Analysis of more than 400,000 women provides case-control evidence for BRCA1 and BRCA2 variant classification.

Clinical genetic testing identifies variants causal for hereditary cancer, information that is used for risk assessment and clinical management. Unfortunately, some variants identified are of uncertain clinical significance (VUS), complicating patient management. Case-control data is one evidence type used to classify VUS, and previous findings indicate that case-control likelihood ratios (LRs) outperform odds ratios for variant classification. As an initiative of the Evidence-based Network for the Interpretation of Germline Mutant Alleles (ENIGMA) Analytical Working Group we analyzed germline sequencing data of BRCA1 and BRCA2 from 96,691 female breast cancer cases and 303,925 unaffected controls from three studies: the BRIDGES study of the Breast Cancer Association Consortium, the Cancer Risk Estimates Related to Susceptibility consortium, and the UK Biobank. We observed 11,227 BRCA1 and BRCA2 variants, with 6,921 being coding, covering 23.4% of BRCA1 and BRCA2 VUS in ClinVar and 19.2% of ClinVar curated (likely) benign or pathogenic variants. Case-control LR evidence was highly consistent with ClinVar assertions for (likely) benign or pathogenic variants; exhibiting 99.1% sensitivity and 95.4% specificity for BRCA1 and 92.2% sensitivity and 86.6% specificity for BRCA2. This approach provides case-control evidence for 785 unclassified variants, that can serve as a valuable element for clinical classification.

Large-scale meta-genome-wide association study reveals common genetic factors linked to radiation-induced acute toxicities across cancer types.

BACKGROUND: This study was designed to identify common genetic susceptibility and shared genetic variants associated with acute radiation-induced toxicity across 4 cancer types (prostate, head and neck, breast, and lung). METHODS: A genome-wide association study meta-analysis was performed using 19 cohorts totaling 12 042 patients. Acute standardized total average toxicity (STATacute) was modelled using a generalized linear regression model for additive effect of genetic variants, adjusted for demographic and clinical covariates (rSTATacute). Linkage disequilibrium score regression estimated shared single-nucleotide variation (SNV-formerly SNP)-based heritability of rSTATacute in all patients and for each cancer type. RESULTS: Shared SNV-based heritability of STATacute among all cancer types was estimated at 10% (SE = 0.02) and was higher for prostate (17%, SE = 0.07), head and neck (27%, SE = 0.09), and breast (16%, SE = 0.09) cancers. We identified 130 suggestive associated SNVs with rSTATacute (5.0 × 10‒8 < P < 1.0 × 10‒5) across 25 genomic regions. rs142667902 showed the strongest association (effect allele A; effect size ‒0.17; P = 1.7 × 10‒7), which is located near DPPA4, encoding a protein involved in pluripotency in stem cells, which are essential for repair of radiation-induced tissue injury. Gene-set enrichment analysis identified 'RNA splicing via endonucleolytic cleavage and ligation' (P = 5.1 × 10‒6, P = .079 corrected) as the top gene set associated with rSTATacute among all patients. In silico gene expression analysis showed that the genes associated with rSTATacute were statistically significantly up-regulated in skin (not sun exposed P = .004 corrected; sun exposed P = .026 corrected). CONCLUSIONS: There is shared SNV-based heritability for acute radiation-induced toxicity across and within individual cancer sites. Future meta-genome-wide association studies among large radiation therapy patient cohorts are worthwhile to identify the common causal variants for acute radiotoxicity across cancer types.

Multi-gene panel testing and association analysis in Cypriot breast cancer cases and controls.

Introduction: It is estimated that around 5% of breast cancer cases carry pathogenic variants in established breast cancer susceptibility genes. However, the underlying prevalence and gene-specific population risk estimates in Cyprus are currently unknown. Methods: We performed sequencing on a population-based case-control study of 990 breast cancer cases and 1094 controls from Cyprus using the BRIDGES sequencing panel. Analyses were conducted separately for protein-truncating and rare missense variants. Results: Protein-truncating variants in established breast cancer susceptibility genes were detected in 3.54% of cases and 0.37% of controls. Protein-truncating variants in BRCA2 and ATM were associated with a high risk of breast cancer, whereas PTVs in BRCA1 and PALB2 were associated with a high risk of estrogen receptor (ER)-negative disease. Among participants with a family history of breast cancer, PTVs in ATM, BRCA2, BRCA1, PALB2 and RAD50 were associated with an increased risk of breast cancer. Furthermore, an additional 19.70% of cases and 17.18% of controls had at least one rare missense variant in established breast cancer susceptibility genes. For BRCA1 and PALB2, rare missense variants were associated with an increased risk of overall and triple-negative breast cancer, respectively. Rare missense variants in BRCA1, ATM, CHEK2 and PALB2 domains, were associated with increased risk of disease subtypes. Conclusion: This study provides population-based prevalence and gene-specific risk estimates for protein-truncating and rare missense variants. These results may have important clinical implications for women who undergo genetic testing and be pivotal for a substantial proportion of breast cancer patients in Cyprus.

