Sequencing for germline mutations in Swedish breast cancer families reveals novel breast cancer risk genes.
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ABSTRACT:
SUBMITTER: Helgadottir HT
PROVIDER: S-EPMC8289997 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:BRCA1 and BRCA2 are the most well-known breast cancer susceptibility genes. Additional genes involved in DNA repair have been identified as predisposing to breast cancer. One such gene, RAD51C, is essential for homologous recombination repair. Several likely pathogenic RAD51C mutations have been identified in BRCA1- and BRCA2-negative breast and ovarian cancer families. We performed complete sequencing of RAD51C in germline DNA of 286 female breast and/or ovarian cancer cases with a family history of breast and ovarian cancers, who had previously tested negative for mutations in BRCA1 and BRCA2. We screened 133 breast cancer cases, 119 ovarian cancer cases, and 34 with both breast and ovarian cancers. Fifteen DNA sequence variants were identified; including four intronic, one 5' UTR, one promoter, three synonymous, and six non-synonymous variants. None were truncating. The in-silico SIFT and Polyphen programs were used to predict possible pathogenicity of the six non-synonomous variants based on sequence conservation. G153D and T287A were predicted to be likely pathogenic. Two additional variants, A126T and R214C alter amino acids in important domains of the protein such that they could be pathogenic. Two-hybrid screening and immunoblot analyses were performed to assess the functionality of these four non-synonomous variants in yeast. The RAD51C-G153D protein displayed no detectable interaction with either XRCC3 or RAD51B, and RAD51C-R214C displayed significantly decreased interaction with both XRCC3 and RAD51B (p<0.001). Immunoblots of RAD51C-Gal4 activation domain fusion peptides showed protein levels of RAD51C-G153D and RAD51C-R214C that were 50% and 60% of the wild-type, respectively. Based on these data, the RAD51C-G153D variant is likely to be pathogenic, while the RAD51C- R214C variant is hypomorphic of uncertain pathogenicity. These results provide further support that RAD51C is a rare breast and ovarian cancer susceptibility gene.
| S-EPMC3182241 | biostudies-literature
Project description:In this study we investigated 45 German breast/ovarian cancer families for germline mutations in the BRCA1 gene. We identified four germline mutations in three breast cancer families and in one breast-ovarian cancer family. among these were one frameshift mutation, one nonsense mutation, one novel splice site mutation, and one missense mutation. The missense mutation was also found in 2.8% of the general population, suggesting that it is not disease associated. The average age of disease onset in those families harbouring causative mutations was between 32.3 and 37.4 years, whereas the family harbouring the missense mutation had an average age of onset of 51.2 years. These findings show that BRCA1 is implicated in a small fraction of breast/ovarian cancer families suggesting the involvement of another susceptibility gene(s).
| S-EPMC1051113 | biostudies-other
Project description:BACKGROUND:Multigene panels are routinely used to assess for predisposing germline mutations in families at high breast cancer risk. The number of variants of unknown significance thereby identified increases with the number of sequenced genes. We aimed to determine whether tumor sequencing can help refine the analysis of germline variants based on second somatic genetic events in the same gene. METHODS:Whole-exome sequencing (WES) was performed on whole blood DNA from 70 unrelated breast cancer patients referred for genetic testing and without a BRCA1, BRCA2, TP53, or CHEK2 mutation. Rare variants were retained in a list of 735 genes. WES was performed on matched tumor DNA to identify somatic second hits (copy number alterations (CNAs) or mutations) in the same genes. Distinct methods (among which immunohistochemistry, mutational signatures, homologous recombination deficiency, and tumor mutation burden analyses) were used to further study the role of the variants in tumor development, as appropriate. RESULTS:Sixty-eight patients (97%) carried at least one germline variant (4.7?±?2.0 variants per patient). Of the 329 variants, 55 (17%) presented a second hit in paired tumor tissue. Of these, 53 were CNAs, resulting in tumor enrichment (28 variants) or depletion (25 variants) of the germline variant. Eleven patients received variant disclosure, with clinical measures for five of them. Seven variants in breast cancer-predisposing genes were considered not implicated in oncogenesis. One patient presented significant tumor enrichment of a germline variant in the oncogene ERBB2, in vitro expression of which caused downstream signaling pathway activation. CONCLUSION:Tumor sequencing is a powerful approach to refine variant interpretation in cancer-predisposing genes in high-risk breast cancer patients. In this series, the strategy provided clinically relevant information for 11 out of 70 patients (16%), adapted to the considered gene and the familial clinical phenotype.
| S-EPMC7161277 | biostudies-literature
Project description: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.).
