCA1 or BRCA2.⁷ The estimated prevalence of BRCA1 mutations is 1 in 250 to 300, and the prevalence of BRCA2 mutations is 1 in 800.^1,4 However, in families of Ashkenazi Jewish ancestry, the population frequency of either a BRCA1 or BRCA2 mutation is approximately 1 in 40.^1,4,6

Women with BRCA1 or BRCA2 mutations have a lifetime risk of breast cancer of up to 87%

Women with BRCA1 or BRCA2 mutations have a lifetime risk of breast cancer of up to 87%, or 5 to 7 times higher than in the general population, with the risk rising steeply beginning at age 30.^1,5,8 In addition, the lifetime risk of ovarian cancer is nearly 59% in BRCA1 mutation carriers and 17% in BRCA2 mutation carriers.²²

A meta-analysis found that BRCA1 mutation carriers diagnosed with cancer in one breast have a 5-year risk of developing cancer in the other breast of 15%, and BRCA2 mutation carriers have a risk of 9%.²³ Overall, the risk of contralateral breast cancer is about 3% per year.^3,4,24

BRCA1 mutations are strongly associated with triple-negative breast cancers.^1,3,4

Hereditary diffuse gastric cancer

Hereditary diffuse gastric cancer is an autosomal-dominant syndrome associated with mutations in the CDH1 gene, although up to 75% of patients with this syndrome do not have an identifiable CDH1 mutation.^9,25,26 In cases in which there is no identifiable CDH1 mutation, the diagnosis is made on the basis of the patient’s medical and family history.

Hereditary diffuse gastric cancer is associated with an increased risk of the lobular subtype of breast cancer as well as diffuse gastric cancer. The cumulative lifetime risk of breast cancer in women with CDH1 mutations is 39% to 52%,^6,9–11,25 and their lifetime risk of diffuse gastric cancer is 83%.⁹ The combined risk of breast cancer and gastric cancer in women with this syndrome is 90% by age 80.⁹

Cowden syndrome (PTEN hamartoma tumor syndrome)

Cowden syndrome (PTEN hamartoma tumor syndrome) is caused by mutations in PTEN, another tumor-suppressor gene.¹¹ The primary clinical concerns are melanoma and breast, endometrial, thyroid (follicular or papillary), colon, and renal cell cancers. Women with a PTEN mutation have a twofold greater risk of developing any type of cancer than men with a PTEN mutation.¹² The cumulative lifetime risk of invasive breast cancer in women with this syndrome is 70% to 85%.^11–13

Peutz-Jeghers syndrome

Peutz-Jeghers syndrome is an autosomal dominant polyposis disorder caused, in most patients, by a mutation in the serine/threonine kinase tumor-suppressor gene STK11.¹⁴

Patients with Peutz-Jeghers syndrome have higher risks of gastrointestinal, breast, gynecologic (uterine, ovarian, and cervical), pancreatic, and lung cancers. In women, the lifetime risk of breast cancer is 44% to 50% by age 70, regardless of the type of mutation.^6,14,15 Breast cancers associated with Peutz-Jeghers syndrome are usually ductal, and the mean age at diagnosis is 37 years.¹⁶

Li-Fraumeni syndrome

Li-Fraumeni syndrome is an autosomal-dominant disorder caused by germline mutations in the TP53 gene, which codes for a transcription factor associated with cell proliferation and apoptosis.²⁷

Think about other highly penetrant genetic mutations in a young breast cancer patient with no mutation in BRCA1 or BRCA2These mutations confer a lifetime cancer risk of 93% in women (mainly breast cancer) and 68% in men.^1,27 Other cancers associated with TP53 mutations include sarcomas, brain cancer, leukemia, and adrenocortical tumors. Germline TP53 mutations are responsible for approximately 1% of all breast cancers.^1,4

Breast cancers can occur at a young age in patients with a TP53 mutation. Women with TP53 mutations are 18 times more likely to develop breast cancer before age 45 compared with the general population.⁴

It is important to consider a TP53 mutation in premenopausal women or women less than 30 years of age with breast cancer who have no mutations in BRCA1 and BRCA2.¹

MANAGING PATIENTS WITH GENETIC PREDISPOSITION TO BREAST CANCER

Management for patients at high risk fall into three broad categories: clinical surveillance, chemoprevention, and surgical risk reduction. The utility and benefit of each depend to a large degree on the patient’s specific mutation, family history, and comorbidities. Decisions must be shared with the patient.

CLOSE CLINICAL SURVEILLANCE

Consensus guidelines for cancer screening in the syndromes described here are available from the National Comprehensive Cancer Network at www.nccn.org and are summarized in Table 2.^26,28 While the guidelines are broadly applicable to all women with these conditions, some individualization is required based on personal and family medical history.

In general, screening begins at the ages listed in Table 2 or 10 years earlier than the age at which cancer developed in the first affected relative, whichever is earlier. However, screening decisions are shared with the patient and are sometimes affected by significant out-of-pocket costs for the patient and anxiety resulting from the test or subsequent test findings, which must all be considered.

Breast self-awareness and clinical breast examination

Although controversial in the general population, breast self-examination is recommended for patients carrying mutations that increase risk.⁶

Discuss breast self-awareness with all women patients at age 18

A discussion about breast self-awareness is recommended for all women at the age of 18. It should include the signs and symptoms of breast cancer, what feels “normal” to the patient, and what is known about modifiable risk factors for breast cancer. The patient should also be told to report any changes in her personal or family history.

Clinical breast examinations should be done every 6 months, as some cancers are found clinically, particularly in young women with dense tissue, and confirmed by diagnostic imaging and targeted ultrasonography.

Radiographic surveillance

Mammography and magnetic resonance imaging (MRI) are also important components of a breast cancer surveillance regimen in women at high risk. Adherence to a well-formulated plan of clinical and radiographic examinations increases early detection in patients who have a hereditary predisposition to breast cancer.

MRI is more sensitive than mammography and reduces the likelihood of finding advanced cancers by up to 70% compared with mammography in women at high risk of breast cancer.^29–31 The sensitivity of breast MRI alone ranges from 71% to 100%, and the sensitivity increases to 89% to 100% when combined with mammography. In contrast, the sensitivity of mammography alone is 25% to 59%.²⁹ MRI has also been shown to be cost-effective when added to mammography and physical examination in women at high risk.^5,32

Adding MRI to the breast cancer screening regimen has been under discussion and has been endorsed by the American Cancer Society in formal recommendations set forth in 2007 for patients with known hereditary cancer syndromes, in untested first-degree relatives of identified genetic mutation carriers, or in women who have an estimated lifetime risk of breast cancer of 20% or more, as determined by models largely dependent on family history.³³

But MRI has a downside—it is less specific than mammography.^29,33 Its lower specificity (77% to 90% vs 95% with mammography alone) leads to additional radiographic studies and tissue samplings for the “suspicious” lesions discovered. From 3% to 15% of screening breast MRIs result in a biopsy, and the proportion of biopsies that reveal cancer is 13% to 40%.³³ Furthermore, by itself, MRI has not been shown to reduce mortality in any high-risk group.

Mammography remains useful in conjunction with MRI due to its ability to detect breast calcifications, which may be the earliest sign of breast cancer, and ability to detect changes in breast architecture. A typical screening program (Table 2) should incorporate both modalities, commonly offset by 6 months (eg, mammography at baseline, then MRI 6 months later, then mammography again 6 months after that, and so on) to increase the detection of interval cancer development.

Chemoprevention

Chemoprevention means taking medications to reduce the risk. Certain selective estrogen receptor modulators and aromatase inhibitors decrease the risk of invasive breast cancer in healthy women at high risk. These drugs include tamoxifen, which can be used before menopause, and raloxifene, anastrozole, and exemestane, which must be used only after menopause.

Because data are limited, we cannot make any generalized recommendations about chemoprevention in patients with hereditary breast cancer syndromes. Decisions about chemoprevention should take into account the patient’s personal and family histories. Often, a medical oncologist or medical breast specialist can help by discussing the risks and benefits for the individual patient.

Tamoxifen has been the most studied, mainly in BRCA mutation carriers.^6,34–37 As in the general population, tamoxifen reduces the incidence of estrogen receptor-positive breast cancers by 50%.^36–38 It has not been shown to significantly reduce breast cancer risk in premenopausal women with BRCA1 mutations,³⁷ most likely because most cancers that occur in this group are estrogen receptor-negative. In patients with a history of breast cancer, however, tamoxifen has been shown to reduce the risk of developing contralateral breast cancer by 45% to 60% in both BRCA1 and BRCA2 mutation carriers.^6,35

There is also little evidence that giving a chemopreventive agent after bilateral salpingo-oophorectomy reduces the risk further in premenopausal BRCA mutation carriers.³⁵ These patients often receive hormonal therapy with estrogen, which currently would preclude the use of tamoxifen. Tamoxifen in postmenopausal women is associated with a small increased risk of venous thromboembolic disease and endometrial cancer.³⁸

Oral contraceptives reduce the risk of ovarian cancer by up to 50% in BRCA1 mutation carriers and up to 60% in BRCA2 mutation carriers.⁶ However, data conflict on their effect on the risk of breast cancer in BRCA1 and BRCA2 mutation carriers.³⁹

Decisions about chemoprevention with agents other than tamoxifen and in syndromes other than hereditary breast and ovarian cancer syndrome must take into consideration the existing lack of data in this area.

SURGICAL PROPHYLAXIS

Surgical prophylactic options for patients at genetic risk of breast cancer are bilateral mastectomy and bilateral salpingo-oophorectomy.

Prophylactic mastectomy

Bilateral risk-reducing mastectomy reduces the risk of breast cancer by at least 90%^24,39,40 and greatly reduces the need for complex surveillance. Patients are often followed annually clinically, with single-view mammography if they have tissue flap reconstruction.

Nipple-sparing and skin-sparing mastectomies, which facilitate reconstruction and cosmetic outcomes, are an option in the risk-reduction setting and have been shown thus far to be safe.^41–43 In patients with breast cancer, the overall breast cancer recurrence rates with nipple-sparing mastectomy are similar to those of traditional mastectomy and breast conservation treatment.⁴¹

In patients at very high risk of breast cancer, risk-reducing operations also reduce the risk of ultimately needing chemotherapy and radiation to treat breast cancer, as the risk of developing breast cancer is significantly lowered.

The timing of risk-reducing mastectomy depends largely on personal and family medical history and personal choice. Bilateral mastectomy at age 25 results in the greatest survival gain for patients with hereditary breast and ovarian cancer syndrome.⁵ Such precise data are not available for other hereditary cancer syndromes, but it is reasonable to consider bilateral mastectomy as an option for any woman with a highly penetrant genetic mutation that predisposes her to breast cancer. Special consideration in the timing of risk-reducing mastectomy must be given to women with Li-Fraumeni syndrome, as this condition is often associated with an earlier age at breast cancer diagnosis (before age 30).¹

Family planning, sexuality, self-image, and the anxiety associated with both cancer risk and surveillance are all factors women consider when deciding whether and when to undergo mastectomy. A survey of 12 high-risk women who elected prophylactic mastectomy elicited feelings of some regret in 3 of them, while all expressed a sense of relief and reduced anxiety related to both cancer risk and screenings.²⁴ Another group of 14 women surveyed after the surgery reported initial distress related to physical appearance, self-image, and intimacy but also reported a significant decrease in anxiety related to breast cancer risk and were largely satisfied with their decision.⁴⁴

Prophylactic salpingo-oophorectomy

In patients who have pathogenic mutations in BRCA1 or 2, prophylactic salpingo-oophorectomy before age 40 decreases the risk of ovarian cancer by up to 96% and breast cancer by 50%.^1,37,45 This operation, in fact, is the only intervention that has been shown to reduce the mortality rate in patients with a hereditary predisposition to cancer.⁴⁶

We recommend that women with hereditary breast and ovarian cancer syndrome strongly consider prophylactic salpingo-oophorectomy by age 40 or when childbearing is complete for the greatest reduction in risk.^1,5 In 2006, Domchek et al⁴⁶ reported an overall decrease in the mortality rate in BRCA1/2-positive patients who underwent this surgery, but not in breast cancer-specific or ovarian cancer-specific mortality.

On the other hand, removing the ovaries before menopause places women at risk of serious complications associated with premature loss of gonadal hormones, including cardiovascular disease, decreased bone density, reduced sexual satisfaction, dyspareunia, hot flashes, and night sweats.⁴⁷ Therefore, it is generally reserved for women who are also at risk of ovarian cancer.

Hormonal therapy, ie, estrogen therapy for patients who choose complete hysterectomy, and estrogen-progesterone therapy for patients who choose to keep their uterus, reduces menopausal symptoms and symptoms of sexual dissatisfaction and has not thus far been shown to increase breast cancer risk.^1,34 However, this information is from nonrandomized studies, which are inherently limited.

It is important to address and modify risk factors for heart disease and osteoporosis in women with premature surgical menopause, as they may be particularly vulnerable to these conditions.

HEREDITARY BREAST CANCER IN MEN

Fewer than 1% of cases of breast cancer arise in men, and fewer than 1% of cases of cancer in men are breast cancer.

Male breast cancer is more likely than female breast cancer to be estrogen receptor- and progesterone receptor-positive. In an analysis of the Surveillance, Epidemiology, and End Results registry between 1973 and 2005, triple-negative breast cancer was found in 23% of female patients but only 7.6% of male patients.²

Male breast cancer is most common in families with BRCA2, and to a lesser degree, BRCA1 mutations. Other genetic disorders including Li-Fraumeni syndrome, hereditary nonpolyposis colorectal cancer, and Klinefelter syndrome also increase the risk of male breast cancer. A genetic predisposition for breast cancer is present in approximately 10% of male breast cancer patients.² Any man with breast cancer, therefore, should be referred for genetic counseling.