The impact of coding germline variants on contralateral breast cancer risk and survival.

Evidence linking coding germline variants in breast cancer (BC)-susceptibility genes other than BRCA1, BRCA2, and CHEK2 with contralateral breast cancer (CBC) risk and breast cancer-specific survival (BCSS) is scarce. The aim of this study was to assess the association of protein-truncating variants (PTVs) and rare missense variants (MSVs) in nine known (ATM, BARD1, BRCA1, BRCA2, CHEK2, PALB2, RAD51C, RAD51D, and TP53) and 25 suspected BC-susceptibility genes with CBC risk and BCSS. Hazard ratios (HRs) and 95% confidence intervals (CIs) were estimated with Cox regression models. Analyses included 34,401 women of European ancestry diagnosed with BC, including 676 CBCs and 3,449 BC deaths; the median follow-up was 10.9 years. Subtype analyses were based on estrogen receptor (ER) status of the first BC. Combined PTVs and pathogenic/likely pathogenic MSVs in BRCA1, BRCA2, and TP53 and PTVs in CHEK2 and PALB2 were associated with increased CBC risk [HRs (95% CIs): 2.88 (1.70-4.87), 2.31 (1.39-3.85), 8.29 (2.53-27.21), 2.25 (1.55-3.27), and 2.67 (1.33-5.35), respectively]. The strongest evidence of association with BCSS was for PTVs and pathogenic/likely pathogenic MSVs in BRCA2 (ER-positive BC) and TP53 and PTVs in CHEK2 [HRs (95% CIs): 1.53 (1.13-2.07), 2.08 (0.95-4.57), and 1.39 (1.13-1.72), respectively, after adjusting for tumor characteristics and treatment]. HRs were essentially unchanged when censoring for CBC, suggesting that these associations are not completely explained by increased CBC risk, tumor characteristics, or treatment. There was limited evidence of associations of PTVs and/or rare MSVs with CBC risk or BCSS for the 25 suspected BC genes. The CBC findings are relevant to treatment decisions, follow-up, and screening after BC diagnosis.

Segregation analysis of 17,425 population-based breast cancer families: Evidence for genetic susceptibility and risk prediction.

Rare pathogenic variants in known breast cancer-susceptibility genes and known common susceptibility variants do not fully explain the familial aggregation of breast cancer. To investigate plausible genetic models for the residual familial aggregation, we studied 17,425 families ascertained through population-based probands, 86% of whom were screened for pathogenic variants in BRCA1, BRCA2, PALB2, CHEK2, ATM, and TP53 via gene-panel sequencing. We conducted complex segregation analyses and fitted genetic models in which breast cancer incidence depended on the effects of known susceptibility genes and other unidentified major genes and a normally distributed polygenic component. The proportion of familial variance explained by the six genes was 46% at age 20-29 years and decreased steadily with age thereafter. After allowing for these genes, the best fitting model for the residual familial variance included a recessive risk component with a combined genotype frequency of 1.7% (95% CI: 0.3%-5.4%) and a penetrance to age 80 years of 69% (95% CI: 38%-95%) for homozygotes, which may reflect the combined effects of multiple variants acting in a recessive manner, and a polygenic variance of 1.27 (95% CI: 0.94%-1.65), which did not vary with age. The proportion of the residual familial variance explained by the recessive risk component was 40% at age 20-29 years and decreased with age thereafter. The model predicted age-specific familial relative risks consistent with those observed by large epidemiological studies. The findings have implications for strategies to identify new breast cancer-susceptibility genes and improve disease-risk prediction, especially at a young age.

Meta-GWAS identifies the heritability of acute radiation-induced toxicities in head and neck cancer.

