Serotonin receptor 5-HT 2A as a potential target for HCC immunotherapy

While recent clinical trials of combination immunotherapies for hepatocellular carcinoma (HCC) have shown promising clinical efficacy and survival improvements breakthroughs, there is still much room for further improvement. A key limiting factor for HCC immunotherapy is the intrinsic immunosuppression within the liver microenvironment, resulting in suboptimal priming of tumour-specific CD8 cytotoxic T cells and thus immune evasion by the tumour. Hence, identifying new key molecular pathways suppressing T cell responses within the liver is critical for the rational design of more effective combination immunotherapies for HCC. Here, we present evidence that targeting 5-HT2A serotonin receptor signalling could be a viable approach for HCC immunotherapy. Disruption of 5-HT2A signalling using either a selective antagonist small molecule, ketanserin, or by knockout of its coding gene Htr2a, augments the cytotoxic effector phenotype of mouse CD8 T cells activated in vitro with immunosuppressive liver NPCs. Similarly, abrogating 5-HT2A signalling in in vitro activated primary human CD8 T cells with ketanserin treatment or CRISPR-mediated disruption of the HTR2A gene also increased expression of the cytotoxic effector molecules Granzyme B and perforin. RNA-seq analysis of 5-HT2A signalling-deficient activated mouse CD8 T cells revealed increased expression of cytotoxicity-related genes such as Granzyme B and reduced expression of transcription factors downstream of MAP kinase signalling pathways. Consistent with these observations, 5-HT2A activation in CD8 T cells rapidly triggered phosphorylation of the p38, ERK, and JNK/SAP MAP kinases. In vivo, systemic ketanserin treatment significantly prolonged survival of HCC tumour-bearing mice and was non-inferior to alphaPDL1 + alphaVEGFA combination antibody treatment. Combining ketanserin with alphaPDL1 + alphaVEGFA antibodies also significantly prolonged survival relative to control-treated mice while preserving the occurrence of complete tumour regression observed with alphaPDL1 + alphaVEGFA treatment alone. Together, our data describe a role for 5-HT2A as a negative regulator of the cytotoxic effector phenotype in CD8 T cells and highlight the therapeutic potential of targeting 5-HT2A for HCC immunotherapy.