In men, a BRCA2 mutation confers a lifetime risk of breast cancer of 5% to 10%.² This is similar to the lifetime risk of breast cancer for the average woman but it is still significant, as the lifetime risk of breast cancer for the average man is 0.1%.^1,2

Five-year survival rates in male breast cancer range from only 36% to 66%, most likely because it is usually diagnosed in later stages, as men are not routinely screened for breast cancer. In men with known hereditary susceptibility, National Comprehensive Cancer Network guidelines recommend that they be educated about and begin breast self-examination at the age of 35 and be clinically examined every 12 months starting at age 35.⁴⁸ There are limited data to support breast imaging in men. High-risk surveillance with MRI screening in this group is not recommended. Prostate cancer screening is recommended for men with BRCA2 mutations starting at age 40, and should be considered for men with BRCA1 mutations starting at age 40.

No specific guidelines exist for pancreatic cancer and melanoma, but screening may be individualized based on cancers observed in the family.

A 25-year-old woman presents to the emergency department with a 7-day history of fatigue and nausea. On presentation she denies having abdominal pain, headache, fever, chills, night sweats, vomiting, diarrhea, melena, hematochezia, or weight loss. She recalls changes in the colors of her eyes and darkening urine over the last few days. Her medical history before this is unremarkable. She takes no prescription, over-the-counter, or herbal medications. She works as a librarian and has no occupational toxic exposures. She is single and has one sister with no prior medical history. She denies recent travel, sick contacts, smoking, recreational drug use, or pets at home.

On physical examination, her vital signs are temperature 37.3°C (99.1°F), heart rate 90 beats per minute, blood pressure 125/80 mm Hg, respiration rate 14 per minute, and oxygen saturation 97% on room air. She has icteric sclera and her skin is jaundiced. Cardiac examination is normal. Lungs are clear to auscultation and percussion bilaterally. Her abdomen is soft with no visceromegaly, masses, or tenderness. Extremities are normal with no edema. She is alert and oriented, but she has mild asterixis of the outstretched hands. The neurologic examination is otherwise unremarkable.

The patient’s basic laboratory values are listed in Table 1. Shortly after admission, she develops changes in her mental status, remaining alert but becoming agitated and oriented to person only. In view of her symptoms and laboratory findings, acute liver failure is suspected.

ACUTE LIVER FAILURE

1. The diagnostic criteria for acute liver failure include all of the following except which one?

• Acute elevation of liver biochemical tests
• Presence of preexisting liver disease
• Coagulopathy, defined by an international normalized ratio (INR) of 1.5 or greater
• Encephalopathy
• Duration of symptoms less than 26 weeks

Acute liver failure is defined by acute onset of worsening liver tests, coagulopathy (INR ≥ 1.5), and encephalopathy in patients with no preexisting liver disease and with symptom duration of less than 26 weeks.¹ With a few exceptions, a history of preexisting liver disease negates the diagnosis of acute liver failure. Our patient meets the diagnostic criteria for acute liver failure.

Immediate management

Once acute liver failure is identified or suspected, the next step is to transfer the patient to the intensive care unit for close monitoring of mental status. Serial neurologic evaluations permit early detection of cerebral edema, which is considered the most common cause of death in patients with acute liver failure. Additionally, close monitoring of electrolytes and plasma glucose is necessary since these patients are susceptible to electrolyte disturbances and hypoglycemia.

Patients with acute liver failure are at increased risk of infections and should be routinely screened by obtaining urine and blood cultures.

Gastrointestinal bleeding is not uncommon in patients with acute liver failure and is usually due to gastric stress ulceration. Prophylaxis with a histamine 2 receptor antagonist or proton pump inhibitor should be considered in order to prevent gastrointestinal bleeding.

Treatment with N-acetylcysteine is beneficial, not only in patients with acute liver failure due to acetaminophen overdose, but also in those with acute liver failure from other causes.

CASE CONTINUES:
TRANSFER TO THE INTENSIVE CARE UNIT

The patient, now diagnosed with acute liver failure, is transferred to the intensive care unit. Arterial blood gas measurement shows:

• pH 7.38 (reference range 7.35–7.45)
• Pco₂ 40 mm Hg (36–46)
• Po₂ 97 mm Hg (85–95)
• Hco₃ 22 mmol/L (22–26).

A chest radiograph is obtained and is clear. Computed tomography (CT) of the brain reveals no edema. Transcranial Doppler ultrasonography does not show any intracranial fluid collections.

Blood and urine cultures are negative. Her hemoglobin level remains stable, and she does not develop signs of bleeding. She is started on a proton pump inhibitor for stress ulcer prophylaxis and is empirically given intravenous N-acetylcysteine until the cause of acute liver failure can be determined.

CAUSES OF ACUTE LIVER FAILURE

2. Which of the following can cause acute liver failure?

• Acetaminophen overdose
• Viral hepatitis
• Autoimmune hepatitis
• Wilson disease
• Alcoholic hepatitis

Drug-induced liver injury is the most common cause of acute liver failure in the United States,^2,3 and of all drugs, acetaminophen overdose is the number-one cause. In acetaminophen-induced liver injury, serum aminotransferase levels are usually elevated to more than 1,000 U/L, while serum bilirubin remains normal in the early stages. Antimicrobial agents, antiepileptic drugs, and herbal supplements have also been implicated in acute liver failure. Our patient has denied taking herbal supplements or medications, including over-the-counter ones.

Acute viral hepatitis can explain the patient’s condition. It is a common cause of acute liver failure in the United States.² Hepatitis A and E are more common in developing countries. Other viruses such as cytomegalovirus, Epstein-Barr virus, herpes simplex virus type 1 and 2, and varicella zoster virus can also cause acute liver failure. Serum aminotransferase levels may exceed 1,000 U/L in patients with viral hepatitis.

A young woman presents with acute liver failure: What is the cause? Is her sister at risk?

Autoimmune hepatitis is a rare cause of acute liver failure, but it should be considered in the differential diagnosis, particularly in middle-aged women with autoimmune disorders such as hypothyroidism. Autoimmune hepatitis can cause marked elevation in aminotransferase levels (> 1,000 U/L).

Wilson disease is an autosomal-recessive disease in which there is excessive accumulation of copper in the liver and other organs because of an inherited defect in the biliary excretion of copper. Wilson disease can cause acute liver failure and should be excluded in any patient, particularly if under age 40 with acute onset of unexplained hepatic, neurologic, or psychiatric disease.

Alcoholic hepatitis usually occurs in patients with a long-standing history of heavy alcohol use. As a result, most patients with alcoholic hepatitis have manifestations of chronic liver disease due to alcohol use. Therefore, by definition, it is not a cause of acute liver failure. Additionally, in patients with alcoholic hepatitis, the aspartate aminotransferase (AST) level is elevated but less than 300 IU/mL, and the ratio of AST to alanine aminotransferase (ALT) is usually more than 2.

CASE CONTINUES: FURTHER TESTING

The results of our patient’s serologic tests are shown in Table 2. Other test results:

• Autoimmune markers including antinuclear antibodies, antimitochondrial antibodies, antismooth muscle antibodies, and liver and kidney microsomal antibodies are negative; her immunoglobulin G (IgG) level is normal
• Serum ceruloplasmin 25 mg/dL (normal 21–45)
• Free serum copper 120 µg/dL (normal 8–12)
• Abdominal ultrasonography is unremarkable, with normal liver parenchyma and no intrahepatic or extrahepatic biliary dilatation
• Doppler ultrasonography of the liver shows patent blood vessels.

3. Based on the new data, which of the following statements is correct?

• Hepatitis B is the cause of acute liver failure in this patient
• Herpetic hepatitis cannot be excluded on the basis of the available data
• Wilson disease is most likely the diagnosis, given her elevated free serum copper
• A normal serum ceruloplasmin level is not sufficient to rule out acute liver failure secondary to Wilson disease

Hepatitis B surface antigen and hepatitis B core antibodies were negative in our patient, excluding hepatitis B virus infection. The positive hepatitis B surface antibody indicates prior immunization.

Herpetic hepatitis is an uncommon but important cause of acute liver failure because the mortality rate is high if the patient is not treated early with acyclovir. Fever, elevated aminotransferases, and leukopenia are common with herpetic hepatitis. Fewer than 50% of patients with herpetic hepatitis have vesicular rash.^4,5 The value of antibody serologic testing is limited due to high rates of false-positive and false-negative results. The gold standard diagnostic tests are viral load (detection of viral RNA by polymerase chain reaction), viral staining on liver biopsy, or both. In our patient, herpes simplex virus polymerase chain reaction testing was negative, which makes herpetic hepatitis unlikely.

Wilson disease is a genetic condition in which the ability to excrete copper in the bile is impaired, resulting in accumulation of copper in the hepatocytes. Subsequently, copper is released into the bloodstream and eventually into the urine.

However, copper excretion into the bile is impaired in patients with acute liver failure regardless of the etiology. Therefore, elevated free serum copper and 24-hour urine copper levels are not specific for the diagnosis of acute liver failure secondary to Wilson disease. Moreover, Kayser-Fleischer rings, which represent copper deposition in the limbus of the cornea, may not be apparent in the early stages of Wilson disease.

Wilson disease involves accumulation of copper in the liver and other organs as the result of a genetic defect

Since it is challenging to diagnose Wilson disease in the context of acute liver failure, Korman et al⁶ compared patients with acute liver failure secondary to Wilson disease with patients with acute liver failure secondary to other conditions. They found that alkaline phosphatase levels are frequently decreased in patients with acute liver failure secondary to Wilson disease,⁶ and that a ratio of alkaline phosphatase to total bilirubin of less than 4 is 94% sensitive and 96% specific for the diagnosis.⁶

Hemolysis is common in acute liver failure due to Wilson disease. This leads to disproportionate elevation of AST compared with ALT, since AST is present in red blood cells. Consequently, the ratio of AST to ALT is usually greater than 2.2, which provides a sensitivity of 94% and a specificity of 86% for the diagnosis.⁶ These two ratios together provide 100% sensitivity and 100% specificity for the diagnosis of Wilson disease in the context of acute liver failure.⁶

Ceruloplasmin. Patients with Wilson disease typically have a low ceruloplasmin level. However, because it is an acute-phase reaction protein, ceruloplasmin can be normal or elevated in patients with acute liver failure from Wilson disease.⁶ Therefore, a normal ceruloplasmin level is not sufficient to rule out acute liver failure secondary to Wilson disease.

CASE CONTINUES: A DEFINITIVE DIAGNOSIS

Our patient undergoes further testing, which reveals the following:

• Her 24-hour urinary excretion of copper is 150 µg (reference value < 30)
• Slit-lamp examination is normal and shows no evidence of Kayser-Fleischer rings
• Her ratio of alkaline phosphatase to total bilirubin is 0.77 based on her initial laboratory results (Table 1)
• Her AST-ALT ratio is 3.4.

The diagnosis in our patient is acute liver failure secondary to Wilson disease.

4. What is the most appropriate next step?

• Liver biopsy
• d-penicillamine by mouth
• Trientine by mouth
• Liver transplant
• Plasmapheresis

Liver biopsy. Accumulation of copper in the liver parenchyma in patients with Wilson disease is sporadic. Therefore, qualitative copper staining on liver biopsy can be falsely negative. Quantitative copper measurement in liver tissue is the gold standard for the diagnosis of Wilson disease. However, the test is time-consuming and is not rapidly available in the context of acute liver failure.

Chelating agents such as d-pencillamine and trientine are used to treat the chronic manifestations of Wilson disease but are not useful for acute liver failure secondary to Wilson disease.

Acute liver failure secondary to Wilson disease is life-threatening, and liver transplant is considered the only definitive life-saving therapy.

Therapeutic plasmapheresis has been reported to be a successful adjunctive therapy to bridge patients with acute liver failure secondary to Wilson disease to transplant.⁷ However, liver transplant is still the only definitive treatment.

CASE CONTINUES: THE PATIENT’S SISTER SEEKS CARE

The patient undergoes liver transplantation, with no perioperative or postoperative complications.

The patient’s 18-year-old sister is now seeking medical attention in the outpatient clinic, concerned that she may have Wilson disease. She is otherwise healthy and denies any symptoms or complaints.

5. What is the next step for the patient’s sister?

• Reassurance
• Prophylaxis with trientine
• Check liver enzyme levels, serum ceruloplasmin level, and urine copper, and order a slit-lamp examination
• Genetic testing

Wilson disease can be asymptomatic in its early stages and may be diagnosed incidentally during routine blood tests that reveal abnormal liver enzyme levels. All patients with a confirmed family history of Wilson disease should be screened even if they are asymptomatic. The diagnosis of Wilson disease should be established in first-degree relatives before specific treatment for the relatives is prescribed.

Based on information in Roberts EA, Schilsky ML; American Association for Study of Liver Diseases (AASLD). Diagnosis and treatment of Wilson disease: an update. Hepatology 2008; 7:2089–2111.

The first step in screening a first-degree relative for Wilson disease is to check liver enzyme levels (specifically aminotransferases, alkaline phosphatase, and bilirubin), serum ceruloplasmin level, and 24-hour urine copper, and order an ophthalmologic slit-lamp examination. If any of these tests is abnormal, liver biopsy should be performed for histopathologic evaluation and quantitative copper measurement. Kayser-Fleischer  rings are seen in only 50% of patients with Wilson disease and hepatic involvement, but they are pathognomic. Guidelines⁸ for screening first-degree relatives of Wilson disease patients are shown in Figure 1.

Genetic analysis. ATP7B, the Wilson disease gene, is located on chromosome 13. At least 300 mutations of the gene have been described,² and the most common mutation is present in only 15% to 30% of the Wilson disease population.^8–10 Routine molecular testing of the ATP7B

CASE CONTINUES: WORKUP OF THE PATIENT’S SISTER

The patient’s sister has no symptoms and her physical examination is normal. Slit-lamp examination reveals no evidence of Kayser-Fleischer rings. Her laboratory values, including complete blood counts, complete metabolic panel, and INR, are within normal ranges. Other test results, however, are abnormal:

• Free serum copper level 27 µg/dL (normal 8–12)
• Serum ceruloplasmin 9.0 mg/dL (normal 20–50)
• 24-hour urinary copper excretion 135 µg (normal < 30).