BACKGROUND AND PURPOSE: We aimed to the genetic components and susceptibility variants associated with acute radiation-induced toxicities (RITs) in patients with head and neck cancer (HNC). MATERIALS AND METHODS: We performed the largest meta-GWAS of seven European cohorts (n = 4,042). Patients were scored weekly during radiotherapy for acute RITs including dysphagia, mucositis, and xerostomia. We analyzed the effect of variants on the average burden (measured as area under curve, AUC) per each RIT, and standardized total average acute toxicity (STATacute) score using a multivariate linear regression. We tested suggestive variants (p < 1.0x10-5) in discovery set (three cohorts; n = 2,640) in a replication set (four cohorts; n = 1,402). We meta-analysed all cohorts to calculate RITs specific SNP-based heritability, and effect of polygenic risk scores (PRSs), and genetic correlations among RITS. RESULTS: From 393 suggestive SNPs identified in discovery set; 37 were nominally significant (preplication < 0.05) in replication set, but none reached genome-wide significance (pcombined < 5 × 10-8). In-silico functional analyses identified "3'-5'-exoribonuclease activity" (FDR = 1.6e-10) for dysphagia, "inositol phosphate-mediated signalling" for mucositis (FDR = 2.20e-09), and "drug catabolic process" for STATacute (FDR = 3.57e-12) as the most enriched pathways by the RIT specific suggestive genes. The SNP-based heritability (±standard error) was 29 ± 0.08 % for dysphagia, 9 ± 0.12 % (mucositis) and 27 ± 0.09 % (STATacute). Positive genetic correlation was rg = 0.65 (p = 0.048) between dysphagia and STATacute. PRSs explained limited variation of dysphagia (3 %), mucositis (2.5 %), and STATacute (0.4 %). CONCLUSION: In HNC patients, acute RITs are modestly heritable, sharing 10 % genetic susceptibility, when PRS explains < 3 % of their variance. We identified numerus suggestive SNPs, which remain to be replicated in larger studies.

Prospective validation of the BOADICEA multifactorial breast cancer risk prediction model in a large prospective cohort study.

BACKGROUND: The multifactorial Breast and Ovarian Analysis of Disease Incidence and Carrier Estimation Algorithm (BOADICEA) breast cancer risk prediction model has been recently extended to consider all established breast cancer risk factors. We assessed the clinical validity of the model in a large independent prospective cohort. METHODS: We validated BOADICEA (V.6) in the Swedish KARolinska Mammography Project for Risk Prediction of Breast Cancer (KARMA) cohort including 66 415 women of European ancestry (median age 54 years, IQR 45-63; 816 incident breast cancers) without previous cancer diagnosis. We calculated 5-year risks on the basis of questionnaire-based risk factors, pedigree-structured first-degree family history, mammographic density (BI-RADS), a validated breast cancer polygenic risk score (PRS) based on 313-SNPs, and pathogenic variant status in 8 breast cancer susceptibility genes: BRCA1, BRCA2, PALB2, CHEK2, ATM, RAD51C, RAD51D and BARD1. Calibration was assessed by comparing observed and expected risks in deciles of predicted risk and the calibration slope. The discriminatory ability was assessed using the area under the curve (AUC). RESULTS: Among the individual model components, the PRS contributed most to breast cancer risk stratification. BOADICEA was well calibrated in predicting the risks for low-risk and high-risk women when all, or subsets of risk factors are included in the risk prediction. Discrimination was maximised when all risk factors are considered (AUC=0.70, 95% CI: 0.66 to 0.73; expected-to-observed ratio=0.88, 95% CI: 0.75 to 1.04; calibration slope=0.97, 95% CI: 0.95 to 0.99). The full multifactorial model classified 3.6% women as high risk (5-year risk ≥3%) and 11.1% as very low risk (5-year risk <0.33%). CONCLUSION: The multifactorial BOADICEA model provides valid breast cancer risk predictions and a basis for personalised decision-making on disease prevention and screening.

No Association Between Polygenic Risk Scores for Cancer and Development of Radiation Therapy Toxicity.

PURPOSE: Our aim was to test whether updated polygenic risk scores (PRS) for susceptibility to cancer affect risk of radiation therapy toxicity. METHODS AND MATERIALS: Analyses included 9,717 patients with breast (n=3,078), prostate (n=5,748) or lung (n=891) cancer from Radiogenomics and REQUITE Consortia cohorts. Patients underwent potentially curative radiation therapy and were assessed prospectively for toxicity. Germline genotyping involved genome-wide single nucleotide polymorphism (SNP) arrays with nontyped SNPs imputed. PRS for each cancer were generated by summing literature-identified cancer susceptibility risk alleles: 352 breast, 136 prostate, and 24 lung. Weighted PRS were generated using log odds ratio (ORs) for cancer susceptibility. Standardized total average toxicity (STAT) scores at 2 and 5 years (breast, prostate) or 6 to 12 months (lung) quantified toxicity. Primary analysis tested late STAT, secondary analyses investigated acute STAT, and individual endpoints and SNPs using multivariable regression. RESULTS: Increasing PRS did not increase risk of late toxicity in patients with breast (OR, 1.000; 95% confidence interval [CI], 0.997-1.002), prostate (OR, 0.99; 95% CI, 0.98-1.00; weighted PRS OR, 0.93; 95% CI, 0.83-1.03), or lung (OR, 0.93; 95% CI, 0.87-1.00; weighted PRS OR, 0.68; 95% CI, 0.45-1.03) cancer. Similar results were seen for acute toxicity. Secondary analyses identified rs138944387 associated with breast pain (OR, 3.05; 95% CI, 1.86-5.01; P = 1.09 × 10-5) and rs17513613 with breast edema (OR, 0.94; 95% CI, 0.92-0.97; P = 1.08 × 10-5). CONCLUSIONS: Patients with increased polygenic predisposition to breast, prostate, or lung cancer can safely undergo radiation therapy with no anticipated excess toxicity risk. Some individual SNPs increase the likelihood of a specific toxicity endpoint, warranting validation in independent cohorts and functional studies to elucidate biologic mechanisms.

