Tumor cell-intrinsic PD-L1 signals promote DNA damage responses that mediate resistance to Chk1 and PARP inhibitors in vivo

Abstract Tumor PD-L1 mediates diverse cell-intrinsic signals that increase cancer pathogenesis including the DNA damage response (DDR). Although immune consequences of tumor-extrinsic PD-L1 blockade are known, novel therapeutic vulnerabilities from inhibiting tumor-intrinsic PD-L1 signals are little studied. To assess tumor-intrinsic PD-L1 DDR control, we depleted PD-L1 by CRISPR/Cas9 or shRNA (PD-L1KO) in mouse B16 (melanoma), 4T1 (breast), ID8agg (ovarian), and human bladder RT4 cancer lines. We identified specific DDR signaling defects in PD-L1KO versus control (con) cells after exposure to distinct DNA damaging agents. 24h gemcitabine (or 2 Gy X-ray) in vitro in PD-L1KO vs. con RT4 cells elevated γH2AX (DNA damage marker) and reduced Chk2 DDR gene expression but not ATM, ATR, Chk1, or BRCA1 (immunoblots), and decreased BRCA1/p-RPA32 nuclear foci formation (confocal) altogether suggesting tumor-intrinsic PD-L1 promotes specific DDR/DNA repair pathways. PD-L1 loss could thus predict synthetic lethality to DDR inhibitors (DDRi) Chk1i or PARPi. PD-L1KORT4, ID8, and 4T1 cells were more sensitive to small molecule Chk1i in vitro (>15 fold vs. con, MTT) and in vivo in NSG mice. Chk1i and PARPi led to γH2AX increase in PD-L1KO B16, RT4, and 4T1 vs. sham treated tumor cells as predicted by DDR defects. Genetic cellular PD-L1 depletion, but not anti-PD-L1 (extrinsic), strongly sensitized B16 and 4T1 tumors to PARPi in vivo. Strikingly, PARPi had no effect on PD-L1KO B16 in RAG2KO mice despite treating WT mice, indicating a strong immune component to DDRi treatment efficacy. Thus, DDRi plus inhibiting tumor intrinsic PD-L1 signals could improve immunotherapy in immunotherapy-sensitive or resistant tumors.