She undergoes liver biopsy for quantitative copper measurement, and the result is very high at 1,118 µg/g dry weight (reference range 10–35). The diagnosis of Wilson disease is established.

TREATING CHRONIC WILSON DISEASE

6. Which of the following is not an appropriate next step for the patient’s sister?

• Tetrathiomolybdate
• d-penicillamine
• Trientine
• Zinc salts
• Prednisone

The goal of medical treatment of chronic Wilson disease is to improve symptoms and prevent progression of the disease.

Chelating agents and zinc salts are the most commonly used medicines in the management of Wilson disease. Chelating agents remove copper from tissue, whereas zinc blocks the intestinal absorption of copper and stimulates the synthesis of endogenous chelators such as metallothioneins. Tetrathiomolybdate is an alternative agent developed to interfere with the distribution of excess body copper to susceptible target sites by reducing free serum copper (Table 3). There are no data to support the use of prednisone in the treatment of Wilson disease.

During treatment with chelating agents, 24-hour urinary excretion of copper is routinely monitored to determine the efficacy of therapy and adherence to treatment. Once de-coppering is achieved, as evidenced by a normalization of 24-hour urine copper excretion, the chelating agent can be switched to zinc salts to prevent intestinal absorption of copper.

Clinical and biochemical stabilization is achieved typically within 2 to 6 months of the initial treatment with chelating agents.⁸ Organ meats, nuts, shellfish, and chocolate are rich in copper and should be avoided.

The patient’s sister is started on trientine 250 mg orally three times daily on an empty stomach at least 1 hour before meals. Treatment is monitored by following 24-hour urine copper measurement. A 24-hour urine copper measurement at 3 months after starting treatment has increased from 54 at baseline to 350 µg, which indicates that the copper is being removed from tissues. The plan is for early substitution of zinc for long-term maintenance once de-coppering is completed.

KEY POINTS

• Acute liver failure is severe acute liver injury characterized by coagulopathy (INR ≥ 1.5) and encephalopathy in a patient with no preexisting liver disease and with duration of symptoms less than 26 weeks.
• Acute liver failure secondary to Wilson disease is uncommon but should be excluded, particularly in young patients.
• The diagnosis of Wilson disease in the setting of acute liver failure is challenging because the serum ceruloplasmin level may be normal in acute liver failure secondary to Wilson disease, and free serum copper and 24-hour urine copper are usually elevated in all acute liver failure patients regardless of the etiology.
• A ratio of alkaline phosphatase to total bilirubin of less than 4 plus an AST-ALT ratio greater than 2.2 in a patient with acute liver failure should be regarded as Wilson disease until proven otherwise (Figure 2).
• Acute liver failure secondary to Wilson disease is usually fatal, and emergency liver transplant is a life-saving procedure.
• Screening of first-degree relatives of Wilson disease patients should include a history and physical examination, liver enzyme tests, complete blood cell count, serum ceruloplasmin level, serum free copper level, slit-lamp examination of the eyes, and 24-hour urinary copper measurement. Genetic tests are supplementary for screening but are not routinely available.

A 25-year-old woman presents to the emergency department with a 7-day history of fatigue and nausea. On presentation she denies having abdominal pain, headache, fever, chills, night sweats, vomiting, diarrhea, melena, hematochezia, or weight loss. She recalls changes in the colors of her eyes and darkening urine over the last few days. Her medical history before this is unremarkable. She takes no prescription, over-the-counter, or herbal medications. She works as a librarian and has no occupational toxic exposures. She is single and has one sister with no prior medical history. She denies recent travel, sick contacts, smoking, recreational drug use, or pets at home.

On physical examination, her vital signs are temperature 37.3°C (99.1°F), heart rate 90 beats per minute, blood pressure 125/80 mm Hg, respiration rate 14 per minute, and oxygen saturation 97% on room air. She has icteric sclera and her skin is jaundiced. Cardiac examination is normal. Lungs are clear to auscultation and percussion bilaterally. Her abdomen is soft with no visceromegaly, masses, or tenderness. Extremities are normal with no edema. She is alert and oriented, but she has mild asterixis of the outstretched hands. The neurologic examination is otherwise unremarkable.

The patient’s basic laboratory values are listed in Table 1. Shortly after admission, she develops changes in her mental status, remaining alert but becoming agitated and oriented to person only. In view of her symptoms and laboratory findings, acute liver failure is suspected.

ACUTE LIVER FAILURE

1. The diagnostic criteria for acute liver failure include all of the following except which one?

• Acute elevation of liver biochemical tests
• Presence of preexisting liver disease
• Coagulopathy, defined by an international normalized ratio (INR) of 1.5 or greater
• Encephalopathy
• Duration of symptoms less than 26 weeks

Acute liver failure is defined by acute onset of worsening liver tests, coagulopathy (INR ≥ 1.5), and encephalopathy in patients with no preexisting liver disease and with symptom duration of less than 26 weeks.¹ With a few exceptions, a history of preexisting liver disease negates the diagnosis of acute liver failure. Our patient meets the diagnostic criteria for acute liver failure.

Immediate management

Once acute liver failure is identified or suspected, the next step is to transfer the patient to the intensive care unit for close monitoring of mental status. Serial neurologic evaluations permit early detection of cerebral edema, which is considered the most common cause of death in patients with acute liver failure. Additionally, close monitoring of electrolytes and plasma glucose is necessary since these patients are susceptible to electrolyte disturbances and hypoglycemia.

Patients with acute liver failure are at increased risk of infections and should be routinely screened by obtaining urine and blood cultures.

Gastrointestinal bleeding is not uncommon in patients with acute liver failure and is usually due to gastric stress ulceration. Prophylaxis with a histamine 2 receptor antagonist or proton pump inhibitor should be considered in order to prevent gastrointestinal bleeding.

Treatment with N-acetylcysteine is beneficial, not only in patients with acute liver failure due to acetaminophen overdose, but also in those with acute liver failure from other causes.

CASE CONTINUES:
TRANSFER TO THE INTENSIVE CARE UNIT

The patient, now diagnosed with acute liver failure, is transferred to the intensive care unit. Arterial blood gas measurement shows:

• pH 7.38 (reference range 7.35–7.45)
• Pco₂ 40 mm Hg (36–46)
• Po₂ 97 mm Hg (85–95)
• Hco₃ 22 mmol/L (22–26).

A chest radiograph is obtained and is clear. Computed tomography (CT) of the brain reveals no edema. Transcranial Doppler ultrasonography does not show any intracranial fluid collections.

Blood and urine cultures are negative. Her hemoglobin level remains stable, and she does not develop signs of bleeding. She is started on a proton pump inhibitor for stress ulcer prophylaxis and is empirically given intravenous N-acetylcysteine until the cause of acute liver failure can be determined.

CAUSES OF ACUTE LIVER FAILURE

2. Which of the following can cause acute liver failure?

• Acetaminophen overdose
• Viral hepatitis
• Autoimmune hepatitis
• Wilson disease
• Alcoholic hepatitis

Drug-induced liver injury is the most common cause of acute liver failure in the United States,^2,3 and of all drugs, acetaminophen overdose is the number-one cause. In acetaminophen-induced liver injury, serum aminotransferase levels are usually elevated to more than 1,000 U/L, while serum bilirubin remains normal in the early stages. Antimicrobial agents, antiepileptic drugs, and herbal supplements have also been implicated in acute liver failure. Our patient has denied taking herbal supplements or medications, including over-the-counter ones.

Acute viral hepatitis can explain the patient’s condition. It is a common cause of acute liver failure in the United States.² Hepatitis A and E are more common in developing countries. Other viruses such as cytomegalovirus, Epstein-Barr virus, herpes simplex virus type 1 and 2, and varicella zoster virus can also cause acute liver failure. Serum aminotransferase levels may exceed 1,000 U/L in patients with viral hepatitis.

A young woman presents with acute liver failure: What is the cause? Is her sister at risk?

Autoimmune hepatitis is a rare cause of acute liver failure, but it should be considered in the differential diagnosis, particularly in middle-aged women with autoimmune disorders such as hypothyroidism. Autoimmune hepatitis can cause marked elevation in aminotransferase levels (> 1,000 U/L).

Wilson disease is an autosomal-recessive disease in which there is excessive accumulation of copper in the liver and other organs because of an inherited defect in the biliary excretion of copper. Wilson disease can cause acute liver failure and should be excluded in any patient, particularly if under age 40 with acute onset of unexplained hepatic, neurologic, or psychiatric disease.

Alcoholic hepatitis usually occurs in patients with a long-standing history of heavy alcohol use. As a result, most patients with alcoholic hepatitis have manifestations of chronic liver disease due to alcohol use. Therefore, by definition, it is not a cause of acute liver failure. Additionally, in patients with alcoholic hepatitis, the aspartate aminotransferase (AST) level is elevated but less than 300 IU/mL, and the ratio of AST to alanine aminotransferase (ALT) is usually more than 2.

CASE CONTINUES: FURTHER TESTING

The results of our patient’s serologic tests are shown in Table 2. Other test results:

• Autoimmune markers including antinuclear antibodies, antimitochondrial antibodies, antismooth muscle antibodies, and liver and kidney microsomal antibodies are negative; her immunoglobulin G (IgG) level is normal
• Serum ceruloplasmin 25 mg/dL (normal 21–45)
• Free serum copper 120 µg/dL (normal 8–12)
• Abdominal ultrasonography is unremarkable, with normal liver parenchyma and no intrahepatic or extrahepatic biliary dilatation
• Doppler ultrasonography of the liver shows patent blood vessels.

3. Based on the new data, which of the following statements is correct?

• Hepatitis B is the cause of acute liver failure in this patient
• Herpetic hepatitis cannot be excluded on the basis of the available data
• Wilson disease is most likely the diagnosis, given her elevated free serum copper
• A normal serum ceruloplasmin level is not sufficient to rule out acute liver failure secondary to Wilson disease

Hepatitis B surface antigen and hepatitis B core antibodies were negative in our patient, excluding hepatitis B virus infection. The positive hepatitis B surface antibody indicates prior immunization.

Herpetic hepatitis is an uncommon but important cause of acute liver failure because the mortality rate is high if the patient is not treated early with acyclovir. Fever, elevated aminotransferases, and leukopenia are common with herpetic hepatitis. Fewer than 50% of patients with herpetic hepatitis have vesicular rash.^4,5 The value of antibody serologic testing is limited due to high rates of false-positive and false-negative results. The gold standard diagnostic tests are viral load (detection of viral RNA by polymerase chain reaction), viral staining on liver biopsy, or both. In our patient, herpes simplex virus polymerase chain reaction testing was negative, which makes herpetic hepatitis unlikely.

Wilson disease is a genetic condition in which the ability to excrete copper in the bile is impaired, resulting in accumulation of copper in the hepatocytes. Subsequently, copper is released into the bloodstream and eventually into the urine.

However, copper excretion into the bile is impaired in patients with acute liver failure regardless of the etiology. Therefore, elevated free serum copper and 24-hour urine copper levels are not specific for the diagnosis of acute liver failure secondary to Wilson disease. Moreover, Kayser-Fleischer rings, which represent copper deposition in the limbus of the cornea, may not be apparent in the early stages of Wilson disease.

Wilson disease involves accumulation of copper in the liver and other organs as the result of a genetic defect

Since it is challenging to diagnose Wilson disease in the context of acute liver failure, Korman et al⁶ compared patients with acute liver failure secondary to Wilson disease with patients with acute liver failure secondary to other conditions. They found that alkaline phosphatase levels are frequently decreased in patients with acute liver failure secondary to Wilson disease,⁶ and that a ratio of alkaline phosphatase to total bilirubin of less than 4 is 94% sensitive and 96% specific for the diagnosis.⁶

Hemolysis is common in acute liver failure due to Wilson disease. This leads to disproportionate elevation of AST compared with ALT, since AST is present in red blood cells. Consequently, the ratio of AST to ALT is usually greater than 2.2, which provides a sensitivity of 94% and a specificity of 86% for the diagnosis.⁶ These two ratios together provide 100% sensitivity and 100% specificity for the diagnosis of Wilson disease in the context of acute liver failure.⁶

Ceruloplasmin. Patients with Wilson disease typically have a low ceruloplasmin level. However, because it is an acute-phase reaction protein, ceruloplasmin can be normal or elevated in patients with acute liver failure from Wilson disease.⁶ Therefore, a normal ceruloplasmin level is not sufficient to rule out acute liver failure secondary to Wilson disease.

CASE CONTINUES: A DEFINITIVE DIAGNOSIS

Our patient undergoes further testing, which reveals the following:

• Her 24-hour urinary excretion of copper is 150 µg (reference value < 30)
• Slit-lamp examination is normal and shows no evidence of Kayser-Fleischer rings
• Her ratio of alkaline phosphatase to total bilirubin is 0.77 based on her initial laboratory results (Table 1)
• Her AST-ALT ratio is 3.4.

The diagnosis in our patient is acute liver failure secondary to Wilson disease.

4. What is the most appropriate next step?

• Liver biopsy
• d-penicillamine by mouth
• Trientine by mouth
• Liver transplant
• Plasmapheresis

Liver biopsy. Accumulation of copper in the liver parenchyma in patients with Wilson disease is sporadic. Therefore, qualitative copper staining on liver biopsy can be falsely negative. Quantitative copper measurement in liver tissue is the gold standard for the diagnosis of Wilson disease. However, the test is time-consuming and is not rapidly available in the context of acute liver failure.

Chelating agents such as d-pencillamine and trientine are used to treat the chronic manifestations of Wilson disease but are not useful for acute liver failure secondary to Wilson disease.

Acute liver failure secondary to Wilson disease is life-threatening, and liver transplant is considered the only definitive life-saving therapy.

Therapeutic plasmapheresis has been reported to be a successful adjunctive therapy to bridge patients with acute liver failure secondary to Wilson disease to transplant.⁷ However, liver transplant is still the only definitive treatment.