Breast cancer risks associated with missense variants in breast cancer susceptibility genes.

BACKGROUND: Protein truncating variants in ATM, BRCA1, BRCA2, CHEK2, and PALB2 are associated with increased breast cancer risk, but risks associated with missense variants in these genes are uncertain. METHODS: We analyzed data on 59,639 breast cancer cases and 53,165 controls from studies participating in the Breast Cancer Association Consortium BRIDGES project. We sampled training (80%) and validation (20%) sets to analyze rare missense variants in ATM (1146 training variants), BRCA1 (644), BRCA2 (1425), CHEK2 (325), and PALB2 (472). We evaluated breast cancer risks according to five in silico prediction-of-deleteriousness algorithms, functional protein domain, and frequency, using logistic regression models and also mixture models in which a subset of variants was assumed to be risk-associated. RESULTS: The most predictive in silico algorithms were Helix (BRCA1, BRCA2 and CHEK2) and CADD (ATM). Increased risks appeared restricted to functional protein domains for ATM (FAT and PIK domains) and BRCA1 (RING and BRCT domains). For ATM, BRCA1, and BRCA2, data were compatible with small subsets (approximately 7%, 2%, and 0.6%, respectively) of rare missense variants giving similar risk to those of protein truncating variants in the same gene. For CHEK2, data were more consistent with a large fraction (approximately 60%) of rare missense variants giving a lower risk (OR 1.75, 95% CI (1.47-2.08)) than CHEK2 protein truncating variants. There was little evidence for an association with risk for missense variants in PALB2. The best fitting models were well calibrated in the validation set. CONCLUSIONS: These results will inform risk prediction models and the selection of candidate variants for functional assays and could contribute to the clinical reporting of gene panel testing for breast cancer susceptibility.

Pathology of Tumors Associated With Pathogenic Germline Variants in 9 Breast Cancer Susceptibility Genes.

IMPORTANCE: Rare germline genetic variants in several genes are associated with increased breast cancer (BC) risk, but their precise contributions to different disease subtypes are unclear. This information is relevant to guidelines for gene panel testing and risk prediction. OBJECTIVE: To characterize tumors associated with BC susceptibility genes in large-scale population- or hospital-based studies. DESIGN, SETTING, AND PARTICIPANTS: The multicenter, international case-control analysis of the BRIDGES study included 42 680 patients and 46 387 control participants, comprising women aged 18 to 79 years who were sampled independently of family history from 38 studies. Studies were conducted between 1991 and 2016. Sequencing and analysis took place between 2016 and 2021. EXPOSURES: Protein-truncating variants and likely pathogenic missense variants in ATM, BARD1, BRCA1, BRCA2, CHEK2, PALB2, RAD51C, RAD51D, and TP53. MAIN OUTCOMES AND MEASURES: The intrinsic-like BC subtypes as defined by estrogen receptor, progesterone receptor, and ERBB2 (formerly known as HER2) status, and tumor grade; morphology; size; stage; lymph node involvement; subtype-specific odds ratios (ORs) for carrying protein-truncating variants and pathogenic missense variants in the 9 BC susceptibility genes. RESULTS: The mean (SD) ages at interview (control participants) and diagnosis (cases) were 55.1 (11.9) and 55.8 (10.6) years, respectively; all participants were of European or East Asian ethnicity. There was substantial heterogeneity in the distribution of intrinsic subtypes by gene. RAD51C, RAD51D, and BARD1 variants were associated mainly with triple-negative disease (OR, 6.19 [95% CI, 3.17-12.12]; OR, 6.19 [95% CI, 2.99-12.79]; and OR, 10.05 [95% CI, 5.27-19.19], respectively). CHEK2 variants were associated with all subtypes (with ORs ranging from 2.21-3.17) except for triple-negative disease. For ATM variants, the association was strongest for the hormone receptor (HR)+ERBB2- high-grade subtype (OR, 4.99; 95% CI, 3.68-6.76). BRCA1 was associated with increased risk of all subtypes, but the ORs varied widely, being highest for triple-negative disease (OR, 55.32; 95% CI, 40.51-75.55). BRCA2 and PALB2 variants were also associated with triple-negative disease. TP53 variants were most strongly associated with HR+ERBB2+ and HR-ERBB2+ subtypes. Tumors occurring in pathogenic variant carriers were of higher grade. For most genes and subtypes, a decline in ORs was observed with increasing age. Together, the 9 genes were associated with 27.3% of all triple-negative tumors in women 40 years or younger. CONCLUSIONS AND RELEVANCE: The results of this case-control study suggest that variants in the 9 BC risk genes differ substantially in their associated pathology but are generally associated with triple-negative and/or high-grade disease. Knowing the age and tumor subtype distributions associated with individual BC genes can potentially aid guidelines for gene panel testing, risk prediction, and variant classification and guide targeted screening strategies.

