Challenges and approaches to calibrating patient phenotype as evidence for cancer gene variant classification under ACMG/AMP guidelines.

Since first publication of the American College of Medical Genetics and Genomics/Association for Medical Pathology (ACMG/AMP) variant classification guidelines, additional recommendations for application of certain criteria have been released (https://clinicalgenome.org/docs/), to improve their application in the diagnostic setting. However, none have addressed use of the PS4 and PP4 criteria, capturing patient presentation as evidence towards pathogenicity. Application of PS4 can be done through traditional case-control studies, or "proband counting" within or across clinical testing cohorts. Review of the existing PS4 and PP4 specifications for Hereditary Cancer Gene Variant Curation Expert Panels revealed substantial differences in the approach to defining specifications. Using BRCA1, BRCA2 and TP53 as exemplar genes, we calibrated different methods proposed for applying the "PS4 proband counting" criterion. For each approach, we considered limitations, non-independence with other ACMG/AMP criteria, broader applicability, and variability in results for different datasets. Our findings highlight inherent overlap of proband-counting methods with ACMG/AMP frequency codes, and the importance of calibration to derive dataset-specific code weights that can account for potential between-dataset differences in ascertainment and other factors. Our work emphasizes the advantages and generalizability of logistic regression analysis over simple proband-counting approaches to empirically determine the relative predictive capacity and weight of various personal clinical features in the context of multigene panel testing, for improved variant interpretation. We also provide a general protocol, including instructions for data formatting and a web-server for analysis of personal history parameters, to facilitate dataset-specific calibration analyses required to use such data for germline variant classification.

Pretest Video Education Versus Genetic Counseling for Patients With Prostate Cancer: ProGen, A Multisite Randomized Controlled Trial.

PURPOSE: Germline genetic testing (GT) is recommended for men with prostate cancer (PC), but testing through traditional models is limited. The ProGen study examined a novel model aimed at providing access to GT while promoting education and informed consent. METHODS: Men with potentially lethal PC (metastatic, localized with a Gleason score of ≥8, persistent prostate-specific antigen after local therapy), diagnosis age ≤55 years, previous malignancy, and family history suggestive of a pathogenic variant (PV) and/or at oncologist's discretion were randomly assigned 3:1 to video education (VE) or in-person genetic counseling (GC). Participants had 67 genes analyzed (Ambry), with results disclosed via telephone by a genetic counselor. Outcomes included GT consent, GT completion, PV prevalence, and survey measures of satisfaction, psychological impact, genetics knowledge, and family communication. Two-sided Fisher's exact tests were used for between-arm comparisons. RESULTS: Over a 2-year period, 662 participants at three sites were randomly assigned and pretest VE (n = 498) or GC (n = 164) was completed by 604 participants (VE, 93.1%; GC, 88.8%), of whom 596 participants (VE, 98.9%; GC, 97.9%) consented to GT and 591 participants completed GT (VE, 99.3%; GC, 98.6%). These differences were not statistically significant although subtle differences in satisfaction and psychological impact were. Notably, 84 PVs were identified in 78 participants (13.2%), with BRCA1/2 PV comprising 32% of participants with a positive result (BRCA2 n = 21, BRCA1 n = 4). CONCLUSION: Both VE and traditional GC yielded high GT uptake without significant differences in outcome measures of completion, GT uptake, genetics knowledge, and family communication. The increased demand for GT with limited genetics resources supports consideration of pretest VE for patients with PC.

A genotype-first approach identifies high incidence of NF1 pathogenic variants with distinct disease associations.

Loss of function variants in the NF1 gene cause neurofibromatosis type 1 (NF1), a genetic disorder characterized by complete penetrance, prevalence of 1 in 3,000, characteristic physical exam findings, and a substantially increased risk for malignancy. However, our understanding of the disorder is entirely based on patients ascertained through phenotype-first approaches. Leveraging a genotype-first approach in two large patient cohorts, we demonstrate unexpectedly high prevalence (1 in 450-750) of NF1 pathogenic variants. Half were identified in individuals lacking clinical features of NF1, with many appearing to have post-zygotic mosaicism for the identified variant. Incidentally discovered variants were not associated with classic NF1 features but were associated with an increased incidence of malignancy compared to a control population. Our findings suggest that NF1 pathogenic variants are substantially more common than previously thought, often characterized by somatic mosaicism and reduced penetrance, and are important contributors to cancer risk in the general population.

