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Background Immunotherapy has transformed treatment paradigms in many malignancies but remains largely ineffective in immunologically ‘cold’ solid tumors. One key barrier is the downregulation of major histocompatibility complex class I (MHC I), which limits antigen presentation and restricts CD8+ T cell-mediated immunity. In contrast, natural killer (NK) cells recognize and eliminate tumor cells in an MHC-independent manner, positioning them as a promising modality for targeting MHC-deficient tumors. However, many tumors still resist NK-mediated killing, suggesting the existence of innate immune checkpoints and tumor-intrinsic mechanisms of evasion.Methods To systematically identify regulators of NK cell resistance, we developed and optimized a curated CRISPR library targeting cell surface and immune-related genes. We performed a pan-cancer CRISPR screen across 12 representative cell lines derived from six cancer types (prostate, breast, lung, bladder, kidney, and ovarian), co-cultured with NK92 cells under two effector-to-target (E:T) conditions to model low and high killing pressures. Over 100 individual screens were collected across multiple time points and biological replicates. Hits were ranked based on enrichment or depletion at endpoint relative to time zero and stratified as common or cancer type specific regulators.Results The screen robustly recovered known innate immune regulators, validating the platform. Knockout of genes such as NECTIN2, PTPN2, and MHC class I components (e.g., HLA-B, TAP1, B2M) significantly enhanced NK cell-mediated killing. On the other end of the spectrum, loss of genes including ICAM1, IFNGR1, IFNGR2, JAK1, and JAK2 impaired NK cell cytotoxicity, suggesting their essential roles in maintaining tumor sensitivity to NK attack. Top hits were validated using individual knockout co-culture assays and NK cell degranulation assays. The majority of prioritized hits demonstrated significant phenotypes, highlighting their role as functional innate immune checkpoints. We also identified novel gene candidates with conserved effects across multiple tumor types, as well as lineage-specific regulators. Early in vivo pilot studies suggest that blocking select novel hits enhances NK anti-tumor activity.Conclusions This study establishes a scalable CRISPR screening platform to uncover innate immune resistance pathways across diverse solid tumors. Our results reveal both conserved and tumor-specific mediators of NK cell resistance and nominate a new class of potential therapeutic targets to augment innate immune responses. These findings provide a roadmap for enhancing NK-based immunotherapies through rational checkpoint modulation in solid tumors.