A genome-wide association study of radiotherapy induced toxicity in head and neck cancer patients identifies a susceptibility locus associated with mucositis.

PURPOSE: A two-stage genome-wide association study was carried out in head and neck cancer (HNC) patients aiming to identify genetic variants associated with either specific radiotherapy-induced (RT) toxicity endpoints or a general proneness to develop toxicity after RT. MATERIALS AND METHODS: The analysis included 1780 HNC patients treated with primary RT for laryngeal or oro/hypopharyngeal cancers. In a non-hypothesis-driven explorative discovery study, associations were tested in 1183 patients treated within The Danish Head and Neck Cancer Group. Significant associations were later tested in an independent Dutch cohort of 597 HNC patients and if replicated, summary data obtained from discovery and replication studies were meta-analysed. Further validation of significantly replicated findings was pursued in an Asian cohort of 235 HNC patients with nasopharynx as the primary tumour site. RESULTS: We found and replicated a significant association between a locus on chromosome 5 and mucositis with a pooled OR for rs1131769*C in meta-analysis = 1.95 (95% CI 1.48-2.41; ppooled = 4.34 × 10-16). CONCLUSION: This first exploratory GWAS in European cohorts of HNC patients identified and replicated a risk locus for mucositis. A larger Meta-GWAS to identify further risk variants for RT-induced toxicity in HNC patients is warranted.

Rare germline copy number variants (CNVs) and breast cancer risk.

Germline copy number variants (CNVs) are pervasive in the human genome but potential disease associations with rare CNVs have not been comprehensively assessed in large datasets. We analysed rare CNVs in genes and non-coding regions for 86,788 breast cancer cases and 76,122 controls of European ancestry with genome-wide array data. Gene burden tests detected the strongest association for deletions in BRCA1 (P = 3.7E-18). Nine other genes were associated with a p-value < 0.01 including known susceptibility genes CHEK2 (P = 0.0008), ATM (P = 0.002) and BRCA2 (P = 0.008). Outside the known genes we detected associations with p-values < 0.001 for either overall or subtype-specific breast cancer at nine deletion regions and four duplication regions. Three of the deletion regions were in established common susceptibility loci. To the best of our knowledge, this is the first genome-wide analysis of rare CNVs in a large breast cancer case-control dataset. We detected associations with exonic deletions in established breast cancer susceptibility genes. We also detected suggestive associations with non-coding CNVs in known and novel loci with large effects sizes. Larger sample sizes will be required to reach robust levels of statistical significance.

First international workshop of the ATM and cancer risk group (4-5 December 2019).

The first International Workshop of the ATM and Cancer Risk group focusing on the role of Ataxia-Telangiectasia Mutated (ATM) gene in cancer was held on December 4 and 5, 2019 at Institut Curie in Paris, France. It was motivated by the fact that germline ATM pathogenic variants have been found to be associated with different cancer types. However, due to the lack of precise age-, sex-, and site-specific risk estimates, no consensus on management guidelines for variant carriers exists, and the clinical utility of ATM variant testing is uncertain. The meeting brought together epidemiologists, geneticists, biologists and clinicians to review current knowledge and on-going challenges related to ATM and cancer risk. This report summarizes the meeting sessions content that covered the latest results in family-based and population-based studies, the importance of accurate variant classification, the effect of radiation exposures for ATM variant carriers, and the characteristics of ATM-deficient tumors. The report concludes that ATM variant carriers outside of the context of Ataxia-Telangiectasia may benefit from effective cancer risk management and therapeutic strategies and that efforts to set up large-scale studies in the international framework to achieve this goal are necessary.