CASE CONTINUES: THE PATIENT’S SISTER SEEKS CARE

The patient undergoes liver transplantation, with no perioperative or postoperative complications.

The patient’s 18-year-old sister is now seeking medical attention in the outpatient clinic, concerned that she may have Wilson disease. She is otherwise healthy and denies any symptoms or complaints.

5. What is the next step for the patient’s sister?

• Reassurance
• Prophylaxis with trientine
• Check liver enzyme levels, serum ceruloplasmin level, and urine copper, and order a slit-lamp examination
• Genetic testing

Wilson disease can be asymptomatic in its early stages and may be diagnosed incidentally during routine blood tests that reveal abnormal liver enzyme levels. All patients with a confirmed family history of Wilson disease should be screened even if they are asymptomatic. The diagnosis of Wilson disease should be established in first-degree relatives before specific treatment for the relatives is prescribed.

Based on information in Roberts EA, Schilsky ML; American Association for Study of Liver Diseases (AASLD). Diagnosis and treatment of Wilson disease: an update. Hepatology 2008; 7:2089–2111.

The first step in screening a first-degree relative for Wilson disease is to check liver enzyme levels (specifically aminotransferases, alkaline phosphatase, and bilirubin), serum ceruloplasmin level, and 24-hour urine copper, and order an ophthalmologic slit-lamp examination. If any of these tests is abnormal, liver biopsy should be performed for histopathologic evaluation and quantitative copper measurement. Kayser-Fleischer  rings are seen in only 50% of patients with Wilson disease and hepatic involvement, but they are pathognomic. Guidelines⁸ for screening first-degree relatives of Wilson disease patients are shown in Figure 1.

Genetic analysis. ATP7B, the Wilson disease gene, is located on chromosome 13. At least 300 mutations of the gene have been described,² and the most common mutation is present in only 15% to 30% of the Wilson disease population.^8–10 Routine molecular testing of the ATP7B

CASE CONTINUES: WORKUP OF THE PATIENT’S SISTER

The patient’s sister has no symptoms and her physical examination is normal. Slit-lamp examination reveals no evidence of Kayser-Fleischer rings. Her laboratory values, including complete blood counts, complete metabolic panel, and INR, are within normal ranges. Other test results, however, are abnormal:

• Free serum copper level 27 µg/dL (normal 8–12)
• Serum ceruloplasmin 9.0 mg/dL (normal 20–50)
• 24-hour urinary copper excretion 135 µg (normal < 30).

She undergoes liver biopsy for quantitative copper measurement, and the result is very high at 1,118 µg/g dry weight (reference range 10–35). The diagnosis of Wilson disease is established.

TREATING CHRONIC WILSON DISEASE

6. Which of the following is not an appropriate next step for the patient’s sister?

• Tetrathiomolybdate
• d-penicillamine
• Trientine
• Zinc salts
• Prednisone

The goal of medical treatment of chronic Wilson disease is to improve symptoms and prevent progression of the disease.

Chelating agents and zinc salts are the most commonly used medicines in the management of Wilson disease. Chelating agents remove copper from tissue, whereas zinc blocks the intestinal absorption of copper and stimulates the synthesis of endogenous chelators such as metallothioneins. Tetrathiomolybdate is an alternative agent developed to interfere with the distribution of excess body copper to susceptible target sites by reducing free serum copper (Table 3). There are no data to support the use of prednisone in the treatment of Wilson disease.

During treatment with chelating agents, 24-hour urinary excretion of copper is routinely monitored to determine the efficacy of therapy and adherence to treatment. Once de-coppering is achieved, as evidenced by a normalization of 24-hour urine copper excretion, the chelating agent can be switched to zinc salts to prevent intestinal absorption of copper.

Clinical and biochemical stabilization is achieved typically within 2 to 6 months of the initial treatment with chelating agents.⁸ Organ meats, nuts, shellfish, and chocolate are rich in copper and should be avoided.

The patient’s sister is started on trientine 250 mg orally three times daily on an empty stomach at least 1 hour before meals. Treatment is monitored by following 24-hour urine copper measurement. A 24-hour urine copper measurement at 3 months after starting treatment has increased from 54 at baseline to 350 µg, which indicates that the copper is being removed from tissues. The plan is for early substitution of zinc for long-term maintenance once de-coppering is completed.

KEY POINTS

• Acute liver failure is severe acute liver injury characterized by coagulopathy (INR ≥ 1.5) and encephalopathy in a patient with no preexisting liver disease and with duration of symptoms less than 26 weeks.
• Acute liver failure secondary to Wilson disease is uncommon but should be excluded, particularly in young patients.
• The diagnosis of Wilson disease in the setting of acute liver failure is challenging because the serum ceruloplasmin level may be normal in acute liver failure secondary to Wilson disease, and free serum copper and 24-hour urine copper are usually elevated in all acute liver failure patients regardless of the etiology.
• A ratio of alkaline phosphatase to total bilirubin of less than 4 plus an AST-ALT ratio greater than 2.2 in a patient with acute liver failure should be regarded as Wilson disease until proven otherwise (Figure 2).
• Acute liver failure secondary to Wilson disease is usually fatal, and emergency liver transplant is a life-saving procedure.
• Screening of first-degree relatives of Wilson disease patients should include a history and physical examination, liver enzyme tests, complete blood cell count, serum ceruloplasmin level, serum free copper level, slit-lamp examination of the eyes, and 24-hour urinary copper measurement. Genetic tests are supplementary for screening but are not routinely available.

A 25-year-old woman presents to the emergency department with a 7-day history of fatigue and nausea. On presentation she denies having abdominal pain, headache, fever, chills, night sweats, vomiting, diarrhea, melena, hematochezia, or weight loss. She recalls changes in the colors of her eyes and darkening urine over the last few days. Her medical history before this is unremarkable. She takes no prescription, over-the-counter, or herbal medications. She works as a librarian and has no occupational toxic exposures. She is single and has one sister with no prior medical history. She denies recent travel, sick contacts, smoking, recreational drug use, or pets at home.

On physical examination, her vital signs are temperature 37.3°C (99.1°F), heart rate 90 beats per minute, blood pressure 125/80 mm Hg, respiration rate 14 per minute, and oxygen saturation 97% on room air. She has icteric sclera and her skin is jaundiced. Cardiac examination is normal. Lungs are clear to auscultation and percussion bilaterally. Her abdomen is soft with no visceromegaly, masses, or tenderness. Extremities are normal with no edema. She is alert and oriented, but she has mild asterixis of the outstretched hands. The neurologic examination is otherwise unremarkable.

The patient’s basic laboratory values are listed in Table 1. Shortly after admission, she develops changes in her mental status, remaining alert but becoming agitated and oriented to person only. In view of her symptoms and laboratory findings, acute liver failure is suspected.

ACUTE LIVER FAILURE

1. The diagnostic criteria for acute liver failure include all of the following except which one?

• Acute elevation of liver biochemical tests
• Presence of preexisting liver disease
• Coagulopathy, defined by an international normalized ratio (INR) of 1.5 or greater
• Encephalopathy
• Duration of symptoms less than 26 weeks

Acute liver failure is defined by acute onset of worsening liver tests, coagulopathy (INR ≥ 1.5), and encephalopathy in patients with no preexisting liver disease and with symptom duration of less than 26 weeks.¹ With a few exceptions, a history of preexisting liver disease negates the diagnosis of acute liver failure. Our patient meets the diagnostic criteria for acute liver failure.

Immediate management

Once acute liver failure is identified or suspected, the next step is to transfer the patient to the intensive care unit for close monitoring of mental status. Serial neurologic evaluations permit early detection of cerebral edema, which is considered the most common cause of death in patients with acute liver failure. Additionally, close monitoring of electrolytes and plasma glucose is necessary since these patients are susceptible to electrolyte disturbances and hypoglycemia.

Patients with acute liver failure are at increased risk of infections and should be routinely screened by obtaining urine and blood cultures.

Gastrointestinal bleeding is not uncommon in patients with acute liver failure and is usually due to gastric stress ulceration. Prophylaxis with a histamine 2 receptor antagonist or proton pump inhibitor should be considered in order to prevent gastrointestinal bleeding.

Treatment with N-acetylcysteine is beneficial, not only in patients with acute liver failure due to acetaminophen overdose, but also in those with acute liver failure from other causes.

CASE CONTINUES:
TRANSFER TO THE INTENSIVE CARE UNIT

The patient, now diagnosed with acute liver failure, is transferred to the intensive care unit. Arterial blood gas measurement shows:

• pH 7.38 (reference range 7.35–7.45)
• Pco₂ 40 mm Hg (36–46)
• Po₂ 97 mm Hg (85–95)
• Hco₃ 22 mmol/L (22–26).

A chest radiograph is obtained and is clear. Computed tomography (CT) of the brain reveals no edema. Transcranial Doppler ultrasonography does not show any intracranial fluid collections.

Blood and urine cultures are negative. Her hemoglobin level remains stable, and she does not develop signs of bleeding. She is started on a proton pump inhibitor for stress ulcer prophylaxis and is empirically given intravenous N-acetylcysteine until the cause of acute liver failure can be determined.

CAUSES OF ACUTE LIVER FAILURE

2. Which of the following can cause acute liver failure?

• Acetaminophen overdose
• Viral hepatitis
• Autoimmune hepatitis
• Wilson disease
• Alcoholic hepatitis

Drug-induced liver injury is the most common cause of acute liver failure in the United States,^2,3 and of all drugs, acetaminophen overdose is the number-one cause. In acetaminophen-induced liver injury, serum aminotransferase levels are usually elevated to more than 1,000 U/L, while serum bilirubin remains normal in the early stages. Antimicrobial agents, antiepileptic drugs, and herbal supplements have also been implicated in acute liver failure. Our patient has denied taking herbal supplements or medications, including over-the-counter ones.

Acute viral hepatitis can explain the patient’s condition. It is a common cause of acute liver failure in the United States.² Hepatitis A and E are more common in developing countries. Other viruses such as cytomegalovirus, Epstein-Barr virus, herpes simplex virus type 1 and 2, and varicella zoster virus can also cause acute liver failure. Serum aminotransferase levels may exceed 1,000 U/L in patients with viral hepatitis.

A young woman presents with acute liver failure: What is the cause? Is her sister at risk?

Autoimmune hepatitis is a rare cause of acute liver failure, but it should be considered in the differential diagnosis, particularly in middle-aged women with autoimmune disorders such as hypothyroidism. Autoimmune hepatitis can cause marked elevation in aminotransferase levels (> 1,000 U/L).

Wilson disease is an autosomal-recessive disease in which there is excessive accumulation of copper in the liver and other organs because of an inherited defect in the biliary excretion of copper. Wilson disease can cause acute liver failure and should be excluded in any patient, particularly if under age 40 with acute onset of unexplained hepatic, neurologic, or psychiatric disease.

Alcoholic hepatitis usually occurs in patients with a long-standing history of heavy alcohol use. As a result, most patients with alcoholic hepatitis have manifestations of chronic liver disease due to alcohol use. Therefore, by definition, it is not a cause of acute liver failure. Additionally, in patients with alcoholic hepatitis, the aspartate aminotransferase (AST) level is elevated but less than 300 IU/mL, and the ratio of AST to alanine aminotransferase (ALT) is usually more than 2.

CASE CONTINUES: FURTHER TESTING

The results of our patient’s serologic tests are shown in Table 2. Other test results:

• Autoimmune markers including antinuclear antibodies, antimitochondrial antibodies, antismooth muscle antibodies, and liver and kidney microsomal antibodies are negative; her immunoglobulin G (IgG) level is normal
• Serum ceruloplasmin 25 mg/dL (normal 21–45)
• Free serum copper 120 µg/dL (normal 8–12)
• Abdominal ultrasonography is unremarkable, with normal liver parenchyma and no intrahepatic or extrahepatic biliary dilatation
• Doppler ultrasonography of the liver shows patent blood vessels.

3. Based on the new data, which of the following statements is correct?

• Hepatitis B is the cause of acute liver failure in this patient
• Herpetic hepatitis cannot be excluded on the basis of the available data
• Wilson disease is most likely the diagnosis, given her elevated free serum copper
• A normal serum ceruloplasmin level is not sufficient to rule out acute liver failure secondary to Wilson disease

Hepatitis B surface antigen and hepatitis B core antibodies were negative in our patient, excluding hepatitis B virus infection. The positive hepatitis B surface antibody indicates prior immunization.

Herpetic hepatitis is an uncommon but important cause of acute liver failure because the mortality rate is high if the patient is not treated early with acyclovir. Fever, elevated aminotransferases, and leukopenia are common with herpetic hepatitis. Fewer than 50% of patients with herpetic hepatitis have vesicular rash.^4,5 The value of antibody serologic testing is limited due to high rates of false-positive and false-negative results. The gold standard diagnostic tests are viral load (detection of viral RNA by polymerase chain reaction), viral staining on liver biopsy, or both. In our patient, herpes simplex virus polymerase chain reaction testing was negative, which makes herpetic hepatitis unlikely.

Wilson disease is a genetic condition in which the ability to excrete copper in the bile is impaired, resulting in accumulation of copper in the hepatocytes. Subsequently, copper is released into the bloodstream and eventually into the urine.

However, copper excretion into the bile is impaired in patients with acute liver failure regardless of the etiology. Therefore, elevated free serum copper and 24-hour urine copper levels are not specific for the diagnosis of acute liver failure secondary to Wilson disease. Moreover, Kayser-Fleischer rings, which represent copper deposition in the limbus of the cornea, may not be apparent in the early stages of Wilson disease.

