Multicenter Prospective Cohort Study of the Diagnostic Yield and Patient Experience of Multiplex Gene Panel Testing For Hereditary Cancer Risk.

PURPOSE: Multiplex gene panel testing (MGPT) allows for the simultaneous analysis of germline cancer susceptibility genes. This study describes the diagnostic yield and patient experiences of MGPT in diverse populations. PATIENTS AND METHODS: This multicenter, prospective cohort study enrolled participants from three cancer genetics clinics-University of Southern California Norris Comprehensive Cancer Center, Los Angeles County and University of Southern California Medical Center, and Stanford Cancer Institute-who met testing guidelines or had a 2.5% or greater probability of a pathogenic variant (N = 2,000). All patients underwent 25- or 28-gene MGPT and results were compared with differential genetic diagnoses generated by pretest expert clinical assessment. Post-test surveys on distress, uncertainty, and positive experiences were administered at 3 months (69% response rate) and 1 year (57% response rate). RESULTS: Of 2,000 participants, 81% were female, 41% were Hispanic, 26% were Spanish speaking only, and 30% completed high school or less education. A total of 242 participants (12%) carried one or more pathogenic variant (positive), 689 (34%) carried one or more variant of uncertain significance (VUS), and 1,069 (53%) carried no pathogenic variants or VUS (negative). More than one third of pathogenic variants (34%) were not included in the differential diagnosis. After testing, few patients (4%) had prophylactic surgery, most (92%) never regretted testing, and most (80%) wanted to know all results, even those of uncertain significance. Positive patients were twice as likely as negative/VUS patients (83% v 41%; P < .001) to encourage their relatives to be tested. CONCLUSION: In a racially/ethnically and socioeconomically diverse cohort, MGPT increased diagnostic yield. More than one third of identified pathogenic variants were not clinically anticipated. Patient regret and prophylactic surgery use were low, and patients appropriately encouraged relatives to be tested for clinically relevant results.

Prevalence of germ-line mutations in cancer genes among pancreatic cancer patients with a positive family history.

PurposePanel-based genetic testing has identified increasing numbers of patients with pancreatic ductal adenocarcinoma (PDAC) who carry germ-line mutations. However, small sample sizes or number of genes evaluated limit prevalence estimates of these mutations. We estimated prevalence of mutations in PDAC patients with positive family history.MethodsWe sequenced 25 cancer susceptibility genes in lymphocyte DNA from 302 PDAC patients in the Mayo Clinic Biospecimen Resource for Pancreatic Research Registry. Kindreds containing at least two first-degree relatives with PDAC met criteria for familial pancreatic cancer (FPC), while the remaining were familial, but not FPC.ResultsThirty-six patients (12%) carried at least one deleterious mutation in one of 11 genes. Of FPC patients, 25/185 (14%) were carriers, while 11/117 (9%) non-FPC patients with family history were carriers. Deleterious mutations (n) identified in PDAC patients were BRCA2 (11), ATM (8), CDKN2A (4), CHEK2 (4), MUTYH/MYH (3 heterozygotes, not biallelic), BRCA1 (2), and 1 each in BARD1, MSH2, NBN, PALB2, and PMS2. Novel mutations were found in ATM, BARD1, and PMS2.ConclusionMultiple susceptibility gene testing in PDAC patients with family history of pancreatic cancer is warranted regardless of FPC status and will inform genetic risk counseling for families.

Community Practice Implementation of a Self-administered Version of PREMM1,2,6 to Assess Risk for Lynch Syndrome.

