Abstract
Germline pathogenic mutations in BRCA1 increase risk of developing breast cancer. Screening for mutations in BRCA1 frequently identifies sequence variants of unknown pathogenicity and recent work has aimed to develop methods for determining pathogenicity. We previously observed that tumor DNA methylation can differentiate BRCA1-mutated from BRCA1-wild type tumors. We hypothesized that we could predict pathogenicity of variants based on DNA methylation profiles of tumors that had arisen in carriers of unclassified variants. We selected 150 FFPE breast tumor DNA samples [47 BRCA1 pathogenic mutation carriers, 65 BRCAx (BRCA1-wild type), 38 BRCA1 test variants] and analyzed a subset (n=54) using the Illumina 450K methylation platform, using the remaining samples for bisulphite pyrosequencing validation. Three validated markers (BACH2, C8orf31, and LOC654342) were combined with sequence bioinformatics in a model to predict pathogenicity of 27 variants (independent test set). Predictions were compared with standard multifactorial likelihood analysis. Prediction was consistent for c.5194-12G>A (IVS 19-12 G>A) (P>0.99); 13 variants were considered not pathogenic or likely not pathogenic using both approaches. We conclude that tumor DNA methylation data alone has potential to be used in prediction of BRCA1 variant pathogenicity but is not independent of estrogen receptor status and grade, which are used in current multifactorial models to predict pathogenicity.
Highlights
Breast cancer remains the most diagnosed cancer in women, with an overall incidence in the UK of 1 in 8
We have previously shown that breast tumors from patients median methylation in BRCA1 tumors for five candidate regions with pathogenic germline BRCA1 mutations have distinct DNA [CD9 (OMIM# 143030), SGK1 (OMIM# 602958), ERCC3 methylation profiles compared to familial breast cancer cases (OMIM# 133510), and 2 regions in FGF2 (OMIM# 134920), with no BRCA1 or BRCA2 mutations (BRCAx).[13]
Multiple studies have identified distinct epigenetic profiles not understood, and it is unknown whether these changes in that correlate with different breast cancer subtypes and/or estrogen receptor (ER) methylation occur at ER target genes or as a consequence of www.tandfonline.com
Summary
Breast cancer remains the most diagnosed cancer in women, with an overall incidence in the UK of 1 in 8 (http://www.cancerresearchuk.org/cancer-info/cancerstats/). These probabilities were converted to LRs and com- line mutations compared to BRCAx, and found that the bined by multiplication The test set samples from variant carriers clustered with the BRCA1 pathogenic or BRCAx subgroups Of these 18 loci, four sites were validated by pyrosequencing in an independent group of samples, but logistic regression suggested that for all but one probe, the association was not truly independent from ER status and grade in the independent samples. This work suggests that the methylation markers will have value for future variant classification for BRCA1 and potentially for other genes with known tumor methylation phenotypes, such as MLH1 (OMIM# 120436) in colon cancer (Cancer Genome Atlas Network, 2012b)
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