Wilson disease involves accumulation of copper in the liver and other organs as the result of a genetic defect

Since it is challenging to diagnose Wilson disease in the context of acute liver failure, Korman et al⁶ compared patients with acute liver failure secondary to Wilson disease with patients with acute liver failure secondary to other conditions. They found that alkaline phosphatase levels are frequently decreased in patients with acute liver failure secondary to Wilson disease,⁶ and that a ratio of alkaline phosphatase to total bilirubin of less than 4 is 94% sensitive and 96% specific for the diagnosis.⁶

Hemolysis is common in acute liver failure due to Wilson disease. This leads to disproportionate elevation of AST compared with ALT, since AST is present in red blood cells. Consequently, the ratio of AST to ALT is usually greater than 2.2, which provides a sensitivity of 94% and a specificity of 86% for the diagnosis.⁶ These two ratios together provide 100% sensitivity and 100% specificity for the diagnosis of Wilson disease in the context of acute liver failure.⁶

Ceruloplasmin. Patients with Wilson disease typically have a low ceruloplasmin level. However, because it is an acute-phase reaction protein, ceruloplasmin can be normal or elevated in patients with acute liver failure from Wilson disease.⁶ Therefore, a normal ceruloplasmin level is not sufficient to rule out acute liver failure secondary to Wilson disease.

CASE CONTINUES: A DEFINITIVE DIAGNOSIS

Our patient undergoes further testing, which reveals the following:

• Her 24-hour urinary excretion of copper is 150 µg (reference value < 30)
• Slit-lamp examination is normal and shows no evidence of Kayser-Fleischer rings
• Her ratio of alkaline phosphatase to total bilirubin is 0.77 based on her initial laboratory results (Table 1)
• Her AST-ALT ratio is 3.4.

The diagnosis in our patient is acute liver failure secondary to Wilson disease.

4. What is the most appropriate next step?

• Liver biopsy
• d-penicillamine by mouth
• Trientine by mouth
• Liver transplant
• Plasmapheresis

Liver biopsy. Accumulation of copper in the liver parenchyma in patients with Wilson disease is sporadic. Therefore, qualitative copper staining on liver biopsy can be falsely negative. Quantitative copper measurement in liver tissue is the gold standard for the diagnosis of Wilson disease. However, the test is time-consuming and is not rapidly available in the context of acute liver failure.

Chelating agents such as d-pencillamine and trientine are used to treat the chronic manifestations of Wilson disease but are not useful for acute liver failure secondary to Wilson disease.

Acute liver failure secondary to Wilson disease is life-threatening, and liver transplant is considered the only definitive life-saving therapy.

Therapeutic plasmapheresis has been reported to be a successful adjunctive therapy to bridge patients with acute liver failure secondary to Wilson disease to transplant.⁷ However, liver transplant is still the only definitive treatment.

CASE CONTINUES: THE PATIENT’S SISTER SEEKS CARE

The patient undergoes liver transplantation, with no perioperative or postoperative complications.

The patient’s 18-year-old sister is now seeking medical attention in the outpatient clinic, concerned that she may have Wilson disease. She is otherwise healthy and denies any symptoms or complaints.

5. What is the next step for the patient’s sister?

• Reassurance
• Prophylaxis with trientine
• Check liver enzyme levels, serum ceruloplasmin level, and urine copper, and order a slit-lamp examination
• Genetic testing

Wilson disease can be asymptomatic in its early stages and may be diagnosed incidentally during routine blood tests that reveal abnormal liver enzyme levels. All patients with a confirmed family history of Wilson disease should be screened even if they are asymptomatic. The diagnosis of Wilson disease should be established in first-degree relatives before specific treatment for the relatives is prescribed.

Based on information in Roberts EA, Schilsky ML; American Association for Study of Liver Diseases (AASLD). Diagnosis and treatment of Wilson disease: an update. Hepatology 2008; 7:2089–2111.

The first step in screening a first-degree relative for Wilson disease is to check liver enzyme levels (specifically aminotransferases, alkaline phosphatase, and bilirubin), serum ceruloplasmin level, and 24-hour urine copper, and order an ophthalmologic slit-lamp examination. If any of these tests is abnormal, liver biopsy should be performed for histopathologic evaluation and quantitative copper measurement. Kayser-Fleischer  rings are seen in only 50% of patients with Wilson disease and hepatic involvement, but they are pathognomic. Guidelines⁸ for screening first-degree relatives of Wilson disease patients are shown in Figure 1.

Genetic analysis. ATP7B, the Wilson disease gene, is located on chromosome 13. At least 300 mutations of the gene have been described,² and the most common mutation is present in only 15% to 30% of the Wilson disease population.^8–10 Routine molecular testing of the ATP7B

CASE CONTINUES: WORKUP OF THE PATIENT’S SISTER

The patient’s sister has no symptoms and her physical examination is normal. Slit-lamp examination reveals no evidence of Kayser-Fleischer rings. Her laboratory values, including complete blood counts, complete metabolic panel, and INR, are within normal ranges. Other test results, however, are abnormal:

• Free serum copper level 27 µg/dL (normal 8–12)
• Serum ceruloplasmin 9.0 mg/dL (normal 20–50)
• 24-hour urinary copper excretion 135 µg (normal < 30).

She undergoes liver biopsy for quantitative copper measurement, and the result is very high at 1,118 µg/g dry weight (reference range 10–35). The diagnosis of Wilson disease is established.

TREATING CHRONIC WILSON DISEASE

6. Which of the following is not an appropriate next step for the patient’s sister?

• Tetrathiomolybdate
• d-penicillamine
• Trientine
• Zinc salts
• Prednisone

The goal of medical treatment of chronic Wilson disease is to improve symptoms and prevent progression of the disease.

Chelating agents and zinc salts are the most commonly used medicines in the management of Wilson disease. Chelating agents remove copper from tissue, whereas zinc blocks the intestinal absorption of copper and stimulates the synthesis of endogenous chelators such as metallothioneins. Tetrathiomolybdate is an alternative agent developed to interfere with the distribution of excess body copper to susceptible target sites by reducing free serum copper (Table 3). There are no data to support the use of prednisone in the treatment of Wilson disease.

During treatment with chelating agents, 24-hour urinary excretion of copper is routinely monitored to determine the efficacy of therapy and adherence to treatment. Once de-coppering is achieved, as evidenced by a normalization of 24-hour urine copper excretion, the chelating agent can be switched to zinc salts to prevent intestinal absorption of copper.

Clinical and biochemical stabilization is achieved typically within 2 to 6 months of the initial treatment with chelating agents.⁸ Organ meats, nuts, shellfish, and chocolate are rich in copper and should be avoided.

The patient’s sister is started on trientine 250 mg orally three times daily on an empty stomach at least 1 hour before meals. Treatment is monitored by following 24-hour urine copper measurement. A 24-hour urine copper measurement at 3 months after starting treatment has increased from 54 at baseline to 350 µg, which indicates that the copper is being removed from tissues. The plan is for early substitution of zinc for long-term maintenance once de-coppering is completed.

KEY POINTS

• Acute liver failure is severe acute liver injury characterized by coagulopathy (INR ≥ 1.5) and encephalopathy in a patient with no preexisting liver disease and with duration of symptoms less than 26 weeks.
• Acute liver failure secondary to Wilson disease is uncommon but should be excluded, particularly in young patients.
• The diagnosis of Wilson disease in the setting of acute liver failure is challenging because the serum ceruloplasmin level may be normal in acute liver failure secondary to Wilson disease, and free serum copper and 24-hour urine copper are usually elevated in all acute liver failure patients regardless of the etiology.
• A ratio of alkaline phosphatase to total bilirubin of less than 4 plus an AST-ALT ratio greater than 2.2 in a patient with acute liver failure should be regarded as Wilson disease until proven otherwise (Figure 2).
• Acute liver failure secondary to Wilson disease is usually fatal, and emergency liver transplant is a life-saving procedure.
• Screening of first-degree relatives of Wilson disease patients should include a history and physical examination, liver enzyme tests, complete blood cell count, serum ceruloplasmin level, serum free copper level, slit-lamp examination of the eyes, and 24-hour urinary copper measurement. Genetic tests are supplementary for screening but are not routinely available.

To the Editor: We previously addressed whether VKORC1 and CYP2C9 pharmacogenomic testing should be considered when prescribing warfarin.¹ Our recommendation, based on available evidence at that time, was that physicians should consider pharmacogenomic testing for any patient who is started on warfarin therapy.

Since the publication of this recommendation, two major trials, COAG (Clarification of Optimal Anticoagulation Through Genetics)² and EU-PACT (European Pharmacogenetics of Anticoagulant Therapy-Warfarin),³ were published along with commentaries debating the clinical utility of warfarin pharmacogenomics.^4–15 Based on these publications, we would like to update our recommendations for pharmacogenomic testing for warfarin therapy.

COAG compared the efficacy of a clinical algorithm or a clinical algorithm plus VKORC1 and CYP2C9 genotyping to guide warfarin dosage. At the end of 4 weeks, the mean percentage of time within the therapeutic international normalized ratio (INR) range was 45.4% for those in the clinical algorithm arm and 45.2% for those in the genotyping arm (95% confidence interval [CI] –3.4 to 3.1, P = .91). For both treatment groups, clinical data that included body surface area, age, target INR, concomitantly prescribed drugs, and smoking status were used to predict warfarin dose, with the genotyping arm including VKORC1 and CYP2C9. Although VKORC1 and CYP2C9 genotyping offered no additional benefit, caution should be used when extrapolating this conclusion to clinical settings in which warfarin therapy is initiated using a standardized starting dose (eg, 5 mg daily) instead of a clinical dosing algorithm.

Of interest, in the COAG trial, among black patients, the mean percentage of time in the therapeutic INR range was significantly less for those in the genotype-guided arm than for those in the clinically guided arm—ie, 35.2% vs 43.5% (95% CI –15.0 to –2.0, P = .01). The percentage of time with therapeutic INR has been identified as a surrogate marker for poor outcomes such as death, stroke, or major hemorrhage, with those with a lower percentage of time in therapeutic INR being at greater risk of an adverse event.¹⁶ Wan et al¹⁷ demonstrated that a 6.9% improvement of time in therapeutic INR decreased the risk of major hemorrhage by one event per 100 patient-years.¹⁷ Therefore, black patients in the COAG genotyping arm may have been at greater risk for an adverse event because of a lower observed percentage of time within the therapeutic INR range.

In the COAG trial, genotyping was done for only one VKORC1 variant and for two CYP2C9 alleles (CYP2C9*2, and CYP2C9*3). Other genetic variants are of clinical importance for warfarin dosing in black patients, and the lack of genotyping for these additional variants may explain why black patients in the genotyping arm performed worse.^5,7,11 In particular, CYP2C9*8 may be an important predictor of warfarin dose in black patients.¹⁸

EU-PACT compared the efficacy of standardized warfarin dosing and that of a clinical algorithm.³ Patients in the standardized dosing arm were prescribed warfarin 10 mg on the first day of treatment (5 mg for those over age 75), and 5 mg on days 2 and 3, with subsequent dosing adjustments based on INR. Patients in the genotyping arm were prescribed warfarin based on an algorithm that incorporated clinical data that included body surface area, age, and concomitantly prescribed drugs, as well as VKORC1 and CYP2C9 genotypes. At the end of 12 weeks, the mean percentage of time in the therapeutic INR range was 60.3% for those in the standardized-dosing arm and 67.4% for those in the genotyping arm (95% CI 3.3 to 10.6, P < .001).² The approximate 7% improvement in percentage of time in the therapeutic INR range may predict a lower risk of hemorrhage for those in the genotyping arm.¹⁷ Although patients in the genotyping arm had a higher percentage of time in the therapeutic INR range, it is unclear whether genotyping alone is superior to standardized dosing because the dosing algorithm used both clinical data and genotype data.

There are substantial differences between the COAG and EU-PACT trials, including dosing schemes, racial diversity, and trial length, and these differences could have contributed to the conflicting results. Based on these two trials, a possible conclusion is that genotype-guided warfarin dosing may be superior to standardized dosing, but may be no better than utilizing a clinical algorithm in white patients. For black patients, additional studies are needed to determine which genetic variants are of importance for guiding warfarin dosing.

We would like to update the recommendations we made in our previously published article,¹ to state that genotyping for CYP2C9 and VKORC1 may be of clinical utility in white patients depending on whether standardized dosing or a clinical algorithm is used to initiate warfarin therapy. Routine genotyping in black patients is not recommended until further studies clarify which genetic variants are of importance for guiding warfarin dosing.

The ongoing Genetics Informatics Trial of Warfarin to Prevent Venous Thrombosis may bring much needed clarity to the clinical utility of warfarin pharmacogenomics. We hope to publish a more detailed update of our 2013 article after completion of that trial.

To the Editor: We previously addressed whether VKORC1 and CYP2C9 pharmacogenomic testing should be considered when prescribing warfarin.¹ Our recommendation, based on available evidence at that time, was that physicians should consider pharmacogenomic testing for any patient who is started on warfarin therapy.

Since the publication of this recommendation, two major trials, COAG (Clarification of Optimal Anticoagulation Through Genetics)² and EU-PACT (European Pharmacogenetics of Anticoagulant Therapy-Warfarin),³ were published along with commentaries debating the clinical utility of warfarin pharmacogenomics.^4–15 Based on these publications, we would like to update our recommendations for pharmacogenomic testing for warfarin therapy.

COAG compared the efficacy of a clinical algorithm or a clinical algorithm plus VKORC1 and CYP2C9 genotyping to guide warfarin dosage. At the end of 4 weeks, the mean percentage of time within the therapeutic international normalized ratio (INR) range was 45.4% for those in the clinical algorithm arm and 45.2% for those in the genotyping arm (95% confidence interval [CI] –3.4 to 3.1, P = .91). For both treatment groups, clinical data that included body surface area, age, target INR, concomitantly prescribed drugs, and smoking status were used to predict warfarin dose, with the genotyping arm including VKORC1 and CYP2C9. Although VKORC1 and CYP2C9 genotyping offered no additional benefit, caution should be used when extrapolating this conclusion to clinical settings in which warfarin therapy is initiated using a standardized starting dose (eg, 5 mg daily) instead of a clinical dosing algorithm.

