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Background: Inherited mutations in DNA-repair genes such as BRCA2 are associated with increased risks of lethal prostate cancer. Although the prevalence of germline mutations in DNA-repair genes among men with localized prostate cancer who are unselected for family predisposition is insufficient to warrant routine testing, the frequency of such mutations in patients with metastatic prostate cancer has not been established., Methods: We recruited 692 men with documented metastatic prostate cancer who were unselected for family history of cancer or age at diagnosis. We isolated germline DNA and used multiplex sequencing assays to assess mutations in 20 DNA-repair genes associated with autosomal dominant cancer-predisposition syndromes., Results: A total of 84 germline DNA-repair gene mutations that were presumed to be deleterious were identified in 82 men (11.8%); mutations were found in 16 genes, including BRCA2 (37 men [5.3%]), ATM (11 [1.6%]), CHEK2 (10 [1.9% of 534 men with data]), BRCA1 (6 [0.9%]), RAD51D (3 [0.4%]), and PALB2 (3 [0.4%]). Mutation frequencies did not differ according to whether a family history of prostate cancer was present or according to age at diagnosis. Overall, the frequency of germline mutations in DNA-repair genes among men with metastatic prostate cancer significantly exceeded the prevalence of 4.6% among 499 men with localized prostate cancer (P<0.001), including men with high-risk disease, and the prevalence of 2.7% in the Exome Aggregation Consortium, which includes 53,105 persons without a known cancer diagnosis (P<0.001)., Conclusions: In our multicenter study, the incidence of germline mutations in genes mediating DNA-repair processes among men with metastatic prostate cancer was 11.8%, which was significantly higher than the incidence among men with localized prostate cancer. The frequencies of germline mutations in DNA-repair genes among men with metastatic disease did not differ significantly according to age at diagnosis or family history of prostate cancer. (Funded by Stand Up To Cancer and others.).

Importance: Germline mutations in BRCA1 and BRCA2 are relatively common in women with ovarian, fallopian tube, and peritoneal carcinoma (OC) causing a greatly increased lifetime risk of these cancers, but the frequency and relevance of inherited mutations in other genes is less well characterized., Objective: To determine the frequency and importance of germline mutations in cancer-associated genes in OC., Design, Setting, and Participants: A study population of 1915 woman with OC and available germline DNA were identified from the University of Washington (UW) gynecologic tissue bank (n = 570) and from Gynecologic Oncology Group (GOG) phase III clinical trials 218 (n = 788) and 262 (n = 557). Patients were enrolled at diagnosis and were not selected for age or family history. Germline DNA was sequenced from women with OC using a targeted capture and multiplex sequencing assay., Main Outcomes and Measures: Mutation frequencies in OC were compared with the National Heart, Lung, and Blood Institute GO Exome Sequencing Project (ESP) and the Exome Aggregation Consortium (ExAC). Clinical characteristics and survival were assessed by mutation status., Results: Overall, the median (range) age at diagnosis was 60 (28-91) years in patients recruited from UW and 61 (23-87) years in patients recruited from the GOG trials. A higher number of black women were recruited from the GOG trials (4.3% vs 1.4%; P = .009); but in patients recruited from UW, there was a higher proportion of fallopian tube carcinomas (13.3% vs 5.7%; P < .001); stage I and II disease (14.6% vs 0% [GOG trials were restricted to advanced-stage cancer]); and nonserous carcinomas (29.9% vs 13.1%, P < .001). Of 1915 patients, 280 (15%) had mutations in BRCA1 (n = 182), or BRCA2 (n = 98), and 8 (0.4%) had mutations in DNA mismatch repair genes. Mutations in BRIP1 (n = 26), RAD51C (n = 11), RAD51D (n = 11), PALB2 (n = 12), and BARD1 (n = 4) were significantly more common in patients with OC than in the ESP or ExAC, present in 3.3%. Race, histologic subtype, and disease site were not predictive of mutation frequency. Patients with a BRCA2 mutation from the GOG trials had longer progression-free survival (hazard ratio [HR], 0.60; 95% CI, 0.45-0.79; P < .001) and overall survival (HR, 0.39; 95% CI, 0.25-0.60; P < .001) compared with those without mutations., Conclusions and Relevance: Of 1915 patients with OC, 347 (18%) carried pathogenic germline mutations in genes associated with OC risk. PALB2 and BARD1 are suspected OC genes and together with established OC genes (BRCA1, BRCA2, BRIP1, RAD51C, RAD51D, MSH2, MLH1, PMS2, and MSH6) bring the total number of genes suspected to cause hereditary OC to 11.