BACKGROUND & AIMS: Lynch syndrome is a genetic disorder that greatly increases risk for colorectal and other cancers, although it is underdiagnosed. Prediction of MLH1, MSH2, and MSH6 (PREMM1,2,6) is a web-based tool that analyzes individuals' personal/family histories of cancer to quantify their likelihood of carrying a germline mutation associated with Lynch syndrome. We investigated the feasibility of systematic risk assessment for Lynch syndrome in a community gastroenterology practice using a patient-completed version of PREMM1,2,6. METHODS: PREMM1,2,6 was adapted into a computer tablet version designed for self-administration by patients. Individuals presenting to a community gastroenterology office and endoscopy facility in California completed the PREMM1,2,6 assessment before their visit (n = 3134). The total study duration (8 months) comprised a 2-month initiation period (May 1-June 30, 2013) and a 6-month study period (July 1-December 31, 2013). Genetic counseling and germline analysis for mutations in genes associated with Lynch syndrome (MLH1, MSH2, MSH6, PMS2, and EPCAM) were offered to individuals with PREMM1,2,6 scores of 5% or higher. Patients and providers completed surveys to evaluate the feasibility and satisfaction with the process. RESULTS: Of the 3134 individuals assessed by PREMM1,2,6 during the 6-month study period, 177 individuals (5.6%) had scores of 5% or higher. Of these, 146 individuals underwent genetic testing, along with 28 additional participants recruited nonconsecutively during the initiation period. Mutations associated with Lynch syndrome were detected in 3 of the 146 individuals (2.1%) with PREMM1,2,6 scores of 5% or higher who underwent germline testing, and 3 of the 28 patients (10.7%) recruited during study initiation with PREMM1,2,6 scores of 5% or higher. Of the participants who underwent genetic analysis, 98.6% stated that they understood the information provided to them. All of the surveyed providers stated that they were satisfied with the incorporation of PREMM1,2,6 into their clinical practice, and that they would continue using it to assess risk for Lynch syndrome. CONCLUSIONS: A patient self-administered version of the PREMM1,2,6 Lynch syndrome risk assessment model can be used systematically in community-based gastroenterology and endoscopy practices.

Diagnostic Distinction of Malignant Melanoma and Benign Nevi by a Gene Expression Signature and Correlation to Clinical Outcomes.

Background: Histopathologic examination alone can be inadequate for diagnosis of certain melanocytic neoplasms. Recently, a 23-gene expression signature was clinically validated as an ancillary diagnostic test to differentiate benign nevi from melanoma. The current study assessed the performance of this test in an independent cohort of melanocytic lesions against clinically proven outcomes.Methods: Archival tissue from primary cutaneous melanomas and melanocytic nevi was obtained from four independent institutions and tested with the gene signature. Cases were selected according to pre-defined clinical outcome measures. Malignant lesions were defined as stage I-III primary cutaneous melanomas that produced distant metastases (metastatic to sites other than proximal sentinel lymph node(s)) following diagnosis of the primary lesion. Melanomas that were metastatic at the time of diagnosis, all re-excisions, and lesions with <10% tumor volume were excluded. Benign lesions were defined as cutaneous melanocytic lesions with no adverse long-term events reported.Results: Of 239 submitted samples, 182 met inclusion criteria and produced a valid gene expression result. This included 99 primary cutaneous melanomas with proven distant metastases and 83 melanocytic nevi. Median time to melanoma metastasis was 18 months. Median follow-up time for nevi was 74.9 months. The gene expression score differentiated melanoma from nevi with a sensitivity of 93.8% and a specificity of 96.2%.Conclusions: The results of gene expression testing closely correlate with long-term clinical outcomes of patients with melanocytic neoplasms.Impact: Collectively, this provides strong evidence that the gene signature adds valuable adjunctive information to aid in the accurate diagnosis of melanoma. Cancer Epidemiol Biomarkers Prev; 26(7); 1107-13. ©2017 AACR.

Cancer Susceptibility Gene Mutations in Individuals With Colorectal Cancer.

Purpose Hereditary factors play an important role in colorectal cancer (CRC) risk, yet the prevalence of germline cancer susceptibility gene mutations in patients with CRC unselected for high-risk features (eg, early age at diagnosis, personal/family history of cancer or polyps, tumor microsatellite instability [MSI], mismatch repair [MMR] deficiency) is unknown. Patients and Methods We recruited 1,058 participants who received CRC care in a clinic-based setting without preselection for age at diagnosis, personal/family history, or MSI/MMR results. All participants underwent germline testing for mutations in 25 genes associated with inherited cancer risk. Each gene was categorized as high penetrance or moderate penetrance on the basis of published estimates of the lifetime cancer risks conferred by pathogenic germline mutations in that gene. Results One hundred five (9.9%; 95% CI, 8.2% to 11.9%) of 1,058 participants carried one or more pathogenic mutations, including 33 (3.1%) with Lynch syndrome (LS). Twenty-eight (96.6%) of 29 available LS CRCs demonstrated abnormal MSI/MMR results. Seventy-four (7.0%) of 1,058 participants carried non-LS gene mutations, including 23 (2.2%) with mutations in high-penetrance genes (five APC, three biallelic MUTYH, 11 BRCA1/2, two PALB2, one CDKN2A, and one TP53), 15 of whom lacked clinical histories suggestive of their underlying mutation. Thirty-eight (3.6%) participants had moderate-penetrance CRC risk gene mutations (19 monoallelic MUTYH, 17 APC*I1307K, two CHEK2). Neither proband age at CRC diagnosis, family history of CRC, nor personal history of other cancers significantly predicted the presence of pathogenic mutations in non-LS genes. Conclusion Germline cancer susceptibility gene mutations are carried by 9.9% of patients with CRC. MSI/MMR testing reliably identifies LS probands, although 7.0% of patients with CRC carry non-LS mutations, including 1.0% with BRCA1/2 mutations.