Of interest, in the COAG trial, among black patients, the mean percentage of time in the therapeutic INR range was significantly less for those in the genotype-guided arm than for those in the clinically guided arm—ie, 35.2% vs 43.5% (95% CI –15.0 to –2.0, P = .01). The percentage of time with therapeutic INR has been identified as a surrogate marker for poor outcomes such as death, stroke, or major hemorrhage, with those with a lower percentage of time in therapeutic INR being at greater risk of an adverse event.¹⁶ Wan et al¹⁷ demonstrated that a 6.9% improvement of time in therapeutic INR decreased the risk of major hemorrhage by one event per 100 patient-years.¹⁷ Therefore, black patients in the COAG genotyping arm may have been at greater risk for an adverse event because of a lower observed percentage of time within the therapeutic INR range.

In the COAG trial, genotyping was done for only one VKORC1 variant and for two CYP2C9 alleles (CYP2C9*2, and CYP2C9*3). Other genetic variants are of clinical importance for warfarin dosing in black patients, and the lack of genotyping for these additional variants may explain why black patients in the genotyping arm performed worse.^5,7,11 In particular, CYP2C9*8 may be an important predictor of warfarin dose in black patients.¹⁸

EU-PACT compared the efficacy of standardized warfarin dosing and that of a clinical algorithm.³ Patients in the standardized dosing arm were prescribed warfarin 10 mg on the first day of treatment (5 mg for those over age 75), and 5 mg on days 2 and 3, with subsequent dosing adjustments based on INR. Patients in the genotyping arm were prescribed warfarin based on an algorithm that incorporated clinical data that included body surface area, age, and concomitantly prescribed drugs, as well as VKORC1 and CYP2C9 genotypes. At the end of 12 weeks, the mean percentage of time in the therapeutic INR range was 60.3% for those in the standardized-dosing arm and 67.4% for those in the genotyping arm (95% CI 3.3 to 10.6, P < .001).² The approximate 7% improvement in percentage of time in the therapeutic INR range may predict a lower risk of hemorrhage for those in the genotyping arm.¹⁷ Although patients in the genotyping arm had a higher percentage of time in the therapeutic INR range, it is unclear whether genotyping alone is superior to standardized dosing because the dosing algorithm used both clinical data and genotype data.

There are substantial differences between the COAG and EU-PACT trials, including dosing schemes, racial diversity, and trial length, and these differences could have contributed to the conflicting results. Based on these two trials, a possible conclusion is that genotype-guided warfarin dosing may be superior to standardized dosing, but may be no better than utilizing a clinical algorithm in white patients. For black patients, additional studies are needed to determine which genetic variants are of importance for guiding warfarin dosing.

We would like to update the recommendations we made in our previously published article,¹ to state that genotyping for CYP2C9 and VKORC1 may be of clinical utility in white patients depending on whether standardized dosing or a clinical algorithm is used to initiate warfarin therapy. Routine genotyping in black patients is not recommended until further studies clarify which genetic variants are of importance for guiding warfarin dosing.

The ongoing Genetics Informatics Trial of Warfarin to Prevent Venous Thrombosis may bring much needed clarity to the clinical utility of warfarin pharmacogenomics. We hope to publish a more detailed update of our 2013 article after completion of that trial.

To the Editor: We previously addressed whether VKORC1 and CYP2C9 pharmacogenomic testing should be considered when prescribing warfarin.¹ Our recommendation, based on available evidence at that time, was that physicians should consider pharmacogenomic testing for any patient who is started on warfarin therapy.

Since the publication of this recommendation, two major trials, COAG (Clarification of Optimal Anticoagulation Through Genetics)² and EU-PACT (European Pharmacogenetics of Anticoagulant Therapy-Warfarin),³ were published along with commentaries debating the clinical utility of warfarin pharmacogenomics.^4–15 Based on these publications, we would like to update our recommendations for pharmacogenomic testing for warfarin therapy.

COAG compared the efficacy of a clinical algorithm or a clinical algorithm plus VKORC1 and CYP2C9 genotyping to guide warfarin dosage. At the end of 4 weeks, the mean percentage of time within the therapeutic international normalized ratio (INR) range was 45.4% for those in the clinical algorithm arm and 45.2% for those in the genotyping arm (95% confidence interval [CI] –3.4 to 3.1, P = .91). For both treatment groups, clinical data that included body surface area, age, target INR, concomitantly prescribed drugs, and smoking status were used to predict warfarin dose, with the genotyping arm including VKORC1 and CYP2C9. Although VKORC1 and CYP2C9 genotyping offered no additional benefit, caution should be used when extrapolating this conclusion to clinical settings in which warfarin therapy is initiated using a standardized starting dose (eg, 5 mg daily) instead of a clinical dosing algorithm.

Of interest, in the COAG trial, among black patients, the mean percentage of time in the therapeutic INR range was significantly less for those in the genotype-guided arm than for those in the clinically guided arm—ie, 35.2% vs 43.5% (95% CI –15.0 to –2.0, P = .01). The percentage of time with therapeutic INR has been identified as a surrogate marker for poor outcomes such as death, stroke, or major hemorrhage, with those with a lower percentage of time in therapeutic INR being at greater risk of an adverse event.¹⁶ Wan et al¹⁷ demonstrated that a 6.9% improvement of time in therapeutic INR decreased the risk of major hemorrhage by one event per 100 patient-years.¹⁷ Therefore, black patients in the COAG genotyping arm may have been at greater risk for an adverse event because of a lower observed percentage of time within the therapeutic INR range.

In the COAG trial, genotyping was done for only one VKORC1 variant and for two CYP2C9 alleles (CYP2C9*2, and CYP2C9*3). Other genetic variants are of clinical importance for warfarin dosing in black patients, and the lack of genotyping for these additional variants may explain why black patients in the genotyping arm performed worse.^5,7,11 In particular, CYP2C9*8 may be an important predictor of warfarin dose in black patients.¹⁸

EU-PACT compared the efficacy of standardized warfarin dosing and that of a clinical algorithm.³ Patients in the standardized dosing arm were prescribed warfarin 10 mg on the first day of treatment (5 mg for those over age 75), and 5 mg on days 2 and 3, with subsequent dosing adjustments based on INR. Patients in the genotyping arm were prescribed warfarin based on an algorithm that incorporated clinical data that included body surface area, age, and concomitantly prescribed drugs, as well as VKORC1 and CYP2C9 genotypes. At the end of 12 weeks, the mean percentage of time in the therapeutic INR range was 60.3% for those in the standardized-dosing arm and 67.4% for those in the genotyping arm (95% CI 3.3 to 10.6, P < .001).² The approximate 7% improvement in percentage of time in the therapeutic INR range may predict a lower risk of hemorrhage for those in the genotyping arm.¹⁷ Although patients in the genotyping arm had a higher percentage of time in the therapeutic INR range, it is unclear whether genotyping alone is superior to standardized dosing because the dosing algorithm used both clinical data and genotype data.

There are substantial differences between the COAG and EU-PACT trials, including dosing schemes, racial diversity, and trial length, and these differences could have contributed to the conflicting results. Based on these two trials, a possible conclusion is that genotype-guided warfarin dosing may be superior to standardized dosing, but may be no better than utilizing a clinical algorithm in white patients. For black patients, additional studies are needed to determine which genetic variants are of importance for guiding warfarin dosing.

We would like to update the recommendations we made in our previously published article,¹ to state that genotyping for CYP2C9 and VKORC1 may be of clinical utility in white patients depending on whether standardized dosing or a clinical algorithm is used to initiate warfarin therapy. Routine genotyping in black patients is not recommended until further studies clarify which genetic variants are of importance for guiding warfarin dosing.

The ongoing Genetics Informatics Trial of Warfarin to Prevent Venous Thrombosis may bring much needed clarity to the clinical utility of warfarin pharmacogenomics. We hope to publish a more detailed update of our 2013 article after completion of that trial.

“Change is the only constant.”

—Heraclitus (c 535–475 bce)

With the cost of health care rising and money to pay for it shrinking, there has never been a greater need to reduce waste.

See related article

Ineffective treatments and adverse drug effects account for much preventable morbidity and expense. New treatments, touted as more potent, are often introduced as replacements for traditional ones that are still effective in many patients, adding to costs and the potential for harm. For the pharmaceutical industry, the search for new “blockbuster” drugs seems to have hit a wall, at least in cardiovascular medicine.¹ Advances often come at the cost of adverse effects, such as bleeding with triple antiplatelet therapy and diabetes with potent statin drugs.

The path to maximizing benefit and reducing harm now appears to lie in stratifying populations and appreciating patient individuality in response to treatment. For many decades we have known that patients vary widely in their response to drugs, owing to personal factors such as body surface area, age, environment, and genetics. And indeed, we treat our patients as individuals, for example by tailoring aminoglycoside dose to weight and renal function.

However, clinical trials typically give us an idea of the benefits only to the average patient. While subgroup analyses identify groups that may benefit more or less from treatment, the additional information they provide is not easily integrated into the clinical model of prescribing, in which one size fits all.

THE PROMISE OF PHARMACOGENETICS

The emerging field of pharmacogenetics promises to give clinicians the tools to make informed treatment decisions based on predictive genetic testing. This genetic testing aims to match treatment to an individual’s genetic profile. This often involves analyzing single-nucleotide polymorphisms in genes for enzymes that metabolize drugs, such as the cytochrome P450 enzymes, to predict efficacy or an adverse event with treatment.

Pharmacogenetics is playing an increasing role in clinical trials, particularly in the early stages of drug development, by helping to reduce the number of patients needed, prove efficacy, and identify subgroups in which alternative treatment can be targeted. At another level, a molecular understanding of disease is leading to truly targeted treatments based on genomics.

Over recent years, genetic testing has been increasingly used in clinical practice, thanks to a convergence of factors such as rapid, low-cost tests, a growing evidence base, and emerging interest among doctors and payers.

An advantage to using genetic testing as opposed to other types of laboratory testing, such as measuring the concentration of the drug in the blood during treatment, is that genetic tests can predict the response to treatment before the treatment is started. Moreover, with therapeutic drug monitoring after treatment has begun, there are sometimes no detectable measures of toxicity. For example, both carbamazepine and the antiviral drug abacavir can—fortunately only rarely—cause Stevens-Johnson syndrome. But before genetic markers were discovered, there was no method of estimating this risk apart from taking a family history.^2,3 Considering the numbers of people involved, it was not feasible until recently to suggest genetic screening for patients starting on these drugs. However, the cost of genotyping and gene sequencing has been falling at a rate inversely faster than Moore’s law (an approximate annual doubling in computer power), and population genomics is becoming a reality.⁴

The US Food and Drug Administration (FDA) recognizes the current and future value of pharmacogenetics in drug safety and development. A number of approved pharmacogenetic biomarkers are listed on the FDA website (Table 1). Black box warnings have been mandated for a number of drugs on the basis of observational evidence.

The FDA also promotes rapid approval for novel drugs with pharmacogenetic “companion diagnostics.” A recent example of this was the approval of ivacaftor for cystic fibrosis patients who have the G551D mutation.⁵ Here, a molecular understanding of the condition led to the development of a targeted treatment. Although the cost of developing this drug was high, the path is now paved for similar advances. Oncologists are familiar with these advances with the emergence of new molecularly targeted treatments, eg, BRAF inhibitors in metastatic melanoma, imatinib in chronic myeloid leukemia, and gefitinib in non-small-cell lung cancer.

PHARMACOGENETICS IN CARDIOVASCULAR MEDICINE

Cardiovascular medicine also stands to benefit from rapid advances in pharmacogenetics.

While no treatment has been developed that targets the molecular basis of cardiovascular disease, a number of genomic biomarkers have emerged that identify patients at risk of adverse reactions or treatment failure. These include genetic tests to predict the maintenance dose and risk of bleeding with warfarin,⁶ the likelihood of myopathy and myositis with simvastatin,⁷ and the risk of recurrent thrombotic events with clopidogrel.^8–10

Using pharmacogenetics in prescribing warfarin and its alternatives

The pharmacogenetics of warfarin has been extensively researched, but genotyping before prescribing this drug is not yet widely done.

In 2007, the FDA updated the labeling of warfarin to include information about the influence of two genes, VKORC1 and CYP2C9, on a patient’s response to this drug. In 2010 this was updated to add that testing for these genes could be used to predict the maintenance dose of the drug. Difficulties with algorithms used to integrate this into clinical practice have hindered adoption of this testing.

With the advent of new anticoagulants such as dabigatran, rivaroxaban, and apixaban, many have expected warfarin and its pharmacogenetics to become obsolete. However, the new agents cost considerably more. Further, they may not offer a very great advantage over warfarin: in the Randomized Evaluation of Long-term Anticoagulant Therapy (RE-LY) trial, the absolute risk reduction in intracranial hemorrhage with dabigatran vs warfarin was small.¹¹ Therefore, dabigatran is probably not cost-effective in populations at low risk of bleeding.¹² A cost-effectiveness analysis comparing warfarin with dabigatran in patients with uncomplicated atrial fibrillation has suggested that dabigatran is, however, cost-effective in patients at moderate risk.¹²

In the RE-LY trial, the international normalized ratios (INRs) of the patients in the warfarin group were in the therapeutic range only 64% of the time. The advantages of dabigatran over warfarin become less pronounced as warfarin control is tightened.¹³ Of note, pharmacogenetics and home monitoring of the INR have both been shown to lead to tighter control of the INR, with greater time within the therapeutic range.^14,15

Moreover, genetic testing can help us reduce the number of bleeding events in patients taking warfarin.¹⁶ Patients who carry the CYP2C9*2 or CYP2C9*3 polymorphism metabolize S-warfarin slower and therefore have a threefold higher risk of hemorrhage after starting warfarin.¹⁷ We could speculate that patients carrying these variants may be better served by the newer anticoagulants, though this has not been tested in any clinical trial.

