Isoxazole-based molecules restore NK cell immune surveillance in hepatocarcinogenesis by targeting TM4SF5 and SLAMF7 linkage

Dynamic communication between hepatocytes and the environment is critical in hepatocellular carcinoma (HCC) development. Clinical immunotherapy against HCC is currently unsatisfactory and needs more systemic considerations, including the identification of new biomarkers and immune checkpoints. Transmembrane 4 L six family member 5 (TM4SF5) is known to promote HCC, but it remains unclear how cancerous hepatocytes avoid immune surveillance and whether avoidance can be blocked. We investigated how TM4SF5-mediated hepatic tumorigenesis avoids surveillance by natural killer (NK) cells, which are prevalent in the liver, and whether the avoidance can be blocked by anti-TM4SF5 agents. We used comprehensive structure activity relationship analysis to identify TM4SF5-specific isoxazole (TSI)-based small molecules that inhibit TM4SF5-mediated effects. TM4SF5 expressed by hepatocytes reduced NK cell cytotoxicity by downregulating stimulatory ligands/receptors, including signaling lymphocytic activation molecule family member 7 (SLAMF7). TM4SF5 bound SLAMF7 depending on N-glycosylation and caused intracellular trafficking of SLAMF7 from the plasma membrane to lysosomes for degradation. TSI treatments in cell lines and animal models of HCC blocked this binding, intracellular trafficking, and downregulation, resulting in higher levels of stimulatory NK cell ligands. In mouse xenograft models, TSI treatment abrogated HCC development by increasing the abundance and dispersion of Slamf7-positive cells in liver tissues, recapitulating the phenotype of Tm4sf5-knockout mice and indicating TSI-mediated restoration of NK cell surveillance. These findings suggest that TSIs can inhibit TM4SF5-mediated liver carcinogenesis by increasing NK cell surveillance.