Background: Breast cancer remains as one of the most lethal types of cancer in women. Among various subtypes, triple-negative breast cancer (TNBC) is the most aggressive and hard to treat type of breast cancer. Mechanistically, increased DNA repair and cell cycle checkpoint activation remain as the foremost reasons behind TNBC tumor resistance to chemotherapy and disease recurrence.
Methods: We evaluated the mechanism of prexasertib-induced regulation of homologous recombination (HR) proteins using 20S proteasome inhibitors and RT-PCR. HR efficiency and DNA damages were evaluated using Dr-GFP and comet assays. DNA morphology and DNA repair focus studies were analyzed using immunofluorescence. UALCAN portal was used to evaluate the expression of RAD51 and survival probability based on tumor stage, subtype, and race in breast cancer patients.
Results: Our results show that prexasertib treatment promotes both post-translational and transcriptional mediated regulation of BRCA1 and RAD51 proteins. Additionally, prexasertib-treated TNBC cells revealed over 55% reduction in HR efficiency compared to control cells. Based on these results, we hypothesized that prexasertib treatment induced homologous recombination deficiency (HRD) and thus should synergize with PARP inhibitors (PARPi) in TNBC cells. As predicted, combined treatment of prexasertib and PARPi olaparib increased DNA strand breaks, γH2AX foci, and nuclear disintegration relative to single-agent treatment. Further, the prexasertib and olaparib combination was synergistic in multiple TNBC cell lines, as indicated by combination index (CI) values. Analysis of TCGA data revealed elevated RAD51 expression in breast tumors compared to normal breast tissues, especially in TNBC subtype. Interestingly, there was a discrepancy in RAD51 expression in racial groups, with African-American and Asian breast cancer patients showing elevated RAD51 expression compared to Caucasian breast cancer patients. Consistent with these observations, African-American and Asian TNBC patients show decreased survival.
Conclusions: Based on these data, RAD51 could be a biomarker for aggressive TNBC and for racial disparity in breast cancer. As positive correlation exists between RAD51 and CHEK1 expression in breast cancer, the in vitro preclinical data presented here provides additional mechanistic insights for further evaluation of the rational combination of prexasertib and olaparib for improved outcomes and reduced racial disparity in TNBC.
Keywords: CHK1 and homologous recombination; Olaparib; Prexasertib; RAD51; TNBC.