Heterogeneity and clonal evolution of acquired PARP inhibitor resistance in TP53- and BRCA1-deficient cells.

Homologous recombination (HR)-deficient cancers are sensitive to inhibitors of Poly-ADP Ribose Polymerase (PARPi), which have shown clinical efficacy in the treatment of high-grade serous cancers (HGSC). However, the majority of patients will relapse, and acquired PARPi resistance is emerging as a pressing clinical problem. Here we generated seven single-cell clones with acquired PARPi resistance derived from a PARPi-sensitive, TP53-/- and BRCA1-/- epithelial cell line generated using CRISPR/Cas9. These clones showed diverse resistance mechanisms, and some clones presented with multiple mechanisms of resistance at the same time. Genomic analysis of the clones revealed unique transcriptional and mutational profiles and increased genomic instability in comparison to a PARPi-sensitive cell line. Clonal evolutionary analyses suggested that acquired PARPi resistance arose via clonal selection from an intrinsically unstable and heterogenous cell population in the sensitive cell line, which contained pre-existing drug tolerant cells. Similarly, clonal and spatial heterogeneity in tumor biopsies from a clinical BRCA1-mutant HGSC patient with acquired PARPi resistance were observed. In an imaging-based drug screening, the clones showed heterogenous responses to targeted therapeutic agents, indicating that not all PARPi-resistant clones can be targeted with just one therapy. Furthermore, PARPi-resistant clones showed mechanism-dependent vulnerabilities to the selected agents, demonstrating that a deeper understanding on the mechanisms of resistance could lead to improved targeting and biomarkers for HGSC with acquired PARPi resistance.