| S-EPMC4157599 | biostudies-literature
Project description:The presence of germline and somatic deleterious mutations in the BRCA1 and BRCA2 genes has important clinical consequences for breast cancer (BC) patients. Analysis of the mutational status in BRCA genes is not yet common in public Latin American institutions; thus, our objective was to implement high-performance technology with highly reliable results with the possibility of analyzing several patients simultaneously, therefore reducing cost and work time. A prospective cohort of 252 unrelated sporadic breast cancer patients from the Mexican-mestizo population were analyzed using next generation sequencing (NGS) based on ion semiconductor sequencing. We found 28 pathogenic mutations (25 in BRCA1 and 13 in BRCA2), 11 of which had not been reported previously in Hispanic or Latin American populations. A total of 38 patients were positive for a pathogenic mutation representing 15% of our Mexican women cohort with breast cancer; 25 for BRCA1; and 13 for BRCA2. Our results revealed that there are mutations not analyzed by mutations panels, and our findings support the suitability of massive sequencing approaches in the public institutions of developing countries. Hence, BRCA screening should be offered to patients with breast cancer regardless of their family history of cancer in order to identify unaffected family carriers.
| S-EPMC6769960 | biostudies-literature
Project description:To estimate the proportion of breast cancer families due to BRCA1 or BRCA2, we performed mutation screening of the entire coding regions of both genes supplemented with linkage analysis of 31 families, 8 containing male breast cancers and 23 site-specific female breast cancer. A combination of protein-truncation test and SSCP or heteroduplex analyses was used for mutation screening complemented, where possible, by the analysis of expression level of BRCA1 and BRCA2 alleles. Six of the eight families with male breast cancer revealed frameshift mutations, two in BRCA1 and four in BRCA2. Although most families with female site-specific breast cancers were thought to be due to mutations in either BRCA1 or BRCA2, we identified only eight mutations in our series of 23 site-specific female breast cancer families (34%), four in BRCA1 and four in BRCA2. According to the posterior probabilities calculated for mutation-negative families, based on linkage data and mutation screening results, we would expect 8-10 site-specific female breast cancer families of our series to be due to neither BRCA1 nor BRCA2. Thus, our results suggest the existence of at least one more major breast cancer-susceptibility gene.
| S-EPMC1712515 | biostudies-other
Project description:Breast cancer (BC) is a genetically heterogeneous disease with high prevalence in Northern Europe. However, there has been no detailed investigation into the Scandinavian somatic landscape. Here, in a homogeneous Swedish cohort, we describe the somatic events underlying BC, leveraging a targeted next-generation sequencing approach. We designed a 20.5 Mb array targeting coding and regulatory regions of genes with a known role in BC (n = 765). The selected genes were either from human BC studies (n = 294) or from within canine mammary tumor associated regions (n = 471). A set of predominantly estrogen receptor positive tumors (ER + 85%) and their normal tissue counterparts (n = 61) were sequenced to ~ 140 × and 85 × mean target coverage, respectively. MuTect2 and VarScan2 were employed to detect single nucleotide variants (SNVs) and copy number aberrations (CNAs), while MutSigCV (SNVs) and GISTIC (CNAs) algorithms estimated the significance of recurrent somatic events. The significantly mutated genes (q ≤ 0.01) were PIK3CA (28% of patients), TP53 (21%) and CDH1 (11%). However, histone modifying genes contained the largest number of variants (KMT2C and ARID1A, together 28%). Mutations in KMT2C were mutually exclusive with PI3KCA mutations (p ≤ 0. 001) and half of these affect the formation of a functional PHD domain. The tumor suppressor CDK10 was deleted in 80% of the cohort while the oncogene MDM4 was amplified. Mutational signature analyses pointed towards APOBEC deaminase activity (COSMIC signature 2) and DNA mismatch repair (COSMIC signature 6). We noticed two significantly distinct patterns related to patient age; TP53 being more mutated in the younger group (29% vs 9% of patients) and CDH23 mutations were absent from the older group. The increased somatic mutation prevalence in the histone modifying genes KMT2C and ARID1A distinguishes the Swedish cohort from previous studies. KMT2C regulates enhancer activation and assists tumor proliferation in a hormone-rich environment, possibly pointing to a role in ER + BC, especially in older cases. Finally, age of onset appears to affect the mutational landscape suggesting that a larger age-diverse population incorporating more molecular subtypes should be studied to elucidate the underlying mechanisms.