Functional and Clinical Characterization of Variants of Uncertain Significance Identifies a Hotspot for Inactivating Missense Variants in RAD51C.

Pathogenic protein-truncating variants of RAD51C, which plays an integral role in promoting DNA damage repair, increase the risk of breast and ovarian cancer. A large number of RAD51C missense variants of uncertain significance (VUS) have been identified, but the effects of the majority of these variants on RAD51C function and cancer predisposition have not been established. Here, analysis of 173 missense variants by a homology-directed repair (HDR) assay in reconstituted RAD51C-/- cells identified 30 nonfunctional (deleterious) variants, including 18 in a hotspot within the ATP-binding region. The deleterious variants conferred sensitivity to cisplatin and olaparib and disrupted formation of RAD51C/XRCC3 and RAD51B/RAD51C/RAD51D/XRCC2 complexes. Computational analysis indicated the deleterious variant effects were consistent with structural effects on ATP-binding to RAD51C. A subset of the variants displayed similar effects on RAD51C activity in reconstituted human RAD51C-depleted cancer cells. Case-control association studies of deleterious variants in women with breast and ovarian cancer and noncancer controls showed associations with moderate breast cancer risk [OR, 3.92; 95% confidence interval (95% CI), 2.18-7.59] and high ovarian cancer risk (OR, 14.8; 95% CI, 7.71-30.36), similar to protein-truncating variants. This functional data supports the clinical classification of inactivating RAD51C missense variants as pathogenic or likely pathogenic, which may improve the clinical management of variant carriers. SIGNIFICANCE: Functional analysis of the impact of a large number of missense variants on RAD51C function provides insight into RAD51C activity and information for classification of the cancer relevance of RAD51C variants.

Clinician-Reported Impact of Germline Multigene Panel Testing on Cancer Risk Management Recommendations.

BACKGROUND: With increased adoption of multi-gene panel testing (MGPT) for hereditary cancer, management guidelines now include a wider range of predisposition genes. Yet little is known about whether MGPT results prompt changes to clinicians' risk management recommendations and whether those recommendations adhere to guidelines. METHODS: We assessed cancer risk management recommendations made by clinicians ordering MGPT for hereditary cancer at a diagnostic laboratory using an internet-based survey. We received paired pre- and posttest responses for 2172 patients (response rate = 14.3%). Unpaired posttest responses were received in 168 additional patients with positive results. All tests were 2-sided. RESULTS: Clinicians reported a change in risk management recommendations for 76.6% of patients who tested positive for a pathogenic or likely pathogenic variant, with changes to surveillance being most common (71.1%), followed by surgical (33.6%), chemoprevention (15.1%), and clinical trial (9.4%) recommendations. Clinicians recommended risk-reducing interventions more often for patients with pathogenic variants in high-risk than moderate-risk genes (P < .001), whereas surveillance recommendations were similar for high-risk and moderate-risk genes. Guideline adherence was high for surveillance (86.3%) and surgical (79.6%) recommendations. Changes to risk management recommendations occurred in 8.8% and 7.6% of patients with uncertain and negative results, respectively. CONCLUSIONS: Clinicians report frequent changes to cancer risk management recommendations based on positive results in both high-risk and moderate-risk genes. Reported introduction of interventions in patients with inconclusive and negative results is rare and adherence to practice guidelines is high in patients with positive results, suggesting a low probability of harm resulting from MGPT.

Suspected clonal hematopoiesis as a natural functional assay of TP53 germline variant pathogenicity.

PURPOSE: Some variants identified by multigene panel testing of DNA from blood present with low variant allele fraction (VAF), often a manifestation of clonal hematopoiesis. Research has shown that the proportion of variants with low VAF is especially high in TP53, the Li-Fraumeni syndrome gene. Based on the hypothesis that variants with low VAF are positively selected as drivers of clonal hematopoiesis, we investigated the use of VAF as a predictor of TP53 germline variant pathogenicity. METHODS: We used data from 260,681 TP53 variants identified at 2 laboratories to compare the distribution of pathogenic and benign variants at different VAF intervals. RESULTS: Likelihood ratios toward pathogenicity associated with a VAF < 26% equated to the American College of Medical Genetics/Association of Molecular Pathology strong strength level and were applicable for 1 in 5 variants of unknown significance. CONCLUSION: In conclusion, detection of variants with low VAF in blood can be considered an in vivo functional assay to aid assessment of TP53 variant pathogenicity.