Assessment of in silico protein sequence analysis in the clinical classification of variants in cancer risk genes.

Missense variants represent a significant proportion of variants identified in clinical genetic testing. In the absence of strong clinical or functional evidence, the American College of Medical Genetics recommends that these findings be classified as variants of uncertain significance (VUS). VUSs may be reclassified to better inform patient care when new evidence is available. It is critical that the methods used for reclassification are robust in order to prevent inappropriate medical management strategies and unnecessary, life-altering surgeries. In an effort to provide evidence for classification, several in silico algorithms have been developed that attempt to predict the functional impact of missense variants through amino acid sequence conservation analysis. We report an analysis comparing internally derived, evidence-based classifications with the results obtained from six commonly used algorithms. We compiled a dataset of 1118 variants in BRCA1, BRCA2, MLH1, and MSH2 previously classified by our laboratory's evidence-based variant classification program. We compared internally derived classifications with those obtained from the following in silico tools: Align-GVGD, CONDEL, Grantham Analysis, MAPP-MMR, PolyPhen-2, and SIFT. Despite being based on similar underlying principles, all algorithms displayed marked divergence in accuracy, specificity, and sensitivity. Overall, accuracy ranged from 58.7 to 90.8% while the Matthews Correlation Coefficient ranged from 0.26-0.65. CONDEL, a weighted average of multiple algorithms, did not perform significantly better than its individual components evaluated here. These results suggest that the in silico algorithms evaluated here do not provide reliable evidence regarding the clinical significance of missense variants in genes associated with hereditary cancer.

An independent validation of a gene expression signature to differentiate malignant melanoma from benign melanocytic nevi.

BACKGROUND: Recently, a 23-gene signature was developed to produce a melanoma diagnostic score capable of differentiating malignant and benign melanocytic lesions. The primary objective of this study was to independently assess the ability of the gene signature to differentiate melanoma from benign nevi in clinically relevant lesions. METHODS: A set of 1400 melanocytic lesions was selected from samples prospectively submitted for gene expression testing at a clinical laboratory. Each sample was tested and subjected to an independent histopathologic evaluation by 3 experienced dermatopathologists. A primary diagnosis (benign or malignant) was assigned to each sample, and diagnostic concordance among the 3 dermatopathologists was required for inclusion in analyses. The sensitivity and specificity of the score in differentiating benign and malignant melanocytic lesions were calculated to assess the association between the score and the pathologic diagnosis. RESULTS: The gene expression signature differentiated benign nevi from malignant melanoma with a sensitivity of 91.5% and a specificity of 92.5%. CONCLUSIONS: These results reflect the performance of the gene signature in a diverse array of samples encountered in routine clinical practice. Cancer 2017;123:617-628. © 2016 American Cancer Society.

Frequency of Germline Mutations in 25 Cancer Susceptibility Genes in a Sequential Series of Patients With Breast Cancer.

