Genome-wide CRISPR screening reveals tumor-intrinsic immune evasion mechanism in ovarian cancer

Ovarian cancer is the most lethal gynecologic malignancy. Previous studies demonstrated high levels of tumor-infiltrating T cells in ovarian cancer, which predict benefit from immunotherapy. However, clinical trials such as KEYNOTE-100 report only ~10% overall response to pembrolizumab. This discrepancy indicates that ovarian cancers harbor heterogeneous, tumor-intrinsic mechanisms of immune evasion.To distinguish immune-resistant cancer cell lines, we established a reporter-based co-culture platform in which pre-activated human T cells were directly co-cultured with ovarian cancer cell lines. We profiled 24 ovarian cancer cell lines engineered with a real-time cytotoxicity reporter to quantify tumor-cell death. Pre-activated T cells from two healthy donors were co-cultured with each cancer cell line, and reporter readouts stratified the set into 11 immune-sensitive and 6 immune-resistant lines. To uncover genetic determinants of resistance, we selected the two most resistant cell lines for whole-genome CRISPR screening. Following co-culture of CRISPR-edited cancer cells with pre-activated T cells, sgRNA abundances were sequenced and analyzed using the MAGeCK-MLE pipeline. Focusing on concordant depletions, we identified 56 genes (P < 0.05; beta < -0.5) whose loss augments T-cell-mediated tumor killing in both cell lines. Among these, we prioritized ITGA8 (integrin alpha8) as a promising surface target. ITGA8 heterodimerizes with beta1 to form alpha8beta1, which engages extracellular-matrix ligands and activates adhesion- and survival-linked signaling axes, including FAK/Src. Given that integrins regulate immune-cell infiltration into tumors and pathways such as TGF-beta signaling, we hypothesize that ITGA8 sustains pro-survival signaling that constrains T-cell activity. Accordingly, pharmacologic or biologic blockade of ITGA8 may offer a surface-accessible strategy to convert immune-resistant ovarian tumors into immune-sensitive disease.