Background: Germline loss-of-function mutations in PALB2 are known to confer a predisposition to breast cancer. However, the lifetime risk of breast cancer that is conferred by such mutations remains unknown., Methods: We analyzed the risk of breast cancer among 362 members of 154 families who had deleterious truncating, splice, or deletion mutations in PALB2. The age-specific breast-cancer risk for mutation carriers was estimated with the use of a modified segregation-analysis approach that allowed for the effects of PALB2 genotype and residual familial aggregation., Results: The risk of breast cancer for female PALB2 mutation carriers, as compared with the general population, was eight to nine times as high among those younger than 40 years of age, six to eight times as high among those 40 to 60 years of age, and five times as high among those older than 60 years of age. The estimated cumulative risk of breast cancer among female mutation carriers was 14% (95% confidence interval [CI], 9 to 20) by 50 years of age and 35% (95% CI, 26 to 46) by 70 years of age. Breast-cancer risk was also significantly influenced by birth cohort (P<0.001) and by other familial factors (P=0.04). The absolute breast-cancer risk for PALB2 female mutation carriers by 70 years of age ranged from 33% (95% CI, 25 to 44) for those with no family history of breast cancer to 58% (95% CI, 50 to 66) for those with two or more first-degree relatives with breast cancer at 50 years of age., Conclusions: Loss-of-function mutations in PALB2 are an important cause of hereditary breast cancer, with respect both to the frequency of cancer-predisposing mutations and to the risk associated with them. Our data suggest the breast-cancer risk for PALB2 mutation carriers may overlap with that for BRCA2 mutation carriers. (Funded by the European Research Council and others.).

Objectives: Few studies have comprehensively tested all ovarian cancer patients for BRCA1 and BRCA2 (BRCA1/2) mutations. We sought to determine if clinically identified mutation carriers differed in clinical characteristics and outcomes from mutation carriers not identified during routine clinical care., Methods: We included women with ovarian, tubal or peritoneal carcinoma. BROCA, an assay using targeted capture and massively parallel sequencing was used to identify mutations in BRCA1/2 and 19 other tumor suppressor genes. We identified subjects with BRCA1/2 mutations using BROCA that had not previously received standard genetic testing (BROCA, n=37) and compared them to subjects with BRCA1/2 mutations identified during routine clinical care (known, n=70), and to those wildtype for 21 genes using BROCA (wildtype, n=291)., Results: BROCA mutation carriers were older than known carriers, median age of 58 (range 41-77), vs. 51 (range 33-76, p=0.003, Mann-Whitney). 58/70 (82.9%) of known carriers had a strong family history, compared with 15/37 (40.5%) of BROCA carriers, p<0.0001, (Fisher's Exact). Median overall survival was significantly worse for BROCA mutation carriers compared to known mutation carriers, (45 vs. 93months, p<0.0001, HR 3.47 (1.79-6.72), Log-rank test). The improved survival for BRCA1/2 mutation carriers (known and BROCA) compared with wildtype cases (69 vs. 44months, p=0.0001, HR 0.58 (0.43-0.77), Log-rank test) was driven by known mutation carriers., Conclusions: Older age, absence of a strong family history, and poor survival are all associated with decreased clinical identification of inherited BRCA1/2 mutations in women with ovarian cancer. Using age and family history to direct genetic testing will miss a significant percentage of mutation carriers. Testing should be initiated at the time of diagnosis to maximize identification of mutations and minimize survival bias., (Published by Elsevier Inc.)