Splicing predictions, minigene analyses, and ACMG-AMP clinical classification of 42 germline PALB2 splice-site variants.

PALB2 loss-of-function variants confer high risk of developing breast cancer. Here we present a systematic functional analysis of PALB2 splice-site variants detected in approximately 113,000 women in the large-scale sequencing project Breast Cancer After Diagnostic Gene Sequencing (BRIDGES; https://bridges-research.eu/). Eighty-two PALB2 variants at the intron-exon boundaries were analyzed with MaxEntScan. Forty-two variants were selected for the subsequent splicing functional assays. For this purpose, three splicing reporter minigenes comprising exons 1-12 were constructed. The 42 potential spliceogenic variants were introduced into the minigenes by site-directed mutagenesis and assayed in MCF-7/MDA-MB-231 cells. Splicing anomalies were observed in 35 variants, 23 of which showed no traces or minimal amounts of the expected full-length transcripts of each minigene. More than 30 different variant-induced transcripts were characterized, 23 of which were predicted to truncate the PALB2 protein. The pathogenicity of all variants was interpreted according to an in-house adaptation of the American College of Medical Genetics and Genomics and the Association for Molecular Pathology (ACMG-AMP) variant classification scheme. Up to 23 variants were classified as pathogenic/likely pathogenic. Remarkably, three ±1,2 variants (c.49-2A>T, c.108+2T>C, and c.211+1G>A) were classified as variants of unknown significance, as they produced significant amounts of either in-frame transcripts of unknown impact on the PALB2 protein function or the minigene full-length transcripts. In conclusion, we have significantly contributed to the ongoing effort of identifying spliceogenic variants in the clinically relevant PALB2 cancer susceptibility gene. Moreover, we suggest some approaches to classify the findings in accordance with the ACMG-AMP rationale. © 2021 The Authors. The Journal of Pathology published by John Wiley & Sons, Ltd on behalf of The Pathological Society of Great Britain and Ireland.

Breast Cancer Risk Genes - Association Analysis in More than 113,000 Women.

BACKGROUND: Genetic testing for breast cancer susceptibility is widely used, but for many genes, evidence of an association with breast cancer is weak, underlying risk estimates are imprecise, and reliable subtype-specific risk estimates are lacking. METHODS: We used a panel of 34 putative susceptibility genes to perform sequencing on samples from 60,466 women with breast cancer and 53,461 controls. In separate analyses for protein-truncating variants and rare missense variants in these genes, we estimated odds ratios for breast cancer overall and tumor subtypes. We evaluated missense-variant associations according to domain and classification of pathogenicity. RESULTS: Protein-truncating variants in 5 genes (ATM, BRCA1, BRCA2, CHEK2, and PALB2) were associated with a risk of breast cancer overall with a P value of less than 0.0001. Protein-truncating variants in 4 other genes (BARD1, RAD51C, RAD51D, and TP53) were associated with a risk of breast cancer overall with a P value of less than 0.05 and a Bayesian false-discovery probability of less than 0.05. For protein-truncating variants in 19 of the remaining 25 genes, the upper limit of the 95% confidence interval of the odds ratio for breast cancer overall was less than 2.0. For protein-truncating variants in ATM and CHEK2, odds ratios were higher for estrogen receptor (ER)-positive disease than for ER-negative disease; for protein-truncating variants in BARD1, BRCA1, BRCA2, PALB2, RAD51C, and RAD51D, odds ratios were higher for ER-negative disease than for ER-positive disease. Rare missense variants (in aggregate) in ATM, CHEK2, and TP53 were associated with a risk of breast cancer overall with a P value of less than 0.001. For BRCA1, BRCA2, and TP53, missense variants (in aggregate) that would be classified as pathogenic according to standard criteria were associated with a risk of breast cancer overall, with the risk being similar to that of protein-truncating variants. CONCLUSIONS: The results of this study define the genes that are most clinically useful for inclusion on panels for the prediction of breast cancer risk, as well as provide estimates of the risks associated with protein-truncating variants, to guide genetic counseling. (Funded by European Union Horizon 2020 programs and others.).

Radiogenomics Consortium Genome-Wide Association Study Meta-Analysis of Late Toxicity After Prostate Cancer Radiotherapy.

