Purpose: Cancer immunotherapy has shown promising clinical outcomes in many patients. However, some patients still fail to respond, and new strategies are needed to overcome resistance. The purpose of this study was to identify novel genes and understand the mechanisms that confer resistance to cancer immunotherapy. Experimental Design: To identify genes mediating resistance to T-cell killing, we performed an open reading frame (ORF) screen of a kinome library to study whether overexpression of a gene in patient-derived melanoma cells could inhibit their susceptibility to killing by autologous tumor-infiltrating lymphocytes (TIL). Results: The RNA-binding protein MEX3B was identified as a top candidate that decreased the susceptibility of melanoma cells to killing by TILs. Further analyses of anti–PD-1–treated melanoma patient tumor samples suggested that higher MEX3B expression is associated with resistance to PD-1 blockade. In addition, significantly decreased levels of IFNg were secreted from TILs incubated with MEX3B-overexpressing tumor cells. Interestingly, this phenotype was rescued upon overexpression of exogenous HLA-A2. Consistent with this, we observed decreased HLA-A expression in MEX3B-overexpressing tumor cells. Finally, luciferase reporter assays and RNA-binding protein immunoprecipitation assays suggest that this is due to MEX3B binding to the 3 0 untranslated region (UTR) of HLA-A to destabilize the mRNA. Conclusions: MEX3B mediates resistance to cancer immunotherapy by binding to the 3 0 UTR of HLA-A to destabilize the HLA-A mRNA and thus downregulate HLA-A expression on the surface of tumor cells, thereby making the tumor cells unable to be recognized and killed by T cells.