Purpose: Impaired MHCI-presentation and insensitivity to interaction. Around 40% of melanomas constitutively express immune effector molecules are common features of immune MHC Class II molecules; hence, melanomas with or without checkpoint blockade (ICB)-resistant tumors and can be, respecnatural constitutive MHC Class II expression (MHCII^constþ or tively, associated with loss of b₂ microglobulin (B2M) or impaired MHCII^const-) were used. IFNg signaling. Patients with ICB-resistant tumors can respond to Results: CD4^þ TIL-mediated cytotoxicity was not affected by alternative immunotherapies, such as infusion of autologous B2M loss but was dependent on the expression of CIITA. tumor-infiltrating lymphocytes (TIL). CD4^þ T cells can exert MHCII^constþ melanomas were killed by tumor-specific CD4^þ TILs cytotoxic functions against tumor cells; however, it is unclear even in the absence of IFNg-mediated MHCII upregulation, where-whether CD4^þ T-cell responses can be exploited to improve the as IFNg was necessary for CD4^þ TIL-mediated cytotoxicity against clinical outcomes of patients affected by ICB-resistant tumors. MHCII^const- melanomas. Notably, although tumor-specific CD4^þ Experimental Design: Here, we exploited CRISPR (clustered TILs did not kill JAK1^KO MHCII^const- melanomas even after IFNg regularly interspaced short palindromic repeats)/Cas9 gene edit-stimulation, sensitivity to CD4^þ TIL-mediated cytotoxicity was ing to reproduce immune-resistant tumor phenotypes via gene maintained by JAK1^KO MHCII^constþ melanomas. knockout (^KO). To determine the role of cytotoxic CD4^þ TILs Conclusions: In conclusion, our data indicate that exploiting in ICB-resistant tumors, we investigated CD4^þ TIL-mediated tumor-specific cytotoxic CD4^þ TILs could help overcome resistance cytotoxicity in matched pairs of TILs and autologous melanoma to ICB mediated by IFNg-signaling loss in MHCII^constþ melanomas. cell lines, used as a model of patient-specific immune-tumor See related commentary by Betof Warner and Luke, p. 3829