BACKGROUND: A total of 10%-20% of patients develop long-term toxicity following radiotherapy for prostate cancer. Identification of common genetic variants associated with susceptibility to radiotoxicity might improve risk prediction and inform functional mechanistic studies. METHODS: We conducted an individual patient data meta-analysis of six genome-wide association studies (n = 3871) in men of European ancestry who underwent radiotherapy for prostate cancer. Radiotoxicities (increased urinary frequency, decreased urinary stream, hematuria, rectal bleeding) were graded prospectively. We used grouped relative risk models to test associations with approximately 6 million genotyped or imputed variants (time to first grade 2 or higher toxicity event). Variants with two-sided Pmeta less than 5 × 10-8 were considered statistically significant. Bayesian false discovery probability provided an additional measure of confidence. Statistically significant variants were evaluated in three Japanese cohorts (n = 962). All statistical tests were two-sided. RESULTS: Meta-analysis of the European ancestry cohorts identified three genomic signals: single nucleotide polymorphism rs17055178 with rectal bleeding (Pmeta = 6.2 × 10-10), rs10969913 with decreased urinary stream (Pmeta = 2.9 × 10-10), and rs11122573 with hematuria (Pmeta = 1.8 × 10-8). Fine-scale mapping of these three regions was used to identify another independent signal (rs147121532) associated with hematuria (Pconditional = 4.7 × 10-6). Credible causal variants at these four signals lie in gene-regulatory regions, some modulating expression of nearby genes. Previously identified variants showed consistent associations (rs17599026 with increased urinary frequency, rs7720298 with decreased urinary stream, rs1801516 with overall toxicity) in new cohorts. rs10969913 and rs17599026 had similar effects in the photon-treated Japanese cohorts. CONCLUSIONS: This study increases the understanding of the architecture of common genetic variants affecting radiotoxicity, points to novel radio-pathogenic mechanisms, and develops risk models for testing in clinical studies. Further multinational radiogenomics studies in larger cohorts are worthwhile.

Cancer Risks Associated With Germline PALB2 Pathogenic Variants: An International Study of 524 Families.

PURPOSE: To estimate age-specific relative and absolute cancer risks of breast cancer and to estimate risks of ovarian, pancreatic, male breast, prostate, and colorectal cancers associated with germline PALB2 pathogenic variants (PVs) because these risks have not been extensively characterized. METHODS: We analyzed data from 524 families with PALB2 PVs from 21 countries. Complex segregation analysis was used to estimate relative risks (RRs; relative to country-specific population incidences) and absolute risks of cancers. The models allowed for residual familial aggregation of breast and ovarian cancer and were adjusted for the family-specific ascertainment schemes. RESULTS: We found associations between PALB2 PVs and risk of female breast cancer (RR, 7.18; 95% CI, 5.82 to 8.85; P = 6.5 × 10-76), ovarian cancer (RR, 2.91; 95% CI, 1.40 to 6.04; P = 4.1 × 10-3), pancreatic cancer (RR, 2.37; 95% CI, 1.24 to 4.50; P = 8.7 × 10-3), and male breast cancer (RR, 7.34; 95% CI, 1.28 to 42.18; P = 2.6 × 10-2). There was no evidence for increased risks of prostate or colorectal cancer. The breast cancer RRs declined with age (P for trend = 2.0 × 10-3). After adjusting for family ascertainment, breast cancer risk estimates on the basis of multiple case families were similar to the estimates from families ascertained through population-based studies (P for difference = .41). On the basis of the combined data, the estimated risks to age 80 years were 53% (95% CI, 44% to 63%) for female breast cancer, 5% (95% CI, 2% to 10%) for ovarian cancer, 2%-3% (95% CI females, 1% to 4%; 95% CI males, 2% to 5%) for pancreatic cancer, and 1% (95% CI, 0.2% to 5%) for male breast cancer. CONCLUSION: These results confirm PALB2 as a major breast cancer susceptibility gene and establish substantial associations between germline PALB2 PVs and ovarian, pancreatic, and male breast cancers. These findings will facilitate incorporation of PALB2 into risk prediction models and optimize the clinical cancer risk management of PALB2 PV carriers.

Prevalence of BRCA1 and BRCA2 pathogenic variants in a large, unselected breast cancer cohort.