| S-EPMC7653953 | biostudies-literature
Project description:BackgroundRAD51C and RAD51D are crucial in homologous recombination (HR) DNA repair. The prevalence of the RAD51C and RAD51D mutations in breast cancer varies across ethnic groups. Associations of RAD51C and RAD51D germline pathogenic variants (GPVs) with breast and ovarian cancer predisposition have been recently reported and are of interest.MethodsWe performed multi-gene panel sequencing to study the prevalence of RAD51C and RAD51D germline mutations among 3728 patients with hereditary breast and/or ovarian cancer (HBOC).ResultsWe identified 18 pathogenic RAD51C and RAD51D mutation carriers, with a mutation frequency of 0.13% (5/3728) and 0.35% (13/3728), respectively. The most common recurrent mutation was RAD51D c.270_271dupTA; p.(Lys91Ilefs*13), with a mutation frequency of 0.30% (11/3728), which was also commonly identified in Asians. Only four out of six cases (66.7%) of this common mutation tested positive for homologous recombination deficiency (HRD).ConclusionsTaking the family studies in our registry and tumor molecular pathology together, we concluded that this relatively common RAD51D variant showed incomplete penetrance in our local Chinese community. Personalized genetic counseling emphasizing family history for families with this variant, as suggested at the UK Cancer Genetics Group (UKCGG) Consensus meeting, would also be appropriate in Chinese families.
| S-EPMC11355318 | biostudies-literature
Project description:Background:Conventional methods used to identify BRCA1 and BRCA2 germline mutations in hereditary cancers, such as Sanger sequencing/multiplex ligation-dependent probe amplification (MLPA), are time-consuming and expensive, due to the large size of the genes. The recent introduction of next-generation sequencing (NGS) benchtop platforms offered a powerful alternative for mutation detection, dramatically improving the speed and the efficiency of DNA testing. Here we tested the performance of the Ion Torrent PGM platform with the Ion AmpliSeq BRCA1 and BRCA2 Panel in our clinical routine of breast/ovarian hereditary cancer syndrome assessment. Methods:We first tested the NGS approach in a cohort of 11 patients (training set) who had previously undergone genetic diagnosis in our laboratory by conventional methods. Then, we applied the optimized pipeline to the consecutive cohort of 136 uncharacterized probands (validation set). Results:By minimal adjustments in the analytical pipeline of Torrent Suite Software we obtained a 100% concordance with Sanger results regarding the identification of single nucleotide alterations, insertions, and deletions with the exception of three large genomic rearrangements (LGRs) contained in the training set. The optimized pipeline applied to the validation set (VS), identified pathogenic and polymorphic variants, including a novel BRCA2 pathogenic variant at exon 3, 100% of which were confirmed by Sanger in their correct zygosity status. To identify LGRs, all negative samples of the VS were subjected to MLPA analysis. Discussion:Our experience strongly supports that the Ion Torrent PGM technology in BRCA1 and BRCA2 germline variant identification, combined with MLPA analysis, is highly sensitive, easy to use, faster, and cheaper than traditional (Sanger sequencing/MLPA) approaches.
| S-EPMC6482939 | biostudies-literature
Project description:When sequencing blood and tumor samples to identify targetable somatic variants for cancer therapy, clinically relevant germline variants may be uncovered. We evaluated the prevalence of deleterious germline variants in cancer susceptibility genes in women with breast cancer referred for neoadjuvant chemotherapy and returned clinically actionable results to patients. Exome sequencing was performed on blood samples from women with invasive breast cancer referred for neoadjuvant chemotherapy. Germline variants within 142 hereditary cancer susceptibility genes were filtered and reviewed for pathogenicity. Return of results was offered to patients with deleterious variants in actionable genes if they were not aware of their result through clinical testing. 124 patients were enrolled (median age 51) with the following subtypes: triple negative (n = 43, 34.7%), HER2+ (n = 37, 29.8%), luminal B (n = 31, 25%), and luminal A (n = 13, 10.5%). Twenty-eight deleterious variants were identified in 26/124 (21.0%) patients in the following genes: ATM (n = 3), BLM (n = 1), BRCA1 (n = 4), BRCA2 (n = 8), CHEK2 (n = 2), FANCA (n = 1), FANCI (n = 1), FANCL (n = 1), FANCM (n = 1), FH (n = 1), MLH3 (n = 1), MUTYH (n = 2), PALB2 (n = 1), and WRN (n = 1). 121/124 (97.6%) patients consented to return of research results. Thirteen (10.5%) had actionable variants, including four that were returned to patients and led to changes in medical management. Deleterious variants in cancer susceptibility genes are highly prevalent in patients with invasive breast cancer referred for neoadjuvant chemotherapy undergoing exome sequencing. Detection of these variants impacts medical management.
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