BIOLOGICAL SIGNIFICANCE OF ALTERATIONS IN BRCA1 AND BRCA2 GENES AND RESPONSE TO DNA DAMAGE AGENTS IN HEREDITARY BREAST CANCER

Background: Although the large number of studies investigating BRCA mutations and their clinical role in different populations and ethnicities, there is a lack of a systematic analysis on these alterations in Italian cohorts, including the analysis of Variants of Unknown biological and clinical Significance (VUS). Moreover, correct management of breast cancer patients tested positive for alterations in BRCA1 or BRCA2 genes is still controversial. We aimed to assess the biological and clinical relevance of BRCA alterations in a consecutive cohort of hereditary breast cancer patients, with particular attention to VUS. Methods: Genetic and clinical data from 366 patients with familial history of breast cancer were reviewed. The association between clinical-pathological, molecular data, and breast cancer patient subgroups was assessed. BRCA1/2 and γ-H2AX expression levels were assessed by qRT-PCR and IHC for all tumors. In silico protein prediction models were computed for VUS with potential clinical significance. Cell proliferation and apoptosis assays for CRISPR/Ca9s-generated mutant MDA-MB-231 cell line were performed to evaluate the sensitivity of specific VUS to DNA damage agents. Results: Overall, 73 breast cancer patients (20%) tested positive for BRCA1/2 alterations. BRCA1 and BRCA2 mutations were reported in 34 (46.5%) and 15 (20.5%) patients, respectively. Two patients (3%) showed two concurrent mutations in both genes. Twenty-two patients (30%) tested positive for VUS. Breast cancer family history and early onset of disease were significantly associated with BRCA1 (p < 0.001) and BRCA2 (p = 0.045 and p = 0.005) mutations. Triple-negative histotype, grading 3, and high Ki-67 levels were significantly associated with BRCA1 mutations (p < 0.001). Molecular, in silico and in vitro experiments confirmed the deleterious effect of BRCA1 c.5509T>C VUS, which was associated with significant high levels of DNA damage and greater sensitivity to Olaparib compared to Cisplatin treatment. Conclusions: Our study supports the deleterious effect of the BRCA1 c.5509T>C VUS in hereditary breast cancer patients, and suggests that breast cancer patient carriers of this variant could benefit from an intense surveillance and from a single agent treatment with Olaparib avoiding various side effects of chemotherapy treatment.