Breast cancer patients with BRCA1/2-driven tumors may benefit from targeted therapy. It is not clear whether current BRCA screening guidelines are effective at identifying these patients. The purpose of our study was to evaluate the prevalence of inherited BRCA1/2 pathogenic variants in a large, clinically representative breast cancer cohort and to estimate the proportion of BRCA1/2 carriers not detected by selectively screening individuals with the highest probability of being carriers according to current clinical guidelines. The study included 5,122 unselected Swedish breast cancer patients diagnosed from 2001 to 2008. Target sequence enrichment (48.48 Fluidigm Access Arrays) and sequencing were performed (Illumina Hi-Seq 2,500 instrument, v4 chemistry). Differences in patient and tumor characteristics of BRCA1/2 carriers who were already identified as part of clinical BRCA1/2 testing routines and additional BRCA1/2 carriers found by sequencing the entire study population were compared using logistic regression models. Ninety-two of 5,099 patients with valid variant calls were identified as BRCA1/2 carriers by screening all study participants (1.8%). Only 416 study participants (8.2%) were screened as part of clinical practice, but this identified 35 out of 92 carriers (38.0%). Clinically identified carriers were younger, less likely postmenopausal and more likely to be associated with familiar ovarian cancer compared to the additional carriers identified by screening all patients. More BRCA2 (34/42, 81.0%) than BRCA1 carriers (23/50, 46%) were missed by clinical screening. In conclusion, BRCA1/2 mutation prevalence in unselected breast cancer patients was 1.8%. Six in ten BRCA carriers were not detected by selective clinical screening of individuals.

Differential Burden of Rare and Common Variants on Tumor Characteristics, Survival, and Mode of Detection in Breast Cancer.

Genetic variants that increase breast cancer risk can be rare or common. This study tests whether the genetic risk stratification of breast cancer by rare and common variants in established loci can discriminate tumors with different biology, patient survival, and mode of detection. Multinomial logistic regression tested associations between genetic risk load [protein-truncating variant (PTV) carriership in 31 breast cancer predisposition genes-or polygenic risk score (PRS) using 162 single-nucleotide polymorphisms], tumor characteristics, and mode of detection (OR). Ten-year breast cancer-specific survival (HR) was estimated using Cox regression models. In this unselected cohort of 5,099 patients with breast cancer diagnosed in Sweden between 2001 and 2008, PTV carriers (n = 597) were younger and associated with more aggressive tumor phenotypes (ER-negative, large size, high grade, high proliferation, luminal B, and basal-like subtype) and worse outcome (HR, 1.65; 1.16-2.36) than noncarriers. After excluding 92 BRCA1/2 carriers, PTV carriership remained associated with high grade and worse survival (HR, 1.76; 1.21-2.56). In 5,007 BRCA1/2 noncarriers, higher PRS was associated with less aggressive tumor characteristics (ER-positive, PR-positive, small size, low grade, low proliferation, and luminal A subtype). Among patients with low mammographic density (<25%), non-BRCA1/2 PTV carriers were more often interval than screen-detected breast cancer (OR, 1.89; 1.12-3.21) than noncarriers. In contrast, higher PRS was associated with lower risk of interval compared with screen-detected cancer (OR, 0.77; 0.64-0.93) in women with low mammographic density. These findings suggest that rare and common breast cancer susceptibility loci are differentially associated with tumor characteristics, survival, and mode of detection.Significance: These findings offer the potential to improve screening practices for breast cancer by providing a deeper understanding of how risk variants affect disease progression and mode of detection. Cancer Res; 78(21); 6329-38. ©2018 AACR.

Inherited mutations in BRCA1 and BRCA2 in an unselected multiethnic cohort of Asian patients with breast cancer and healthy controls from Malaysia.

BACKGROUND: Genetic testing for BRCA1 and BRCA2 is offered typically to selected women based on age of onset and family history of cancer. However, current internationally accepted genetic testing referral guidelines are built mostly on data from cancer genetics clinics in women of European descent. To evaluate the appropriateness of such guidelines in Asians, we have determined the prevalence of germ line variants in an unselected cohort of Asian patients with breast cancer and healthy controls. METHODS: Germ line DNA from a hospital-based study of 2575 unselected patients with breast cancer and 2809 healthy controls were subjected to amplicon-based targeted sequencing of exonic and proximal splice site junction regions of BRCA1 and BRCA2 using the Fluidigm Access Array system, with sequencing conducted on a Illumina HiSeq2500 platform. Variant calling was performed with GATK UnifiedGenotyper and were validated by Sanger sequencing. RESULTS: Fifty-five (2.1%) BRCA1 and 66 (2.6%) BRCA2 deleterious mutations were identified among patients with breast cancer and five (0.18%) BRCA1 and six (0.21%) BRCA2 mutations among controls. One thousand one hundred and eighty-six (46%) patients and 97 (80%) carriers fulfilled the National Comprehensive Cancer Network guidelines for genetic testing. CONCLUSION: Five per cent of unselected Asian patients with breast cancer carry deleterious variants in BRCA1 or BRCA2. While current referral guidelines identified the majority of carriers, one in two patients would be referred for genetic services. Given that such services are largely unavailable in majority of low-resource settings in Asia, our study highlights the need for more efficient guidelines to identify at-risk individuals in Asia.