Couch, Fergus J.

Abstract Breast cancer is the most common cancer among women in the United States, with up to 30% of those diagnosed displaying a family history of breast cancer. To date, 18% of the familial risk of breast cancer can be explained by SNPs. This review summarizes the discovery of risk-associated SNPs using candidate gene and genome-wide association studies (GWAS), including discovery and replication in large collaborative efforts such as The Collaborative Oncologic Gene-environment Study and OncoArray. We discuss the evolution of GWAS studies, efforts to discover additional SNPs, and methods for identifying causal variants. We summarize findings associated with overall breast cancer, pathologic subtypes, and mutation carriers (BRCA1, BRCA2, and CHEK2). In addition, we summarize the development of polygenic risk scores (PRS) using the risk-associated SNPs and show how PRS can contribute to estimation of individual risks for developing breast cancer. Cancer Epidemiol Biomarkers Prev; 27(4); 380–94. ©2018 AACR. See all articles in this CEBP Focus section, “Genome-Wide Association Studies in Cancer.”

Background Missense (amino-acid changing) variants found in cancer predisposition genes often create difficulties when clinically interpreting genetic testing results. Although bioinformatics has developed approaches to predicting the impact of these variants, many of these approaches have not been readily applicable in the clinical setting. Bioinformatics approaches for predicting the impact of these variants have not yet found their footing in clinical practice because 1) interpreting the medical relevance of predictive scores is difficult; 2) the relationship between bioinformatics “predictors” (sequence conservation, protein structure) and cancer susceptibility is not understood. Methodology/Principal Findings We present a computational method that produces a probabilistic likelihood ratio predictive of whether a missense variant impairs protein function. We apply the method to a tumor suppressor gene, BRCA2, whose loss of function is important to cancer susceptibility. Protein likelihood ratios are computed for 229 unclassified variants found in individuals from high-risk breast/ovarian cancer families. We map the variants onto a protein structure model, and suggest that a cluster of predicted deleterious variants in the BRCA2 OB1 domain may destabilize BRCA2 and a protein binding partner, the small acidic protein DSS1. We compare our predictions with variant “re-classifications” provided by Myriad Genetics, a biotechnology company that holds the patent on BRCA2 genetic testing in the U.S., and with classifications made by an established medical genetics model [ 1 ]. Our approach uses bioinformatics data that is independent of these genetics-based classifications and yet shows significant agreement with them. Preliminary results indicate that our method is less likely to make false positive errors than other bioinformatics methods, which were designed to predict the impact of missense mutations in general. Conclusions/Significance Missense mutations are the most common disease-producing genetic variants. We present a fast, scalable bioinformatics method that integrates information about protein sequence, conservation, and structure in a likelihood ratio that can be integrated with medical genetics likelihood ratios. The protein likelihood ratio, together with medical genetics likelihood ratios, can be used by clinicians and counselors to communicate the relevance of a VUS to the individual who has that VUS. The approach described here is generalizable to regions of any tumor suppressor gene that have been structurally determined by X-ray crystallography or for which a protein homology model can be built.

Abstract Purpose: The relevance of inherited pathogenic variants in cancer predisposition genes to pancreatic cancer (PC) is not well understood. Several small studies have identified pathogenic variants in 4% to 14% of unselected PC patients using multigene panels of predisposition genes, but only BRCA2, ATM, and PALB2 have been clearly implicated in this disease. We aimed to assess the clinical and molecular characteristics of PC patients referred for hereditary cancer genetic testing, and to estimate the risk of PC associated with pathogenic variants in panel-based cancer predisposition genes. Methods: PC patients (n=1,819) were ascertained from a large cohort of over 140,000 patients undergoing multigene panel testing (MGPT) of predisposition genes between March 2012 and June 2016 at a single diagnostic laboratory. Clinical histories and molecular results were reviewed and summarized. Gene-level variant frequencies among PC cases were compared to those from the Exome Aggregation Consortium (ExAC) to calculate gene-specific pancreatic cancer risk ratios. Results: PC patients were predominantly Caucasian (76.5%) and female (58.9%), with a median age at diagnosis of 61 years (51.7). Of these, 33.5% reported additional cancer primaries, and 44.8% reported a family history of PC. Overall, 15.4% of PC patients were found to have at least one pathogenic/likely pathogenic variant in panel-based predisposition genes. Genes with the highest frequencies of pathogenic/likely pathogenic variants included BRCA2 (3.9%), ATM (3.6%), CHEK2 (excluding p.Ile157Thr) (2.0%), PALB2 (1.5%), VHL (1.4%), CDKN2A (1.2%), BRCA1 (0.8%), and MSH6 (0.8%). 21.8% of BRCA1 and BRCA2 carriers did not meet BRCA1/2 testing criteria and 61.5% of MSH6 carriers did not meet Lynch syndrome testing criteria. No CDKN2A families met diagnostic criteria for familial atypical multiple mole melanoma syndrome, and 44% did not report any personal or family history of melanoma. To estimate associations between pathogenic variants and pancreatic cancer, Caucasian PC cases were compared to non-Finnish European, non-TCGA ExAC reference controls. Pathogenic variants in ATM, BRCA2, CDKN2A, MSH6, and PALB2 were significantly associated with high PC risks. Pathogenic variants in BRCA1 were associated with a moderate risk of PC (RR=2.7). Conclusions: These findings shed light on the spectrum of mutations that can be expected for PC patients referred for cancer predisposition testing. The results confirm the associations of CDKN2A and BRCA2 variants with PC, and expand on the phenotypic spectrum associated with these variants. Furthermore, these results suggest that ATM, PALB2, and MSH6 may be high-risk PC genes, warranting further investigation in case-control and family-based studies. Citation Format: Holly LaDuca, Chunling Hu, Hermela Shimelis, Eric Polley, Jenna Lilyquist, Mary Helen Black, Brigette Tippin Davis, David E. Goldgar, Jill Dolinsky, Fergus J. Couch. What have we learned from pancreatic cancer patients undergoing multigene panel testing [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 4286. doi:10.1158/1538-7445.AM2017-4286

