@article {Lambing2021.10.16.464638, author = {Silke Lambing and Stefan Holdenrieder and Patrick M{\"u}ller and Christian Hagen and Stephan Garbe and Martin Schlee and Jasper G. van den Boorn and Eva Bartok and Gunther Hartmann and Marcel Renn}, title = {Ionizing radiation improves RIG-I mediated immunotherapy through enhanced p53 activation in malignant melanoma}, elocation-id = {2021.10.16.464638}, year = {2021}, doi = {10.1101/2021.10.16.464638}, publisher = {Cold Spring Harbor Laboratory}, abstract = {The activation of the innate immune receptor RIG-I is a promising approach in immunooncology and currently under investigation in clinical trials. RIG-I agonists elicit a strong immune activation in both tumor and immune cells and induce both direct and indirect immune cell-mediated tumor cell death which involves tumor-specific cytotoxic T-cell response and type I interferon-driven innate cytotoxic immunity. Besides RIG-I, irradiation is known to induce cytotoxic DNA damage resulting in tumor debulking followed by the induction of tumor-specific immunity. To date, it is unclear whether the molecular antitumor effects of RIG-I and irradiation are additive or even synergize. Here, we investigated the combination of RIG-I activation with radiotherapy in melanoma. We found that low dose x-ray irradiation enhanced the extent and immunogenicity of RIG-I mediated tumor cell death in human and murine melanoma cell lines and in the murine B16 melanoma model in vivo. Pathway analysis of transcriptomic data revealed a central role for p53 downstream of the combined treatment, which was corroborated using p53-/- B16 cells. In vivo, the additional effect of irradiation on immune cell activation and inhibition of tumor growth was lost in mice carrying p53-knockout B16 tumors, while the response to RIG-I stimulation in those mice was maintained. Thus, our results identify p53 as pivotal for the synergy of RIG-I with irradiation, resulting in potent induction of immunogenic tumor cell death. Consequently, low dose radiotherapy holds great promise to further improve the efficacy or RIG-I ligands especially in patients with malignant melanoma or other tumors exhibiting a functional p53 pathway.Competing Interest StatementM.S and G.H. are inventors on a patent covering synthetic RIG-I ligand. MR and GH were co-founders of Rigontec GmbH.}, URL = {https://www.biorxiv.org/content/early/2021/10/16/2021.10.16.464638}, eprint = {https://www.biorxiv.org/content/early/2021/10/16/2021.10.16.464638.full.pdf}, journal = {bioRxiv} }