Figure S1: Characterisation of CAPAN1-B2S* Figure S2: Characterisation of SUM149-B1S* Figure S3: Sensitivity of ddPCR assay. Figure S4: Olaparib and talazoparib select for secondary mutant tumour cells. Figure S5: DLD1 tumour cells have a fitness advantage over DLD1.BRCA2-/- cells in vitro. Figure S6: Olaparib has little efficacy in mixed CAPAN-1 xenografts. Figure S7: Exome sequencing of CAPAN-1.B2.S* shows retention of TP53 mutations. Figure S8: Exome sequencing of SUM149.B1.S* show retention of TP53 mutation. Figure S9: BRCA-proficient and -deficient cells exhibit sensitivity to additional DNA damaging agents. Figure S10: AZD-1775 causes an active S phase reduction in both CAPAN1 and CAPAN-1.B2.S* cells.
ARTICLE ABSTRACT
Although PARP inhibitors target BRCA1- or BRCA2-mutant tumor cells, drug resistance is a problem. PARP inhibitor resistance is sometimes associated with the presence of secondary or “revertant” mutations in BRCA1 or BRCA2. Whether secondary mutant tumor cells are selected for in a Darwinian fashion by treatment is unclear. Furthermore, how PARP inhibitor resistance might be therapeutically targeted is also poorly understood. Using CRISPR mutagenesis, we generated isogenic tumor cell models with secondary BRCA1 or BRCA2 mutations. Using these in heterogeneous in vitro culture or in vivo xenograft experiments in which the clonal composition of tumor cell populations in response to therapy was monitored, we established that PARP inhibitor or platinum salt exposure selects for secondary mutant clones in a Darwinian fashion, with the periodicity of PARP inhibitor administration and the pretreatment frequency of secondary mutant tumor cells influencing the eventual clonal composition of the tumor cell population. In xenograft studies, the presence of secondary mutant cells in tumors impaired the therapeutic effect of a clinical PARP inhibitor. However, we found that both PARP inhibitor–sensitive and PARP inhibitor–resistant BRCA2 mutant tumor cells were sensitive to AZD-1775, a WEE1 kinase inhibitor. In mice carrying heterogeneous tumors, AZD-1775 delivered a greater therapeutic benefit than olaparib treatment. This suggests that despite the restoration of some BRCA1 or BRCA2 gene function in “revertant” tumor cells, vulnerabilities still exist that could be therapeutically exploited. Mol Cancer Ther; 16(9); 2022–34. ©2017 AACR.