Abstract A better understanding of gene-specific risks for development of breast cancer will lead to improved screening, prevention, and therapeutic strategies for individuals identified to carry germline mutations. We performed targeted massively-parallel sequencing to identify mutations and large genomic rearrangements in 26 known or proposed breast cancer susceptibility genes in 2134 BRCA-negative women with familial breast cancer (FBC). A case-control analysis was performed comparing the frequency of internally classified mutations identified in FBC cases to that in non-Finnish European controls from the Exome Aggregation Consortium (ExAC) excluding samples from The Cancer Genome Atlas. Including large genomic rearrangements, mutations were identified in 8.2% of FBC cases compared to 6.2% of ExAC controls, including mutations in high-penetrance genes (0.6% in cases vs. 0.1% in controls), moderate-penetrance genes (3.7% vs 1.7%), and seven cases with two mutations (0.3%). The remainder of FBC cases and ExAC controls had mutations in proposed breast cancer genes (1.6% of cases vs 2.4% of controls), Lynch syndrome genes (0.5% vs. 0.5%) or were heterozygous MUTYH carriers (1.5% vs. 1.5%). Case-control analysis demonstrated significant associations with FBC for ATM, PALB2, and TP53 mutations (OR>3.0, p<10-4), BARD1 mutations (OR=3.2, p=0.012), and CHEK2 truncating mutations (OR=1.6, p=0.041). Our results therefore demonstrate that only approximately 4% of BRCA1/2 negative FBC patients have mutations in genes definitively associated with breast cancer at this time. Large case-control studies are needed to fully evaluate the breast cancer risks associated with moderate penetrance and proposed breast cancer susceptibility genes. Citation Format: Kara N. Maxwell, Thomas Paul Slavin, Jenna M. Lilyquist, Joseph Vijai, Susan L. Neuhausen, Steven N. Hart, Vignesh Ravichandran, Tinu Thomas, Ann Maria, Kasmintan A. Schrader, Raymond Moore, Chunling Hu, Brad Wubbenhorst, Brandon M. Wenz, Kurt D'Andrea, Susan M. Domchek, Mark E. Robson, Paulo Peterlongo, Paolo Radice, James M. Ford, Judy E. Garber, Csilla Szabo, Kenneth Offit, Katherine L. Nathanson, Fergus J. Couch, Jeffrey N. Weitzel. Risks of familial breast cancer associated with known and proposed breast cancer susceptibility genes [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 4265. doi:10.1158/1538-7445.AM2017-4265

Abstract While the population-based risk for breast cancer in males remains relatively low (1:1000), inherited predisposition can significantly raise this to as high as 10%, in men who carry a mutation in the BRCA2 gene. Lifetime breast cancer risks of 1-2% have also been reported in men who carry a mutation in BRCA1. These risks, as well as elevated risks of prostate, pancreatic, and melanoma cancers, are important to discuss in families diagnosed with Hereditary Breast and Ovarian Cancer Syndrome, which is often viewed as being clinically relevant only to the women in an affected family. However, beyond the BRCA1/2 genes, limited data is available on hereditary predisposition to male breast cancer. We analyzed clinical histories and molecular results from multi-gene panel testing for hereditary cancer predisposition in a cohort of 715 men affected by breast cancer. A total of 708 male breast cancer patients were eligible for inclusion in the final analysis. Molecular testing included analysis of 5 to 59 genes for DNA coding sequence and copy number variants by next-generation sequencing and microarray. Genetic variants identified were classified according to a 5-tier system using previously validated algorithms1,2. Only those variants classified as pathogenic or likely pathogenic were included in the analyses as positive for a mutation. Overall, a mutation was detected in 18% of these men. Four subjects carried a mutation in two different breast cancer predisposition genes. BRCA2 and CHEK2 were the most frequently mutated genes. The risk of breast cancer was significantly elevated compared to public controls in individuals with a mutation in BRCA2 (odds ratio (OR)=13.9; p=1.92x10-16), CHEK2 (OR=3.8; p=6.24x10-24) and PALB2 (OR=6.6, p=0.01). Average age was similar amongst men with (63.5±2.7 years) and without (62.3±1.2 years) mutations, as were clinical and family histories of additional cancers. The high overall diagnostic yield suggests the utility of testing all male breast cancer patient regardless of age or family history by multigene panel testing, and provides data to support risk-based counseling and medical recommendations for screening and/or prevention in male mutation carriers. 1. 1. LaDuca et al. Genet Med. 2014 Nov;16(11):830-7 2. 2. Richards et al. Genet Med. 2015 May;17(5):405-24 Citation Format: Elizabeth C. Chao, Mary Pritzlaff, Summerour Pia, Rachel McFarland, Shuwei Li, Jill Dolinsky, David Goldgar, Hermela Shimelis, Fergus Couch, LaDuca Holly. Hereditary risks of male breast cancer in a multi-gene panel testing cohort [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 3406. doi:10.1158/1538-7445.AM2017-3406