PURPOSE: Testing for germline mutations in BRCA1/2 is standard for select patients with breast cancer to guide clinical management. Next-generation sequencing (NGS) allows testing for mutations in additional breast cancer predisposition genes. The frequency of germline mutations detected by using NGS has been reported in patients with breast cancer who were referred for BRCA1/2 testing or with triple-negative breast cancer. We assessed the frequency and predictors of mutations in 25 cancer predisposition genes, including BRCA1/2, in a sequential series of patients with breast cancer at an academic institution to examine the utility of genetic testing in this population. METHODS: Patients with stages I to III breast cancer who were seen at a single cancer center between 2010 and 2012, and who agreed to participate in research DNA banking, were included (N = 488). Personal and family cancer histories were collected and germline DNA was sequenced with NGS to identify mutations. RESULTS: Deleterious mutations were identified in 10.7% of women, including 6.1% in BRCA1/2 (5.1% in non-Ashkenazi Jewish patients) and 4.6% in other breast/ovarian cancer predisposition genes including CHEK2 (n = 10), ATM (n = 4), BRIP1 (n = 4), and one each in PALB2, PTEN, NBN, RAD51C, RAD51D, MSH6, and PMS2. Whereas young age (P < .01), Ashkenazi Jewish ancestry (P < .01), triple-negative breast cancer (P = .01), and family history of breast/ovarian cancer (P = .01) predicted for BRCA1/2 mutations, no factors predicted for mutations in other breast cancer predisposition genes. CONCLUSION: Among sequential patients with breast cancer, 10.7% were found to have a germline mutation in a gene that predisposes women to breast or ovarian cancer, using a panel of 25 predisposition genes. Factors that predict for BRCA1/2 mutations do not predict for mutations in other breast/ovarian cancer susceptibility genes when these genes are analyzed as a single group. Additional cohorts will be helpful to define individuals at higher risk of carrying mutations in genes other than BRCA1/2.

Patients Tested at a Laboratory for Hereditary Cancer Syndromes Show an Overlap for Multiple Syndromes in Their Personal and Familial Cancer Histories.

OBJECTIVE: Hereditary cancer testing guidelines are based on the premise that the common hereditary cancer syndromes have distinct, recognizable phenotypes. However, many syndromes present with overlapping cancers. The aim of this analysis was to identify the proportion of patients tested for Lynch syndrome (LS) or hereditary breast and ovarian cancer (HBOC) who met testing criteria for the other syndrome. METHOD: We analyzed a commercial laboratory database of patients tested for LS and HBOC in a clinical setting from 2006 to 2013. Patient cancer histories were analyzed using the 2012 NCCN criteria for LS and the 2013 NCCN criteria for HBOC. RESULTS: In all, 7% of the patients tested for HBOC met criteria for LS testing. The majority of these patients had a family history of colorectal (30.9%) and/or endometrial cancer (22.7%). Conversely, 29.5% of the patients tested for LS met criteria for HBOC testing. In this group, 30.5% of the patients had a personal history of breast cancer, and 12.6% had a personal history of ovarian cancer. CONCLUSIONS: Our data demonstrate a substantial phenotypic overlap among patients for multiple common inherited cancer syndromes, which likely complicates diagnosis and test selection. This supports the value of multigene panels to identify pathogenic mutations in the absence of a clinically specific phenotype.

Identification of a Variety of Mutations in Cancer Predisposition Genes in Patients With Suspected Lynch Syndrome.

BACKGROUND & AIMS: Multigene panels are commercially available tools for hereditary cancer risk assessment that allow for next-generation sequencing of numerous genes in parallel. However, it is not clear if these panels offer advantages over traditional genetic testing. We investigated the number of cancer predisposition gene mutations identified by parallel sequencing in individuals with suspected Lynch syndrome. METHODS: We performed germline analysis with a 25-gene, next-generation sequencing panel using DNA from 1260 individuals who underwent clinical genetic testing for Lynch syndrome from 2012 through 2013. All patients had a history of Lynch syndrome-associated cancer and/or polyps. We classified all identified germline alterations for pathogenicity and calculated the frequencies of pathogenic mutations and variants of uncertain clinical significance (VUS). We also analyzed data on patients' personal and family history of cancer, including fulfillment of clinical guidelines for genetic testing. RESULTS: Of the 1260 patients, 1112 met National Comprehensive Cancer Network (NCCN) criteria for Lynch syndrome testing (88%; 95% confidence interval [CI], 86%-90%). Multigene panel testing identified 114 probands with Lynch syndrome mutations (9.0%; 95% CI, 7.6%-10.8%) and 71 with mutations in other cancer predisposition genes (5.6%; 95% CI, 4.4%-7.1%). Fifteen individuals had mutations in BRCA1 or BRCA2; 93% of these met the NCCN criteria for Lynch syndrome testing and 33% met NCCN criteria for BRCA1 and BRCA2 analysis (P = .0017). An additional 9 individuals carried mutations in other genes linked to high lifetime risks of cancer (5 had mutations in APC, 3 had bi-allelic mutations in MUTYH, and 1 had a mutation in STK11); all of these patients met NCCN criteria for Lynch syndrome testing. A total of 479 individuals had 1 or more VUS (38%; 95% CI, 35%-41%). CONCLUSIONS: In individuals with suspected Lynch syndrome, multigene panel testing identified high-penetrance mutations in cancer predisposition genes, many of which were unexpected based on patients' histories. Parallel sequencing also detected a high number of potentially uninformative germline findings, including VUS.

