Genome-Wide CRISPR Screening Identifies SLC47A1 (MATE1) As a Gene that Confers Resistance to Prexasertib, a CHK1 Inhibitor, in Ovarian Cancer Cells

Authors

• P MONDAL
• D CHEN
• AV MITCHELL
• L JENKINS
• MM GOTTESMAN

Abstract

High-grade serous ovarian cancer (HGSOC) is the most lethal gynecologic malignancy in the United States, with limited treatment options. HGSOC often harbors p53 mutations, leading to DNA repair defects and genomic instability by disrupting the G1/S checkpoint. Transformed cells then rely heavily on the G2/M checkpoint to repair damaged DNA. The ATR-CHK1-WEE1 pathway is crucial for this process, with CHK1 phosphorylating and deactivating CDC25C phosphatases, triggering G2/M checkpoint activation. Prexasertib, a selective CHK1 inhibitor, emerged as a promising treatment for BRCA wild-type HGSOC, but resistance to monotherapy has quickly developed. In a phase 2 trial, only half of the heavily pretreated patients saw clinical benefits, highlighting the need to understand resistance mechanisms. Using whole genome CRISPR-cas9 screening, we identified that upregulation of Solute Carrier Family 47 Member 1 (SLC47A1) increased resistance to prexasertib. CRISPR activation also identified ABCB1 and ABCG2, multidrug efflux pumps, as top hits, suggesting drug efflux as a key driver of resistance. Multiple cell lines with overexpression or knockdown of SLC47A1 were used to study prexasertib resistance. We discovered CHK1 overexpression, the primary target of prexasertib, as a resistance mechanism. Other components in the CHK1 pathway (CDC25A, CDK2, CCNE1/2) were also involved in determining cell sensitivity or resistance. Overexpression of ABCB1, ABCG2, or SLC47A1 conferred resistance, while SLC47A1 depletion increased sensitivity to prexasertib. MALDI-mass spectrophotometry revealed reduced prexasertib accumulation in cells with SLC47A1 overexpression. These findings offer new insights into resistance mechanisms and could guide future efforts to improve prexasertib efficacy in patients.

Scientific Focus Area: Cancer Biology

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