Germline Pathogenic Variants in Cancer Predisposition Genes Among Women With Invasive Lobular Carcinoma of the Breast.

PURPOSE: To determine the contribution of germline pathogenic variants (PVs) in hereditary cancer testing panel genes to invasive lobular carcinoma (ILC) of the breast. MATERIALS AND METHODS: The study included 2,999 women with ILC from a population-based cohort and 3,796 women with ILC undergoing clinical multigene panel testing (clinical cohort). Frequencies of germline PVs in breast cancer predisposition genes (ATM, BARD1, BRCA1, BRCA2, BRIP1, CDH1, CHEK2, PALB2, PTEN, RAD51C, RAD51D, and TP53) were compared between women with ILC and unaffected female controls and between women with ILC and infiltrating ductal carcinoma (IDC). RESULTS: The frequency of PVs in breast cancer predisposition genes among women with ILC was 6.5% in the clinical cohort and 5.2% in the population-based cohort. In case-control analysis, CDH1 and BRCA2 PVs were associated with high risks of ILC (odds ratio [OR] > 4) and CHEK2, ATM, and PALB2 PVs were associated with moderate (OR = 2-4) risks. BRCA1 PVs and CHEK2 p.Ile157Thr were not associated with clinically relevant risks (OR < 2) of ILC. Compared with IDC, CDH1 PVs were > 10-fold enriched, whereas PVs in BRCA1 were substantially reduced in ILC. CONCLUSION: The study establishes that PVs in ATM, BRCA2, CDH1, CHEK2, and PALB2 are associated with an increased risk of ILC, whereas BRCA1 PVs are not. The similar overall PV frequencies for ILC and IDC suggest that cancer histology should not influence the decision to proceed with genetic testing. Similar to IDC, multigene panel testing may be appropriate for women with ILC, but CDH1 should be specifically discussed because of low prevalence and gastric cancer risk.

Case-case analysis addressing ascertainment bias for multigene panel testing implicates BRCA1 and PALB2 in endometrial cancer.

Hereditary endometrial cancer (EC) is most commonly attributed to pathogenic variants in mismatch repair genes. Evidence supports the existence of additional genetic risk factors in the context of multiple cancer diagnoses and/or family history of EC. EC patients (n = 5292) referred for diagnostic multigene cancer panel testing were annotated for presence of a pathogenic gene variant; personal history of prior, concurrent, or subsequent cancer of another type; reported family history of Lynch syndrome or EC. The Pearson χ2 test was used to assess differences in gene variant prevalence between case sub-groups defined by personal and/or family history of cancer/s, using cases with no family history of Lynch/EC as reference. Another cancer diagnosis was reported for 55% of EC cases. EC cases with a prior and reported family history of Lynch cancer were enriched for variants in MLH1 (p = 3.5 × 10-7 ), MSH2 (p = 3.1 × 10-7 ), and PMS2 (p = .02). Consistent with expectations for a breast cancer gene also predisposing to EC, the variant frequency was increased in EC patients with prior BC and family history of EC for BRCA1 (p = 1.7 × 10-5 ) and PALB2 (p = .0002). Strategic case-case analyses to address cohort ascertainment bias have provided a rationale to direct future studies of candidate hereditary EC genes.

An updated quantitative model to classify missense variants in the TP53 gene: A novel multifactorial strategy.

Multigene panel testing has led to an increase in the number of variants of uncertain significance identified in the TP53 gene, associated with Li-Fraumeni syndrome. We previously developed a quantitative model for predicting the pathogenicity of P53 missense variants based on the combination of calibrated bioinformatic information and somatic to germline ratio. Here, we extended this quantitative model for the classification of P53 predicted missense variants by adding new pieces of evidence (personal and family history parameters, loss-of-function results, population allele frequency, healthy individual status by age 60, and breast tumor pathology). We also annotated which missense variants might have an effect on splicing based on bioinformatic predictions. This updated model plus annotation led to the classification of 805 variants into a clinically relevant class, which correlated well with existing ClinVar classifications, and resolved a large number of conflicting and uncertain classifications. We propose this model as a reliable approach to TP53 germline variant classification and emphasize its use in contributing to optimize TP53-specific ACMG/AMP guidelines.