Erratum to: Incidence of BRCA1 and BRCA2 non-founder mutations in patients of Ashkenazi Jewish ancestry.

In Table 2 of the original publication, the HGVS and legacy nomenclature were mismatched and the HGVS nomenclature did not correlate with data listed in the table. The corrected table is listed below.

Analytical validation of a melanoma diagnostic gene signature using formalin-fixed paraffin-embedded melanocytic lesions.

AIM: These studies were to validate the analytical performance of a gene expression signature that differentiates melanoma and nevi, using RNA expression from 14 signature genes and nine normalization genes that generates a melanoma diagnostic score (MDS). MATERIALS & METHODS: Formalin-fixed paraffin-embedded melanocytic lesions were evaluated in these studies. RESULTS: The overall SD of the assay was determined to be 0.69 MDS units. Individual amplicons within the signature had an average amplification efficiency of 92% and a SD less than 0.5 CT. The MDS was reproducible across a 2000-fold dilution range of input RNA. Melanin, an inhibitor of PCR, does not interfere with the signature. CONCLUSION: These studies indicate this signature is robust and reproducible and is analytically validated on formalin-fixed paraffin-embedded melanocytic lesions.

Targeted deletion of collagen V in tendons and ligaments results in a classic Ehlers-Danlos syndrome joint phenotype.

Collagen V mutations underlie classic Ehlers-Danlos syndrome, and joint hypermobility is an important clinical manifestation. We define the function of collagen V in tendons and ligaments, as well as the role of alterations in collagen V expression in the pathobiology in classic Ehlers-Danlos syndrome. A conditional Col5a1(flox/flox) mouse model was bred with Scleraxis-Cre mice to create a targeted tendon and ligament Col5a1-null mouse model, Col5a1(Δten/Δten). Targeting was specific, resulting in collagen V-null tendons and ligaments. Col5a1(Δten/Δten) mice demonstrated decreased body size, grip weakness, abnormal gait, joint laxity, and early-onset osteoarthritis. These gross changes were associated with abnormal fiber organization, as well as altered collagen fibril structure with increased fibril diameters and decreased fibril number that was more severe in a major joint stabilizing ligament, the anterior cruciate ligament (ACL), than in the flexor digitorum longus tendon. The ACL also had a higher collagen V content than did the flexor digitorum longus tendon. The collagen V-null ACL and flexor digitorum longus tendon both had significant alterations in mechanical properties, with ACL exhibiting more severe changes. The data demonstrate critical differential regulatory roles for collagen V in tendon and ligament structure and function and suggest that collagen V regulatory dysfunction is associated with an abnormal joint phenotype, similar to the hypermobility phenotype in classic Ehlers-Danlos syndrome.

Clinical validation of a gene expression signature that differentiates benign nevi from malignant melanoma.

BACKGROUND: Histopathologic examination is sometimes inadequate for accurate and reproducible diagnosis of certain melanocytic neoplasms. As a result, more sophisticated and objective methods have been sought. The goal of this study was to identify a gene expression signature that reliably differentiated benign and malignant melanocytic lesions and evaluate its potential clinical applicability. Herein, we describe the development of a gene expression signature and its clinical validation using multiple independent cohorts of melanocytic lesions representing a broad spectrum of histopathologic subtypes. METHODS: Using quantitative reverse-transcription polymerase chain reaction (PCR) on a selected set of 23 differentially expressed genes, and by applying a threshold value and weighting algorithm, we developed a gene expression signature that produced a score that differentiated benign nevi from malignant melanomas. RESULTS: The gene expression signature classified melanocytic lesions as benign or malignant with a sensitivity of 89% and a specificity of 93% in a training cohort of 464 samples. The signature was validated in an independent clinical cohort of 437 samples, with a sensitivity of 90% and specificity of 91%. CONCLUSIONS: The performance, objectivity, reliability and minimal tissue requirements of this test suggest that it could have clinical application as an adjunct to histopathology in the diagnosis of melanocytic neoplasms.