It is also worth appreciating that the conditions requiring anticoagulation, such as atrial fibrillation, also have a strong genetic basis. Variants in chromosomes 4q25, 1q21, and 16q22 have all been associated with atrial fibrillation.¹⁸ The risk of atrial fibrillation is five to six times higher in carriers of multiple variants within all of these loci.¹⁹ Genetic variants at 4q25 have been associated with the response to specific antiarrhythmic drug treatments,²⁰ response to pulmonary vein isolation, ^21,22 and direct-current cardioversion.²³ One can imagine a future in which patients with palpitations, carrying multiple gene risk variants, will choose prolonged monitoring at home to confirm a diagnosis. They would then be provided with a personalized best management strategy, using their personal preferences, clinical data, and genetic profile to make a treatment decision.

Using pharmacogenetics in prescribing clopidogrel and its alternatives

The pharmacogenetics of clopidogrel is of particular interest, as it has the potential of establishing a rational basis for using newer antiplatelet drugs such as ticagrelor and prasugrel, which are considerably more expensive than generic clopidogrel.

Most of the people who do not respond to clopidogrel carry the common cytochrome P450 2C19 variants CYP2C19*2 or CYP2C19*3.⁹ These variants are present in particularly high frequency in Asians and African Americans, who often do not feature in large randomized trials.

Newer antiplatelet agents have failed to demonstrate consistent superiority to clopidogrel without a tradeoff of more bleeding. However, in the Trial to Assess Improvement in Therapeutic Outcomes by Optimizing Platelet Inhibition With Prasugrel-Thrombolysis in Myocardial Infarction 39 (TRITON TIMI-38),^24,25 patients with the *2 variant receiving prasugrel had lower cardiovascular event rates than *2 carriers receiving clopidogrel.

Similarly, the patients who benefit the most from ticagrelor are carriers of the 2C19 nonresponder variants. In a large study, clopidogrel responders who did not carry either 2C19 nonresponder genetic variants or ABCB1 variants had cardiovascular outcomes similar to those of patients receiving ticagrelor.²⁶

Clinicians have been cautious in prescribing potent antiplatelet agents to all patients because of the risk of bleeding. One could assume that by reserving newer agents for clopidogrel nonresponders, the bleeding risk could be minimized and overall benefit could be preserved with this strategy.

Cost may also be contained. The cost-effectiveness of such an approach with prasugrel has been tested with computer modeling and appears favorable.²⁷ On the other hand, a similar yet limited analysis did not find genotype-driven use of ticagrelor to be cost-effective.²⁸ This was mostly due to fewer deaths in patients receiving ticagrelor. However, the cost estimate for genotype-guided therapy was overestimated, as heterozygotes in the model were treated with ticagrelor instead of a high dose of clopidogrel.

It now appears that heterozygotes, ie, patients with one copy of the nonresponder variant, can achieve similar platelet inhibition with clopidogrel 225 mg daily as noncarriers on 75 mg daily.²⁹ Since genotype-guided antiplatelet therapy has not been tested in a randomized outcomes trial, this tailored strategy has not been widely accepted.

THE FUTURE

The barriers to adoption of pharmacogenetics are considerable. Clinicians need to be educated about it, reimbursement needs to be worked out, and the pharmaceutical industry needs to get behind it. Nevertheless, the future of pharmacogenetics is extremely promising.

Research networks are forming to support the use of pharmacogenetics in clinical practice. The Pharmgkb (www.pharmgkb.org) database serves as a hub for educating clinicians and researchers as well as curating data for reference. Vanderbilt University is piloting the BioVu project, in which DNA and genotype data on patients are being stored and matched to the electronic clinical records.³⁰ These projects not only provide clinically useful information on the current state of the art of pharmacogenetics, they also aid in disseminating new information about genotype-phenotype relationships.

Analytical software that uses “natural language processing” is being applied to clinician-generated notes to derive new observations and associations between genetic variants and clinical phenotypes. Integrating this information in real-time decision-support modules in the electronic health record provides a feedback loop for a rapid assimilation of new knowledge. Similar innovative decision-support modules are being established by Cleveland Clinic’s Center for Personalized Healthcare.³¹

The rise of ‘omic’ sciences

Pharmacogenetics and pharmacogenomics are part of a larger set of “omic” sciences. The suffix “-omics” implies a larger, more holistic view and is being applied to a number of fields—for example, the study of proteins (proteomics) and the study of metabolites (metabolomics). Profiling proteins and metabolites delivers a deluge of information on a patient that can be clustered, using pattern-recognition software, into population subgroups. Patterns of multiple proteins or metabolites are extracted from this spectral data to identify disease or response to treatment (pharmacometabolomics).

Metabolomics has been shown to predict the response to statins,³² diagnose myocardial infarction,³³ and reclassify cardiovascular risk status.³⁴ In addition, whereas traditional laboratory chemistry is reductionist, using single biomarkers for single-disease diagnosis, omic technologies hold the potential to reveal information on a number of possible health or disease states. The identification of “healthy” profiles using these technologies can potentially provide positive feedback to patients undertaking lifestyle changes and treatment.

The instrument costs for proteomic and metabolomic profiling are relatively high. However, the ongoing running costs are minimal, estimated at as low as less than $13 per test, as there are no expensive reagents.³³ High-volume testing therefore becomes very cost-effective.

Although omic science appears futuristic, proteomics and metabolomics are already used in many clinical laboratories to rapidly identify bacteria. These methods have already revolutionized the way laboratories identify microbes, since they are automated, reduce workload, and give very fast results.

The cost of genetic testing is falling

Critics of pharmacogenetics claim that the predictive value of genetic testing is poor, that evidence is lacking, and that the cost is too high. In all new technologies, the first iteration is coarse, but performance improves with use. The first major barrier is adoption. Projects like BioVu are establishing the infrastructure for a feedback loop to iteratively improve upon the status quo and provide the evidence base clinicians demand.

The cost of genetic testing is falling rapidly, with whole-genome sequencing and annotation now costing less than $5,000. The cost of a pharmacogenetic test can be as low as $100 using low-cost nanotechnology, and the test needs to be performed only once in a patient’s lifetime.²⁷

As other related molecular technologies such as proteomics and metabolomics become available and are integrated with genomics, the predictive ability of this science will improve.

AWAY FROM ONE-SIZE-FITS-ALL MEDICINE

Over the last decade there has been a trend away from “one size fits all” to customized “markets of one” in everything from consumer products to education to medicine. Mass customizing, also known as personalization, has been embraced by the internet community as a means to increase efficiency and reduce cost. This occurs by eliminating waste in redundant work or production of ineffective products.

Personalization on the Internet has been enabled through the use of informatics, mathematics, and supercomputing. The same tools that have personalized the delivery of consumer products are also being applied to the field of pharmacogenetics. Applied in an evidence-based fashion, these new technologies should profoundly improve patient care now and in the future.

“Change is the only constant.”

—Heraclitus (c 535–475 bce)

With the cost of health care rising and money to pay for it shrinking, there has never been a greater need to reduce waste.

See related article

Ineffective treatments and adverse drug effects account for much preventable morbidity and expense. New treatments, touted as more potent, are often introduced as replacements for traditional ones that are still effective in many patients, adding to costs and the potential for harm. For the pharmaceutical industry, the search for new “blockbuster” drugs seems to have hit a wall, at least in cardiovascular medicine.¹ Advances often come at the cost of adverse effects, such as bleeding with triple antiplatelet therapy and diabetes with potent statin drugs.

The path to maximizing benefit and reducing harm now appears to lie in stratifying populations and appreciating patient individuality in response to treatment. For many decades we have known that patients vary widely in their response to drugs, owing to personal factors such as body surface area, age, environment, and genetics. And indeed, we treat our patients as individuals, for example by tailoring aminoglycoside dose to weight and renal function.

However, clinical trials typically give us an idea of the benefits only to the average patient. While subgroup analyses identify groups that may benefit more or less from treatment, the additional information they provide is not easily integrated into the clinical model of prescribing, in which one size fits all.

THE PROMISE OF PHARMACOGENETICS

The emerging field of pharmacogenetics promises to give clinicians the tools to make informed treatment decisions based on predictive genetic testing. This genetic testing aims to match treatment to an individual’s genetic profile. This often involves analyzing single-nucleotide polymorphisms in genes for enzymes that metabolize drugs, such as the cytochrome P450 enzymes, to predict efficacy or an adverse event with treatment.

Pharmacogenetics is playing an increasing role in clinical trials, particularly in the early stages of drug development, by helping to reduce the number of patients needed, prove efficacy, and identify subgroups in which alternative treatment can be targeted. At another level, a molecular understanding of disease is leading to truly targeted treatments based on genomics.

Over recent years, genetic testing has been increasingly used in clinical practice, thanks to a convergence of factors such as rapid, low-cost tests, a growing evidence base, and emerging interest among doctors and payers.

An advantage to using genetic testing as opposed to other types of laboratory testing, such as measuring the concentration of the drug in the blood during treatment, is that genetic tests can predict the response to treatment before the treatment is started. Moreover, with therapeutic drug monitoring after treatment has begun, there are sometimes no detectable measures of toxicity. For example, both carbamazepine and the antiviral drug abacavir can—fortunately only rarely—cause Stevens-Johnson syndrome. But before genetic markers were discovered, there was no method of estimating this risk apart from taking a family history.^2,3 Considering the numbers of people involved, it was not feasible until recently to suggest genetic screening for patients starting on these drugs. However, the cost of genotyping and gene sequencing has been falling at a rate inversely faster than Moore’s law (an approximate annual doubling in computer power), and population genomics is becoming a reality.⁴

The US Food and Drug Administration (FDA) recognizes the current and future value of pharmacogenetics in drug safety and development. A number of approved pharmacogenetic biomarkers are listed on the FDA website (Table 1). Black box warnings have been mandated for a number of drugs on the basis of observational evidence.

The FDA also promotes rapid approval for novel drugs with pharmacogenetic “companion diagnostics.” A recent example of this was the approval of ivacaftor for cystic fibrosis patients who have the G551D mutation.⁵ Here, a molecular understanding of the condition led to the development of a targeted treatment. Although the cost of developing this drug was high, the path is now paved for similar advances. Oncologists are familiar with these advances with the emergence of new molecularly targeted treatments, eg, BRAF inhibitors in metastatic melanoma, imatinib in chronic myeloid leukemia, and gefitinib in non-small-cell lung cancer.

PHARMACOGENETICS IN CARDIOVASCULAR MEDICINE

Cardiovascular medicine also stands to benefit from rapid advances in pharmacogenetics.

While no treatment has been developed that targets the molecular basis of cardiovascular disease, a number of genomic biomarkers have emerged that identify patients at risk of adverse reactions or treatment failure. These include genetic tests to predict the maintenance dose and risk of bleeding with warfarin,⁶ the likelihood of myopathy and myositis with simvastatin,⁷ and the risk of recurrent thrombotic events with clopidogrel.^8–10

Using pharmacogenetics in prescribing warfarin and its alternatives

The pharmacogenetics of warfarin has been extensively researched, but genotyping before prescribing this drug is not yet widely done.

In 2007, the FDA updated the labeling of warfarin to include information about the influence of two genes, VKORC1 and CYP2C9, on a patient’s response to this drug. In 2010 this was updated to add that testing for these genes could be used to predict the maintenance dose of the drug. Difficulties with algorithms used to integrate this into clinical practice have hindered adoption of this testing.

With the advent of new anticoagulants such as dabigatran, rivaroxaban, and apixaban, many have expected warfarin and its pharmacogenetics to become obsolete. However, the new agents cost considerably more. Further, they may not offer a very great advantage over warfarin: in the Randomized Evaluation of Long-term Anticoagulant Therapy (RE-LY) trial, the absolute risk reduction in intracranial hemorrhage with dabigatran vs warfarin was small.¹¹ Therefore, dabigatran is probably not cost-effective in populations at low risk of bleeding.¹² A cost-effectiveness analysis comparing warfarin with dabigatran in patients with uncomplicated atrial fibrillation has suggested that dabigatran is, however, cost-effective in patients at moderate risk.¹²

In the RE-LY trial, the international normalized ratios (INRs) of the patients in the warfarin group were in the therapeutic range only 64% of the time. The advantages of dabigatran over warfarin become less pronounced as warfarin control is tightened.¹³ Of note, pharmacogenetics and home monitoring of the INR have both been shown to lead to tighter control of the INR, with greater time within the therapeutic range.^14,15

Moreover, genetic testing can help us reduce the number of bleeding events in patients taking warfarin.¹⁶ Patients who carry the CYP2C9*2 or CYP2C9*3 polymorphism metabolize S-warfarin slower and therefore have a threefold higher risk of hemorrhage after starting warfarin.¹⁷ We could speculate that patients carrying these variants may be better served by the newer anticoagulants, though this has not been tested in any clinical trial.

It is also worth appreciating that the conditions requiring anticoagulation, such as atrial fibrillation, also have a strong genetic basis. Variants in chromosomes 4q25, 1q21, and 16q22 have all been associated with atrial fibrillation.¹⁸ The risk of atrial fibrillation is five to six times higher in carriers of multiple variants within all of these loci.¹⁹ Genetic variants at 4q25 have been associated with the response to specific antiarrhythmic drug treatments,²⁰ response to pulmonary vein isolation, ^21,22 and direct-current cardioversion.²³ One can imagine a future in which patients with palpitations, carrying multiple gene risk variants, will choose prolonged monitoring at home to confirm a diagnosis. They would then be provided with a personalized best management strategy, using their personal preferences, clinical data, and genetic profile to make a treatment decision.

Using pharmacogenetics in prescribing clopidogrel and its alternatives

The pharmacogenetics of clopidogrel is of particular interest, as it has the potential of establishing a rational basis for using newer antiplatelet drugs such as ticagrelor and prasugrel, which are considerably more expensive than generic clopidogrel.

Most of the people who do not respond to clopidogrel carry the common cytochrome P450 2C19 variants CYP2C19*2 or CYP2C19*3.⁹ These variants are present in particularly high frequency in Asians and African Americans, who often do not feature in large randomized trials.

Newer antiplatelet agents have failed to demonstrate consistent superiority to clopidogrel without a tradeoff of more bleeding. However, in the Trial to Assess Improvement in Therapeutic Outcomes by Optimizing Platelet Inhibition With Prasugrel-Thrombolysis in Myocardial Infarction 39 (TRITON TIMI-38),^24,25 patients with the *2 variant receiving prasugrel had lower cardiovascular event rates than *2 carriers receiving clopidogrel.

