Lionel Ouonkap

The immune system plays a critical role in the detection and elimination of cells with tumorigenic potential. Central to this immunosurveillance are natural killer (NK) cells, a type of innate lymphoid cell responsive to signals of cell stress, infection, and cellular transformation. NK cells, like CD8+ T cells, are a type of cytotoxic lymphocyte (CTL) that detects and kills targeted cells through the release of cytotoxins (e.g., perforins, granzymes) or through the induction of apoptosis. This study aimed to identify novel cancer-expressed ligands that activate and inhibit NK cells using in vitro CRISPR activation and knockout screens. We utilized pooled gain and loss-of-function screening to overexpress or delete surface proteins in cancer cell lines co-cultured with human NK cells with the aim of determining ligands that drive or inhibit NK cell cytotoxicity. These hits have then been validated using overexpression studies in the K562 screening cell lines. The next step will be the identification of tumor cell lines known to be resistant or sensitive to NK cell cytotoxicity, knock out genes of interest (GOI), and use in vitro and in vivo assays to determine functional effects on NK cell cytotoxicity and tumor clearance. Using this technique, we were able to validate the overexpression of already known critical genes for cancer survival such as MUC1 and MUC4 genes. Validated results of these screens will be used to suggest new targets for checkpoint inhibition. If successful, this study will identify potential targets for novel therapeutics, allow for the engineering of dysregulation-resistant CAR NK cells, as well as allow clinicians to identify when using or targeting NK instead of T cells (or vice versa) might be a more optimal strategy in patient care.