Pyrosequencing Assay for BRCA1 Methylation Analysis Results from a Cross-Validation Study

Epithelial ovarian cancers (EOCs) harboring germline or somatic pathogenic variants in BRCA1 and BRCA2 genes show sensitivity to poly(ADP-ribose) polymerase inhibition. It has been suggested that BRCA1 promoter methylation is perhaps a better determinant of therapy response, because of its intrinsic dynamic feature, with respect to genomic scars or gene mutation. Conflicting evidence was reported so far, and the lack of a validated assay to measure promoter methylation was considered a main confounding factor in data interpretation. To contribute to the validation process of a pyrose-quencing assay for BRCA1 promoter methylation, 109 EOCs from two Italian centers were reciprocally blindly investigated. By comparing two different pyrosequencing assays, addressing a partially over-lapping region of BRCA1 promoter, an almost complete concordance of results was obtained. Moreover, the clinical relevance of this approach was also supported by the finding of BRCA1 transcript down -regulation in BRCA1-methylated EOCs. These findings could lead to the development of a simple and cheap pyrosequencing assay for diagnostics, easily applicable to formalin-fixed, paraffin-embedded tissues. This technique may be implemented in routine clinical practice in the near future to identify EOCs sensitive to poly(ADP-ribose) polymerase inhibitor therapy, thus increasing the subset of women affected by EOCs who could benefit from such treatment.