Objective: RAD51D, a gene in the Fanconi Anemia-BRCA homologous recombination pathway, has recently been shown to harbor germline mutations responsible for ovarian carcinoma in multiply affected families. We aimed to extend these results to ovarian carcinoma in the general population., Methods: We sequenced RAD51D in germline DNA from 360 individuals with primary ovarian, peritoneal or fallopian tube carcinoma who were not selected for age of cancer onset or family history. We also sequenced RAD51D in 459 probands from 226 high risk breast cancer families who were wild type for 21 breast and ovarian cancer genes., Results: Of 360 cases, three (0.8%) carried loss-of-function mutations in RAD51D. All three subjects had ovarian carcinoma; one was also diagnosed with a synchronous endometrial carcinoma. Only one of the three subjects had a family history of breast or ovarian cancer. Combined with previous data for this series, 23.9% of women with unselected ovarian, fallopian tube, or peritoneal carcinoma carried a germline loss-of-function mutation in any of 13 tumor suppressor genes. Among the 449 women and 10 men with familial breast cancer, none carried a loss of function mutation in RAD51D., Conclusions: These data support the previous observation that loss-of-function mutations in RAD51D predispose to ovarian carcinoma but not to breast carcinoma. We conclude that inherited ovarian cancer is highly heterogeneous genetically, and that approximately one in four ovarian carcinoma patients carry a germline mutation in a known tumor suppressor gene that confers high risk., (Copyright © 2012 Elsevier Inc. All rights reserved.)

Inherited loss-of-function mutations in BRCA1 and BRCA2 and other tumor suppressor genes predispose to ovarian carcinomas, but the overall burden of disease due to inherited mutations is not known. Using targeted capture and massively parallel genomic sequencing, we screened for germ-line mutations in 21 tumor suppressor genes in genomic DNA from women with primary ovarian, peritoneal, or fallopian tube carcinoma. Subjects were consecutively enrolled at diagnosis and not selected for age or family history. All classes of mutations, including point mutations and large genomic deletions and insertions, were detected. Of 360 subjects, 24% carried germ-line loss-of-function mutations: 18% in BRCA1 or BRCA2 and 6% in BARD1, BRIP1, CHEK2, MRE11A, MSH6, NBN, PALB2, RAD50, RAD51C, or TP53. Six of these genes were not previously implicated in inherited ovarian carcinoma. Primary carcinomas were generally characterized by genomic loss of normal alleles of the mutant genes. Of women with inherited mutations, >30% had no family history of breast or ovarian carcinoma, and >35% were 60 y or older at diagnosis. More patients with ovarian carcinoma carry cancer-predisposing mutations and in more genes than previously appreciated. Comprehensive genetic testing for inherited carcinoma is warranted for all women with ovarian, peritoneal, or fallopian tube carcinoma, regardless of age or family history. Clinical genetic testing is currently done gene by gene, with each test costing thousands of dollars. In contrast, massively parallel sequencing allows such testing for many genes simultaneously at low cost.

We describe the Multiple-Dye Cleavase Fragment Length Polymorphism (MD-CFLP) method set up for a sensitive and preliminary rapid screening of BRCA1 mutations. We analysed exons 11 and 16, which are known to cover slightly more than 70% of the whole coding region of the gene, subdivided into 4 amplicons and labelled with different fluorescent dUTPs. MD-CFLP was first utilised on a panel of 30 DNA samples in which the presence of single-base substitutions or small deletions/insertions had been previously identified by direct sequencing as gold standard, in order to define the optimal conditions in terms of PCR amplification and temperature of digestion. In a second step, we blindly analysed 21 DNA samples by MD-CFLP to verify its reliability. The sensitivity and specificity of MD-CFLP were both 100% in the first study, and 80% and 94%, respectively, in the blind sample assay. Our results demonstrate the capability of the MD-CFLP method to detect DNA sequence alterations in fragments of more than 1 kb. We conclude that CFLP is a powerful tool in mutational analysis, offering reliable results in a shorter time and at a lower cost than conventional methods, and its potential can be enhanced when internal fluorescent labelling and laser detection are used., (Copyright 2001 Cancer Research Campaign http://www.bjcancer.com.)