Risk of Late-Onset Breast Cancer in Genetically Predisposed Women.

PURPOSE: The prevalence of germline pathogenic variants (PVs) in established breast cancer predisposition genes in women in the general population over age 65 years is not well-defined. However, testing guidelines suggest that women diagnosed with breast cancer over age 65 years might have < 2.5% likelihood of a PV in a high-penetrance gene. This study aimed to establish the frequency of PVs and remaining risks of breast cancer for each gene in women over age 65 years. METHODS: A total of 26,707 women over age 65 years from population-based studies (51.5% with breast cancer and 48.5% unaffected) were tested for PVs in germline predisposition gene. Frequencies of PVs and associations between PVs in each gene and breast cancer were assessed, and remaining lifetime breast cancer risks were estimated for non-Hispanic White women with PVs. RESULTS: The frequency of PVs in predisposition genes was 3.18% for women with breast cancer and 1.48% for unaffected women over age 65 years. PVs in BRCA1, BRCA2, and PALB2 were found in 3.42% of women diagnosed with estrogen receptor (ER)-negative, 1.0% with ER-positive, and 3.01% with triple-negative breast cancer. Frequencies of PVs were lower among women with no first-degree relatives with breast cancer. PVs in CHEK2, PALB2, BRCA2, and BRCA1 were associated with increased risks (odds ratio = 2.9-4.0) of breast cancer. Remaining lifetime risks of breast cancer were ≥ 15% for those with PVs in BRCA1, BRCA2, and PALB2. CONCLUSION: This study suggests that all women diagnosed with triple-negative breast cancer or ER-negative breast cancer should receive genetic testing and that women over age 65 years with BRCA1 and BRCA2 PVs and perhaps with PALB2 and CHEK2 PVs should be considered for magnetic resonance imaging screening.

Association of Family Cancer History With Pathogenic Variants in Specific Breast Cancer Susceptibility Genes.

PURPOSE: Family cancer history is an important component of genetic testing guidelines that estimate which patients with breast cancer are most likely to carry a germline pathogenic variant (PV). However, we do not know whether more extensive family history is differentially associated with PVs in specific genes. METHODS: All women diagnosed with breast cancer in 2013-2017 and reported to statewide SEER registries of Georgia and California were linked to clinical genetic testing results and family history from two laboratories. Family history was defined as strong (suggestive of PVs in high-penetrance genes such as BRCA1/2 or TP53, including male breast, ovarian, pancreatic, sarcoma, or multiple female breast cancers), moderate (any other cancer history), or none. Among established breast cancer susceptibility genes (ATM, BARD1, BRCA1, BRCA2, CDH1, CHEK2, NF1, PALB2, PTEN, RAD51C, RAD51D, and TP53), we evaluated PV prevalence according to family history extent and breast cancer subtype. We used a multivariable model to test for interaction between affected gene and family history extent for ATM, BRCA1/2, CHEK2, and PALB2. RESULTS: A total of 34,865 women linked to genetic results. Higher PV prevalence with increasing family history extent (P < .001) was observed only with BRCA1 (3.04% with none, 3.22% with moderate, and 4.06% with strong history) and in triple-negative breast cancer with PALB2 (0.75% with none, 2.23% with moderate, and 2.63% with strong history). In a multivariable model adjusted for age and subtype, there was no interaction between family history extent and PV prevalence for any gene except PALB2 (P = .037). CONCLUSION: Extent of family cancer history is not differentially associated with PVs across established breast cancer susceptibility genes and cannot be used to personalize genes selected for testing.

Racial and Ethnic Differences in Multigene Hereditary Cancer Panel Test Results for Women With Breast Cancer.