Comparison of locus-specific databases for BRCA1 and BRCA2 variants reveals disparity in variant classification within and among databases.

Genetic variants of uncertain clinical significance (VUSs) are a common outcome of clinical genetic testing. Locus-specific variant databases (LSDBs) have been established for numerous disease-associated genes as a research tool for the interpretation of genetic sequence variants to facilitate variant interpretation via aggregated data. If LSDBs are to be used for clinical practice, consistent and transparent criteria regarding the deposition and interpretation of variants are vital, as variant classifications are often used to make important and irreversible clinical decisions. In this study, we performed a retrospective analysis of 2017 consecutive BRCA1 and BRCA2 genetic variants identified from 24,650 consecutive patient samples referred to our laboratory to establish an unbiased dataset representative of the types of variants seen in the US patient population, submitted by clinicians and researchers for BRCA1 and BRCA2 testing. We compared the clinical classifications of these variants among five publicly accessible BRCA1 and BRCA2 variant databases: BIC, ClinVar, HGMD (paid version), LOVD, and the UMD databases. Our results show substantial disparity of variant classifications among publicly accessible databases. Furthermore, it appears that discrepant classifications are not the result of a single outlier but widespread disagreement among databases. This study also shows that databases sometimes favor a clinical classification when current best practice guidelines (ACMG/AMP/CAP) would suggest an uncertain classification. Although LSDBs have been well established for research applications, our results suggest several challenges preclude their wider use in clinical practice.

Validation of a molecular and pathological model for five-year mortality risk in patients with early stage lung adenocarcinoma.

INTRODUCTION: The aim of this study was to validate a molecular expression signature [cell cycle progression (CCP) score] that identifies patients with a higher risk of cancer-related death after surgical resection of early stage (I-II) lung adenocarcinoma in a large patient cohort and evaluate the effectiveness of combining CCP score and pathological stage for predicting lung cancer mortality. METHODS: Formalin-fixed paraffin-embedded surgical tumor samples from 650 patients diagnosed with stage I and II adenocarcinoma who underwent definitive surgical treatment without adjuvant chemotherapy were analyzed for 31 proliferation genes by quantitative real-time polymerase chain reaction. The prognostic discrimination of the expression score was assessed by Cox proportional hazards analysis using 5-year lung cancer-specific death as primary outcome. RESULTS: The CCP score was a significant predictor of lung cancer-specific mortality above clinical covariates [hazard ratio (HR) = 1.46 per interquartile range (95% confidence interval = 1.12-1.90; p = 0.0050)]. The prognostic score, a combination of CCP score and pathological stage, was a more significant indicator of lung cancer mortality risk than pathological stage in the full cohort (HR = 2.01; p = 2.8 × 10) and in stage I patients (HR = 1.67; p = 0.00027). Using the 85th percentile of the prognostic score as a threshold, there was a significant difference in lung cancer survival between low-risk and high-risk patient groups (p = 3.8 × 10). CONCLUSIONS: This study validates the CCP score and the prognostic score as independent predictors of lung cancer death in patients with early stage lung adenocarcinoma treated with surgery alone. Patients with resected stage I lung adenocarcinoma and a high prognostic score may be candidates for adjuvant therapy to reduce cancer-related mortality.

Frequency of mutations in individuals with breast cancer referred for BRCA1 and BRCA2 testing using next-generation sequencing with a 25-gene panel.

BACKGROUND: Next-generation sequencing (NGS) allows for simultaneous sequencing of multiple cancer susceptibility genes and, for an individual, may be more efficient and less expensive than sequential testing. The authors assessed the frequency of deleterious germline mutations among individuals with breast cancer who were referred for BRCA1 and BRCA2 (BRCA1/2) gene testing using a panel of 25 genes associated with inherited cancer predisposition. METHODS: This was a cross-sectional study using NGS in 2158 individuals, including 1781 who were referred for commercial BRCA1/2 gene testing (cohort 1) and 377 who had detailed personal and family history and had previously tested negative for BRCA1/2 mutations (cohort 2). RESULTS: Mutations were identified in 16 genes, most frequently in BRCA1, BRCA2, CHEK2, ATM, and PALB2. Among the participants in cohort 1, 9.3% carried a BRCA1/2 mutation, 3.9% carried a mutation in another breast/ovarian cancer susceptibility gene, and 0.3% carried an incidental mutation in another cancer susceptibility gene unrelated to breast or ovarian cancer. In cohort 2, the frequency of mutations in breast/ovarian-associated genes other than BRCA1/2 was 2.9%, and an additional 0.8% had an incidental mutation. In cohort 1, Lynch syndrome-related mutations were identified in 7 individuals. In contrast to BRCA1/2 mutations, neither age at breast cancer diagnosis nor family history of ovarian or young breast cancer predicted for other mutations. The frequency of mutations in genes other than BRCA1/2 was lower in Ashkenazi Jews compared with non-Ashkenazi individuals (P=.026). CONCLUSIONS: Using an NGS 25-gene panel, the frequency of mutations in genes other than BRCA1/2 was 4.3%, and most mutations (3.9%) were identified in genes associated with breast/ovarian cancer.

