A combination of PG3 and PARP inhibitors exhibits antitumor effects in both BRCA1-mutated and BRCA1-wild-type TNBC

DNA double-strand break (DSB) repair can be mediated by non-homologous end joining (NHEJ) and homologous recombination (HR). Homologous recombination repair (HRR) is important because it accurately repairs DNA DSBs using a sister chromatid as a template, which is crucial for maintaining genome stability and preventing mutations. Deficiency in HRR makes cancer cells sensitive to DNA damage drugs. BRCA1/2 and RAD51 proteins play very important roles in HRR. PARP is a key enzyme in the repair of DNA single-strand breaks (SSB). Unrepaired SSBs will convert to DSBs in cells. Therefore, BRCA1/2 -mutated cancer cells are sensitive to PARP inhibitors. However, PARP inhibitor resistance develops quickly, mainly through mechanisms that restore homologous recombination repair. This can happen through secondary mutations that restore function in genes of BRCA1 or BRCA2, and overexpression of RAD51. RAD51 overexpression is reported in many cancers, such as breast, prostate, and glioblastoma, and has been involved in chemotherapy resistance. Inhibition of RAD51 creates an HR-deficient status, which can sensitize cancer cells to PARP inhibitor treatments. It has been reported that ISR (integrated stress response) leads to downregulation of RAD51. We reported before that PG3 treatment induced potent ISR. Hence, we hypothesized that PG3 can sensitize PRAP-resistant tumor cells (both BRCA1-mutant and wild-type) to PARP inhibitors by downregulating RAD51. The combination treatments of PG3 and Olaparib/Talazoparib showed synergistic inhibitory effects on triple-negative breast cancer cells, BRCA1-mutated SUM149 and MD-MB-436, and BRCA1-wildtype MD-MB-231 and MD-MB-468. Western blots showed that two BRCA1-mutated cell lines, SUM149 and MD-MB436, express very low levels of BRCA1 protein compared to wild-type cell lines MD-MB231and MD-MB468. On the other hand, SUM149 and MD-MB436 show much higher RAD51 expression than MD-MB231 and MD-MB468. PG3 downregulates RAD51 in both SUM149 and MD-MB231 cells, but not in MD-MB436 and MD-MB468 cells. Transcriptional factors c-Myc, E2F1, and FoxM1 regulate RAD51 gene expression. We found that PG3 induced downregulation of c-Myc, E2F1, and FoxM1 in both SUM149 and MB231 cells, but not in MB436 and MB468 cells. That is consistent with previous publications. PG3 also downregulates wild-type BRCA1 in MD-MB231 cells but not in MD-MB468 cells. We found that olaparib treatment induced upregulation of RAD51 in SUM149 cells and upregulation of both BRCA1 and RAD51 in MD-MB231 cells. PG3 blocked the olaparib-induced upregulation of RAD51 in SUM149cells, and the upregulation of BRCA1 and RAD51 in MD-MB231 cells. The combined treatment induced more DNA damage than olaparib or PG3 alone in SUM149 and MB231 cells, as indicated by increased gammaH2AX level.