To evaluate the racial and ethnic differences in prevalence of germline pathogenic variants (PVs) and the effect of race and ethnicity on breast cancer (BC) risk among carriers, results of multigene testing of 77 900 women with BC (non-Hispanic White [NHW] = 57 003; Ashkenazi-Jewish = 4798; Black = 6722; Hispanic = 5194; and Asian = 4183) were analyzed, and the frequency of PVs in each gene were compared between BC patients (cases) and race- and ethnicity-matched gnomAD reference controls. Compared with NHWs, BRCA1 PVs were enriched in Ashkenazi-Jews and Hispanics, whereas CHEK2 PVs were statistically significantly lower in Blacks, Hispanics, and Asians (all 2-sided P < .05). In case-control studies, BARD1 PVs were associated with high risks (odds ratio > 4.00) of BC in Blacks, Hispanics, and Asians; ATM PVs were associated with increased risk of BC among all races and ethnicities except Asians, whereas CHEK2 and BRIP1 PVs were associated with increased risk of BC among NHWs and Hispanics only. These findings suggest a need for personalized management of BC risk in PV carriers based on race and ethnicity.

Genotype-phenotype correlations among TP53 carriers: Literature review and analysis of probands undergoing multi-gene panel testing and single-gene testing.

Pathogenic germline variants in the TP53 gene predispose to a wide range of cancers, known collectively as Li-Fraumeni syndrome (LFS). There has been much research aimed to identify genotype-phenotype correlations, that is, differences between variant location and/or effect and cancer spectrum. These correlations, should they exist, have potential to impact clinical management of carriers. Review of previously published studies showed a variety of study designs and inconsistency in reported findings. Here, we used pooled data from 427 TP53 carriers who had undergone multigene panel testing and 154 TP53 carriers identified by single-gene testing to investigate correlations between TP53 genotype (truncating variants, hotspot variants, other missense variants with dominant-negative effect, missense variants without dominant-negative effect) and a number of LFS-selected malignancies. Our results suggest that carriers of truncating and hotspot variants might be more likely to present with LFS cancers and have shorter time to first cancer diagnosis compared to carriers of other variant types. However, the differences observed were minor, and we conclude that there is currently insufficient evidence to consider location and/or molecular effect of pathogenic variants to assist with clinical management of TP53 carriers. Larger studies are necessary to confirm the correlations suggested by our analysis.

A Rare TP53 Mutation Predominant in Ashkenazi Jews Confers Risk of Multiple Cancers.

Germline mutations in TP53 cause a rare high penetrance cancer syndrome, Li-Fraumeni syndrome (LFS). Here, we identified a rare TP53 tetramerization domain missense mutation, c.1000G>C;p.G334R, in a family with multiple late-onset LFS-spectrum cancers. Twenty additional c.1000G>C probands and one c.1000G>A proband were identified, and available tumors showed biallelic somatic inactivation of TP53. The majority of families were of Ashkenazi Jewish descent, and the TP53 c.1000G>C allele was found on a commonly inherited chromosome 17p13.1 haplotype. Transient transfection of the p.G334R allele conferred a mild defect in colony suppression assays. Lymphoblastoid cell lines from the index family in comparison with TP53 normal lines showed that although classical p53 target gene activation was maintained, a subset of p53 target genes (including PCLO, PLTP, PLXNB3, and LCN15) showed defective transactivation when treated with Nutlin-3a. Structural analysis demonstrated thermal instability of the G334R-mutant tetramer, and the G334R-mutant protein showed increased preponderance of mutant conformation. Clinical case review in comparison with classic LFS cohorts demonstrated similar rates of pediatric adrenocortical tumors and other LFS component cancers, but the latter at significantly later ages of onset. Our data show that TP53 c.1000G>C;p.G334R is found predominantly in Ashkenazi Jewish individuals, causes a mild defect in p53 function, and leads to low penetrance LFS. SIGNIFICANCE: TP53 c.1000C>G;p.G334R is a pathogenic, Ashkenazi Jewish-predominant mutation associated with a familial multiple cancer syndrome in which carriers should undergo screening and preventive measures to reduce cancer risk.

Suggested application of HER2+ breast tumor phenotype for germline TP53 variant classification within ACMG/AMP guidelines.