BRCA1, BRCA2, PALB2, and CDKN2A mutations in familial pancreatic cancer: a PACGENE study.

PURPOSE: Familial pancreatic cancer kindreds contain at least two affected first-degree relatives. Comprehensive data are needed to assist clinical risk assessment and genetic testing. METHODS: Germ-line DNA samples from 727 unrelated probands with positive family history (521 met criteria for familial pancreatic cancer) were tested in compliance with the Clinical Laboratory Improvement Amendments for mutations in BRCA1 and BRCA2 (including analysis of deletions and rearrangements), PALB2, and CDKN2A. We compared prevalence of deleterious mutations between familial pancreatic cancer probands and nonfamilial pancreatic cancer probands (kindreds containing at least two affected biological relatives, but not first-degree relatives). We also examined the impact of family history on breast and ovarian cancers and melanoma. RESULTS: Prevalence of deleterious mutations (excluding variants of unknown significance) among familial pancreatic cancer probands was: BRCA1, 1.2%; BRCA2, 3.7%; PALB2, 0.6%; and CDKN2A, 2.5%. Four novel deleterious mutations were detected. Familial pancreatic cancer probands carry more mutations in the four genes (8.0%) than nonfamilial pancreatic cancer probands (3.5%) (odds ratio: 2.40; 95% confidence interval: 1.06-5.44; P = 0.03). The probability of testing positive for deleterious mutations in any of the four genes ranges up to 10.4%, depending on family history of cancers. BRCA2 and CDKN2A account for the majority of mutations in familial pancreatic cancer. CONCLUSION: Genetic testing of multiple relevant genes in probands with a positive family history is warranted, particularly for familial pancreatic cancer.

Incidence of BRCA1 and BRCA2 non-founder mutations in patients of Ashkenazi Jewish ancestry.

An estimated 1:40 individuals of Ashkenazi Jewish (AJ) ancestry carry one of three common founder mutations in BRCA1 or BRCA2, resulting in the inherited cancer condition, Hereditary Breast and Ovarian Cancer (HBOC) syndrome. Targeted testing for these three mutations (BRCA1 187delAG, BRCA1 5385insC, and BRCA2 6174delT) is therefore recommended for all AJ breast and ovarian cancer patients, regardless of age of diagnosis or family history. Comprehensive analysis of both genes is recommended for a subset of AJ patients in whom founder mutations are not identified, but estimates of the yield from comprehensive analysis in this population vary widely. We sought to determine the proportion of non-founder mutations as a percentage of all mutations in BRCA1 and BRCA2 among AJ patients to inform decisions about HBOC testing strategies in this population. We analyzed the genetic testing results for 37,952 AJ patients for whom clinical testing of BRCA1 and BRCA2 was performed at Myriad Genetic Laboratories from January 2006 through August 2013. Analysis was limited to AJ-only patients for whom the initial test order was either (1) comprehensive testing, or (2) founder mutation testing with instructions to automatically "reflex" to comprehensive analysis if negative. Cases were excluded if a separate follow-up order was placed to reflex to comprehensive analysis only after the founder mutation testing was reported out as negative. Among all BRCA1 and BRCA2 mutations detected in these groups, the percentage of non-founder mutations was 13 % (104/802) and 7.2 % (198/2,769). One-hundred and eighty-nine unique non-founder mutations were detected, 76 in BRCA1 and 113 in BRCA2. Non-founder mutations make up between 7.2 and 13.0 % of all BRCA1 and BRCA2 mutations in Ashkenazi Jews. A wide range of mutations are present, most of which are also seen in non-AJ individuals.