TerminateNB Cell Line Panel CancerPAM

Efficient tumor-specific gene editing remains a major challenge in CRISPR-based therapies. Here, we introduce CancerPAM, a multi-omics pipeline that identifies and ranks tumor-specific CRISPR knock-in sites with high specificity and efficiency. We applied CancerPAM to sequencing data from cell lines and patients with neuroblastoma, a pediatric solid tumor characterized by an immunosuppressive tumor microenvironment (TME) that limits CAR T cell infiltration. Using CRISPR-mediated integration of pro-inflammatory cytokine transgenes (CXCL10, CXCL11, IFNG) at tumor-specific loci, we successfully remodeled the TME, enhancing immune infiltration and CAR T cell function. In vitro and in vivo studies demonstrated improved CAR T cell trafficking and tumor control in CXCL10-expressing tumors. Our findings establish CancerPAM as a versatile tool for precise CRISPR-based genome engineering and highlight tumor-intrinsic cytokine expression as a promising strategy to augment immunotherapies in solid tumors. Further advances in CRISPR delivery and knock-in efficiency will be crucial for clinical translation of this approach.