Similarly, the patients who benefit the most from ticagrelor are carriers of the 2C19 nonresponder variants. In a large study, clopidogrel responders who did not carry either 2C19 nonresponder genetic variants or ABCB1 variants had cardiovascular outcomes similar to those of patients receiving ticagrelor.²⁶

Clinicians have been cautious in prescribing potent antiplatelet agents to all patients because of the risk of bleeding. One could assume that by reserving newer agents for clopidogrel nonresponders, the bleeding risk could be minimized and overall benefit could be preserved with this strategy.

Cost may also be contained. The cost-effectiveness of such an approach with prasugrel has been tested with computer modeling and appears favorable.²⁷ On the other hand, a similar yet limited analysis did not find genotype-driven use of ticagrelor to be cost-effective.²⁸ This was mostly due to fewer deaths in patients receiving ticagrelor. However, the cost estimate for genotype-guided therapy was overestimated, as heterozygotes in the model were treated with ticagrelor instead of a high dose of clopidogrel.

It now appears that heterozygotes, ie, patients with one copy of the nonresponder variant, can achieve similar platelet inhibition with clopidogrel 225 mg daily as noncarriers on 75 mg daily.²⁹ Since genotype-guided antiplatelet therapy has not been tested in a randomized outcomes trial, this tailored strategy has not been widely accepted.

THE FUTURE

The barriers to adoption of pharmacogenetics are considerable. Clinicians need to be educated about it, reimbursement needs to be worked out, and the pharmaceutical industry needs to get behind it. Nevertheless, the future of pharmacogenetics is extremely promising.

Research networks are forming to support the use of pharmacogenetics in clinical practice. The Pharmgkb (www.pharmgkb.org) database serves as a hub for educating clinicians and researchers as well as curating data for reference. Vanderbilt University is piloting the BioVu project, in which DNA and genotype data on patients are being stored and matched to the electronic clinical records.³⁰ These projects not only provide clinically useful information on the current state of the art of pharmacogenetics, they also aid in disseminating new information about genotype-phenotype relationships.

Analytical software that uses “natural language processing” is being applied to clinician-generated notes to derive new observations and associations between genetic variants and clinical phenotypes. Integrating this information in real-time decision-support modules in the electronic health record provides a feedback loop for a rapid assimilation of new knowledge. Similar innovative decision-support modules are being established by Cleveland Clinic’s Center for Personalized Healthcare.³¹

The rise of ‘omic’ sciences

Pharmacogenetics and pharmacogenomics are part of a larger set of “omic” sciences. The suffix “-omics” implies a larger, more holistic view and is being applied to a number of fields—for example, the study of proteins (proteomics) and the study of metabolites (metabolomics). Profiling proteins and metabolites delivers a deluge of information on a patient that can be clustered, using pattern-recognition software, into population subgroups. Patterns of multiple proteins or metabolites are extracted from this spectral data to identify disease or response to treatment (pharmacometabolomics).

Metabolomics has been shown to predict the response to statins,³² diagnose myocardial infarction,³³ and reclassify cardiovascular risk status.³⁴ In addition, whereas traditional laboratory chemistry is reductionist, using single biomarkers for single-disease diagnosis, omic technologies hold the potential to reveal information on a number of possible health or disease states. The identification of “healthy” profiles using these technologies can potentially provide positive feedback to patients undertaking lifestyle changes and treatment.

The instrument costs for proteomic and metabolomic profiling are relatively high. However, the ongoing running costs are minimal, estimated at as low as less than $13 per test, as there are no expensive reagents.³³ High-volume testing therefore becomes very cost-effective.

Although omic science appears futuristic, proteomics and metabolomics are already used in many clinical laboratories to rapidly identify bacteria. These methods have already revolutionized the way laboratories identify microbes, since they are automated, reduce workload, and give very fast results.

The cost of genetic testing is falling

Critics of pharmacogenetics claim that the predictive value of genetic testing is poor, that evidence is lacking, and that the cost is too high. In all new technologies, the first iteration is coarse, but performance improves with use. The first major barrier is adoption. Projects like BioVu are establishing the infrastructure for a feedback loop to iteratively improve upon the status quo and provide the evidence base clinicians demand.

The cost of genetic testing is falling rapidly, with whole-genome sequencing and annotation now costing less than $5,000. The cost of a pharmacogenetic test can be as low as $100 using low-cost nanotechnology, and the test needs to be performed only once in a patient’s lifetime.²⁷

As other related molecular technologies such as proteomics and metabolomics become available and are integrated with genomics, the predictive ability of this science will improve.

AWAY FROM ONE-SIZE-FITS-ALL MEDICINE

Over the last decade there has been a trend away from “one size fits all” to customized “markets of one” in everything from consumer products to education to medicine. Mass customizing, also known as personalization, has been embraced by the internet community as a means to increase efficiency and reduce cost. This occurs by eliminating waste in redundant work or production of ineffective products.

Personalization on the Internet has been enabled through the use of informatics, mathematics, and supercomputing. The same tools that have personalized the delivery of consumer products are also being applied to the field of pharmacogenetics. Applied in an evidence-based fashion, these new technologies should profoundly improve patient care now and in the future.

“Change is the only constant.”

—Heraclitus (c 535–475 bce)

With the cost of health care rising and money to pay for it shrinking, there has never been a greater need to reduce waste.

See related article

Ineffective treatments and adverse drug effects account for much preventable morbidity and expense. New treatments, touted as more potent, are often introduced as replacements for traditional ones that are still effective in many patients, adding to costs and the potential for harm. For the pharmaceutical industry, the search for new “blockbuster” drugs seems to have hit a wall, at least in cardiovascular medicine.¹ Advances often come at the cost of adverse effects, such as bleeding with triple antiplatelet therapy and diabetes with potent statin drugs.

The path to maximizing benefit and reducing harm now appears to lie in stratifying populations and appreciating patient individuality in response to treatment. For many decades we have known that patients vary widely in their response to drugs, owing to personal factors such as body surface area, age, environment, and genetics. And indeed, we treat our patients as individuals, for example by tailoring aminoglycoside dose to weight and renal function.

However, clinical trials typically give us an idea of the benefits only to the average patient. While subgroup analyses identify groups that may benefit more or less from treatment, the additional information they provide is not easily integrated into the clinical model of prescribing, in which one size fits all.

THE PROMISE OF PHARMACOGENETICS

The emerging field of pharmacogenetics promises to give clinicians the tools to make informed treatment decisions based on predictive genetic testing. This genetic testing aims to match treatment to an individual’s genetic profile. This often involves analyzing single-nucleotide polymorphisms in genes for enzymes that metabolize drugs, such as the cytochrome P450 enzymes, to predict efficacy or an adverse event with treatment.

Pharmacogenetics is playing an increasing role in clinical trials, particularly in the early stages of drug development, by helping to reduce the number of patients needed, prove efficacy, and identify subgroups in which alternative treatment can be targeted. At another level, a molecular understanding of disease is leading to truly targeted treatments based on genomics.

Over recent years, genetic testing has been increasingly used in clinical practice, thanks to a convergence of factors such as rapid, low-cost tests, a growing evidence base, and emerging interest among doctors and payers.

An advantage to using genetic testing as opposed to other types of laboratory testing, such as measuring the concentration of the drug in the blood during treatment, is that genetic tests can predict the response to treatment before the treatment is started. Moreover, with therapeutic drug monitoring after treatment has begun, there are sometimes no detectable measures of toxicity. For example, both carbamazepine and the antiviral drug abacavir can—fortunately only rarely—cause Stevens-Johnson syndrome. But before genetic markers were discovered, there was no method of estimating this risk apart from taking a family history.^2,3 Considering the numbers of people involved, it was not feasible until recently to suggest genetic screening for patients starting on these drugs. However, the cost of genotyping and gene sequencing has been falling at a rate inversely faster than Moore’s law (an approximate annual doubling in computer power), and population genomics is becoming a reality.⁴

The US Food and Drug Administration (FDA) recognizes the current and future value of pharmacogenetics in drug safety and development. A number of approved pharmacogenetic biomarkers are listed on the FDA website (Table 1). Black box warnings have been mandated for a number of drugs on the basis of observational evidence.

The FDA also promotes rapid approval for novel drugs with pharmacogenetic “companion diagnostics.” A recent example of this was the approval of ivacaftor for cystic fibrosis patients who have the G551D mutation.⁵ Here, a molecular understanding of the condition led to the development of a targeted treatment. Although the cost of developing this drug was high, the path is now paved for similar advances. Oncologists are familiar with these advances with the emergence of new molecularly targeted treatments, eg, BRAF inhibitors in metastatic melanoma, imatinib in chronic myeloid leukemia, and gefitinib in non-small-cell lung cancer.

PHARMACOGENETICS IN CARDIOVASCULAR MEDICINE

Cardiovascular medicine also stands to benefit from rapid advances in pharmacogenetics.

While no treatment has been developed that targets the molecular basis of cardiovascular disease, a number of genomic biomarkers have emerged that identify patients at risk of adverse reactions or treatment failure. These include genetic tests to predict the maintenance dose and risk of bleeding with warfarin,⁶ the likelihood of myopathy and myositis with simvastatin,⁷ and the risk of recurrent thrombotic events with clopidogrel.^8–10

Using pharmacogenetics in prescribing warfarin and its alternatives

The pharmacogenetics of warfarin has been extensively researched, but genotyping before prescribing this drug is not yet widely done.

In 2007, the FDA updated the labeling of warfarin to include information about the influence of two genes, VKORC1 and CYP2C9, on a patient’s response to this drug. In 2010 this was updated to add that testing for these genes could be used to predict the maintenance dose of the drug. Difficulties with algorithms used to integrate this into clinical practice have hindered adoption of this testing.

With the advent of new anticoagulants such as dabigatran, rivaroxaban, and apixaban, many have expected warfarin and its pharmacogenetics to become obsolete. However, the new agents cost considerably more. Further, they may not offer a very great advantage over warfarin: in the Randomized Evaluation of Long-term Anticoagulant Therapy (RE-LY) trial, the absolute risk reduction in intracranial hemorrhage with dabigatran vs warfarin was small.¹¹ Therefore, dabigatran is probably not cost-effective in populations at low risk of bleeding.¹² A cost-effectiveness analysis comparing warfarin with dabigatran in patients with uncomplicated atrial fibrillation has suggested that dabigatran is, however, cost-effective in patients at moderate risk.¹²

In the RE-LY trial, the international normalized ratios (INRs) of the patients in the warfarin group were in the therapeutic range only 64% of the time. The advantages of dabigatran over warfarin become less pronounced as warfarin control is tightened.¹³ Of note, pharmacogenetics and home monitoring of the INR have both been shown to lead to tighter control of the INR, with greater time within the therapeutic range.^14,15

Moreover, genetic testing can help us reduce the number of bleeding events in patients taking warfarin.¹⁶ Patients who carry the CYP2C9*2 or CYP2C9*3 polymorphism metabolize S-warfarin slower and therefore have a threefold higher risk of hemorrhage after starting warfarin.¹⁷ We could speculate that patients carrying these variants may be better served by the newer anticoagulants, though this has not been tested in any clinical trial.

It is also worth appreciating that the conditions requiring anticoagulation, such as atrial fibrillation, also have a strong genetic basis. Variants in chromosomes 4q25, 1q21, and 16q22 have all been associated with atrial fibrillation.¹⁸ The risk of atrial fibrillation is five to six times higher in carriers of multiple variants within all of these loci.¹⁹ Genetic variants at 4q25 have been associated with the response to specific antiarrhythmic drug treatments,²⁰ response to pulmonary vein isolation, ^21,22 and direct-current cardioversion.²³ One can imagine a future in which patients with palpitations, carrying multiple gene risk variants, will choose prolonged monitoring at home to confirm a diagnosis. They would then be provided with a personalized best management strategy, using their personal preferences, clinical data, and genetic profile to make a treatment decision.

Using pharmacogenetics in prescribing clopidogrel and its alternatives

The pharmacogenetics of clopidogrel is of particular interest, as it has the potential of establishing a rational basis for using newer antiplatelet drugs such as ticagrelor and prasugrel, which are considerably more expensive than generic clopidogrel.

Most of the people who do not respond to clopidogrel carry the common cytochrome P450 2C19 variants CYP2C19*2 or CYP2C19*3.⁹ These variants are present in particularly high frequency in Asians and African Americans, who often do not feature in large randomized trials.

Newer antiplatelet agents have failed to demonstrate consistent superiority to clopidogrel without a tradeoff of more bleeding. However, in the Trial to Assess Improvement in Therapeutic Outcomes by Optimizing Platelet Inhibition With Prasugrel-Thrombolysis in Myocardial Infarction 39 (TRITON TIMI-38),^24,25 patients with the *2 variant receiving prasugrel had lower cardiovascular event rates than *2 carriers receiving clopidogrel.

Similarly, the patients who benefit the most from ticagrelor are carriers of the 2C19 nonresponder variants. In a large study, clopidogrel responders who did not carry either 2C19 nonresponder genetic variants or ABCB1 variants had cardiovascular outcomes similar to those of patients receiving ticagrelor.²⁶

Clinicians have been cautious in prescribing potent antiplatelet agents to all patients because of the risk of bleeding. One could assume that by reserving newer agents for clopidogrel nonresponders, the bleeding risk could be minimized and overall benefit could be preserved with this strategy.

Cost may also be contained. The cost-effectiveness of such an approach with prasugrel has been tested with computer modeling and appears favorable.²⁷ On the other hand, a similar yet limited analysis did not find genotype-driven use of ticagrelor to be cost-effective.²⁸ This was mostly due to fewer deaths in patients receiving ticagrelor. However, the cost estimate for genotype-guided therapy was overestimated, as heterozygotes in the model were treated with ticagrelor instead of a high dose of clopidogrel.