Early onset breast cancer is the most common malignancy in women with Li-Fraumeni syndrome, caused by germline TP53 pathogenic variants. It has repeatedly been suggested that breast tumors from TP53 carriers are more likely to be HER2+ than those of noncarriers, but this information has not been incorporated into variant interpretation models for TP53. Breast tumor pathology is already being used quantitatively for assessing pathogenicity of germline variants in other genes, and it has been suggested that this type of evidence can be incorporated into current American College of Medical Genetics and Genomics/Association for Molecular Pathology (ACMG/AMP) guidelines for germline variant classification. Here, by reviewing published data and using internal datasets separated by different age groups, we investigated if breast tumor HER2+ status has utility as a predictor of TP53 germline variant pathogenicity, considering age at diagnosis. Overall, our results showed that the identification of HER2+ breast tumors diagnosed before the age of 40 can be conservatively incorporated into the current TP53-specific ACMG/AMP PP4 criterion, following a point system detailed in this manuscript. Further larger studies will be needed to reassess the value of HER2+ breast tumors diagnosed at a later age.

Mutation prevalence tables for hereditary cancer derived from multigene panel testing.

Multigene panel testing for cancer predisposition mutations is becoming routine in clinical care. However, the gene content of panels offered by testing laboratories vary significantly, and data on mutation detection rates by gene and by the panel is limited, causing confusion among clinicians on which test to order. Using results from 147,994 multigene panel tests conducted at Ambry Genetics, we built an interactive prevalence tool to explore how differences in ethnicity, age of onset, and personal and family history of different cancers affect the prevalence of pathogenic mutations in 31 cancer predisposition genes, across various clinically available hereditary cancer gene panels. Over 13,000 mutation carriers were identified in this high-risk population. Most were non-Hispanic white (74%, n = 109,537), but also Black (n = 10,875), Ashkenazi Jewish (n = 10,464), Hispanic (n = 10,028), and Asian (n = 7,090). The most prevalent cancer types were breast (50%), ovarian (6.6%), and colorectal (4.7%), which is expected based on genetic testing guidelines and clinician referral for testing. The Hereditary Cancer Multi-Gene Panel Prevalence Tool presented here can be used to provide insight into the prevalence of mutations on a per-gene and per-multigene panel basis, while conditioning on multiple custom phenotypic variables to include race and cancer type.

Diagnosing hereditary cancer predisposition in men with prostate cancer.

PURPOSE: We describe the pathogenic variant spectrum and identify predictors of positive results among men referred for clinical genetic testing for prostate cancer. METHODS: One thousand eight hundred twelve men with prostate cancer underwent clinical multigene panel testing between April 2012 and September 2017. Stepwise logistic regression determined the most reliable predictors of positive results among clinical variables reported on test requisition forms. RESULTS: A yield of 9.4-12.1% was observed among men with no prior genetic testing. In this group, the positive rate of BRCA1 and BRCA2 was 4.6%; the positive rate for the mismatch repair genes was 2.8%. Increasing Gleason score (odds ratio [OR] 1.19; 95% confidence interval [CI] 0.97-1.45); personal history of breast or pancreatic cancer (OR 3.62; 95% CI 1.37-9.46); family history of breast, ovarian, or pancreatic cancer (OR 2.32 95% CI 1.48-3.65); and family history of Lynch syndrome-associated cancers (OR 1.97; 95% CI 1.23-3.15) were predictors of positive results. CONCLUSION: These results support multigene panel testing as the primary genetic testing approach for hereditary prostate cancer and are supportive of recommendations for consideration of germline testing in men with prostate cancer. Expanding the criteria for genetic testing should be considered as many pathogenic variants are actionable for treatment of advanced prostate cancer.

Ovarian and Breast Cancer Risks Associated With Pathogenic Variants in RAD51C and RAD51D.

