A genome-wide CRISPR screen identifies ZNF251 critical for resistance to PARP inhibitors

The poly (ADP-ribose) polymerases (PARPs) inhibitors are an exciting new class of agents that have shown efficacy in treating various cancers, especially these harboring BRCA1/2 mutations. The cancer associated BRCA1/2 mutations disrupt DNA double strand break (DSB) repair by homologous recombination (HR). PARP inhibitors (PARPi) have been applied to trigger synthetic lethality in BRCA1/2-mutated cancer cells by promoting accumulation of toxic DSBs. Unfortunately, PARP inhibitor (PARPi) resistance is common and develops through multiple mechanisms. Restoration of HR and/or stabilizing replication forks are two major mechanisms of PARPi resistance in BRCA1/2-mutated cells. To further understand the mechanisms of drug resistance to PARPi, we undertook an unbiased approach with a CRISPR-pooled library to screen new genes whose loss-of-function confers resistance to PARPi olaparib. We identified ZNF251, a transcription factor, and confirmed its loss-of-function led to the PARPi resistance in BRCA1-mutated breast and ovarian cancer lines. Elevated activities of both HR and non-homologous end joining (NHEJ) repair were detected in cancer cells harboring BRCA1 mutation and ZNF251 deletion (BRCA1mut+ZNF251del) and were associated with enhanced expression of RAD51 and Ku70/Ku80, respectively. Furthermore, we showed that DNA-PKcs inhibitor restored sensitivity of BRCA1mut+ZNF251del cells to PARPi. Taken together, our study identified a novel gene whose loss of function conferred resistance to PARPi, providing new insight into signaling pathways that contribute to acquired resistance in BRCA1-mutated breast and ovarian cancers. GeCKO CRISPR library was purchased from Addgene (#1000000048), amplified, and packaged as lentivirus based on the instructions on Addgene website. The CRISPR screen was performed as described previously(Hou et al. Cancer research 2017). In brief, MDA-MB-436 cells were transduced with lentivirus carrying GeCKO library, and puromycin selection was performed for 2 days. Then we treated transduced MDA-MB-436 cells with olaparib for 14 days and the survived cells were harvested. The genomic DNA was extracted, and PCR was carried out before deep sequencing of sgRNA sequence in the survived cells genome.