BACKGROUND: The purpose of this study was to estimate precise age-specific tubo-ovarian carcinoma (TOC) and breast cancer (BC) risks for carriers of pathogenic variants in RAD51C and RAD51D. METHODS: We analyzed data from 6178 families, 125 with pathogenic variants in RAD51C, and 6690 families, 60 with pathogenic variants in RAD51D. TOC and BC relative and cumulative risks were estimated using complex segregation analysis to model the cancer inheritance patterns in families while adjusting for the mode of ascertainment of each family. All statistical tests were two-sided. RESULTS: Pathogenic variants in both RAD51C and RAD51D were associated with TOC (RAD51C: relative risk [RR] = 7.55, 95% confidence interval [CI] = 5.60 to 10.19; P = 5 × 10-40; RAD51D: RR = 7.60, 95% CI = 5.61 to 10.30; P = 5 × 10-39) and BC (RAD51C: RR = 1.99, 95% CI = 1.39 to 2.85; P = 1.55 × 10-4; RAD51D: RR = 1.83, 95% CI = 1.24 to 2.72; P = .002). For both RAD51C and RAD51D, there was a suggestion that the TOC relative risks increased with age until around age 60 years and decreased thereafter. The estimated cumulative risks of developing TOC to age 80 years were 11% (95% CI = 6% to 21%) for RAD51C and 13% (95% CI = 7% to 23%) for RAD51D pathogenic variant carriers. The estimated cumulative risks of developing BC to 80 years were 21% (95% CI = 15% to 29%) for RAD51C and 20% (95% CI = 14% to 28%) for RAD51D pathogenic variant carriers. Both TOC and BC risks for RAD51C and RAD51D pathogenic variant carriers varied by cancer family history and could be as high as 32-36% for TOC, for carriers with two first-degree relatives diagnosed with TOC, or 44-46% for BC, for carriers with two first-degree relatives diagnosed with BC. CONCLUSIONS: These estimates will facilitate the genetic counseling of RAD51C and RAD51D pathogenic variant carriers and justify the incorporation of RAD51C and RAD51D into cancer risk prediction models.

The Contribution of Germline Predisposition Gene Mutations to Clinical Subtypes of Invasive Breast Cancer From a Clinical Genetic Testing Cohort.

BACKGROUND: The germline cancer predisposition genes associated with increased risk of each clinical subtype of breast cancer, defined by estrogen receptor (ER), progesterone receptor (PR), and HER2, are not well defined. METHODS: A total of 54 555 invasive breast cancer patients with 56 480 breast tumors were subjected to clinical hereditary cancer multigene panel testing. Heterogeneity for predisposition genes across clinical breast cancer subtypes was assessed by comparing mutation frequencies by gene among tumor subtypes and by association studies between each tumor subtype and reference controls. RESULTS: Mutations in 15 cancer predisposition genes were detected in 8.6% of patients with ER+/HER2-; 8.9% with ER+/HER2+; 7.7% with ER-/HER2+; and 14.4% of ER-/PR-/HER2- tumors. BRCA1, BRCA2, BARD1, and PALB2 mutations were enriched in ER- and HER2- tumors; RAD51C and RAD51D mutations were enriched in ER- tumors only; TP53 mutations were enriched in HER2+ tumors, and ATM and CHEK2 mutations were enriched in both ER+ and/or HER2+ tumors. All genes were associated with moderate (odds ratio > 2.00) or strong (odds ratio > 5.00) risks of at least one subtype of breast cancer in case-control analyses. Mutations in ATM, BARD1, BRCA1, BRCA2, CHEK2, PALB2, RAD51C, RAD51D, and TP53 had predicted lifetime absolute risks of at least 20.0% for breast cancer. CONCLUSIONS: Germline mutations in hereditary cancer panel genes confer subtype-specific risks of breast cancer. Combined tumor subtype, age at breast cancer diagnosis, and family history of breast and/or ovarian cancer information provides refined categorical estimates of mutation prevalence for women considering genetic testing.

Differences in patient ascertainment affect the use of gene-specified ACMG/AMP phenotype-related variant classification criteria: Evidence for TP53.

The American College of Medical Genetics and Genomics/Association for Molecular Pathology (ACMG/AMP) guidelines for variant classification are widely used for clinical interpretation of gene test results. These guidelines may be specified to genes/syndromes of interest to improve their utility in the clinical setting. As part of these specifications, phenotype-related criteria can be detailed and weighted depending on the personal history of disease for a given variant carrier. We investigated how ascertainment can affect the significance and/or weight of patient phenotype as a predictor of germline-variant pathogenicity, using the Li-Fraumeni Syndrome gene TP53 as an example. Likelihood ratios in favor of variant pathogenicity were determined for a report of the personal history of several TP53-related cancers, using data from 2,656 probands undergoing single-gene testing (SGT) and 15,483 undergoing multi-gene panel testing (MGPT). Overall, TP53-associated cancers were more predictive of pathogenicity, and demonstrated greater evidence weight, in the MGPT versus SGT dataset. This observation is almost certainly explained by differences in proband ascertainment for the two streams of testing, and these findings have implications for germline-variant classification using ACMG/AMP guidelines.