Abstract 5515: Combination of recombinant murine IL-12 and anti-PD-1 induced synergistic anti-tumor response
Authors: Tsou Peiyi , Lin Wan-Ying , Lee Meng-Na , Chen San-Chi 
Published: June 2022

Abstract
Introduction: Immune-checkpoint inhibitors have become the standard therapy in many types of cancers. The combination of anti-PD-1 and anti-CTLA4 demonstrated better anti-tumor response, but toxicity is a concern. Cytokines including IL-2, IL-12, IL-15, IL-21 that target immune cells have been developed to enhance immune response against tumors. Clinical trials for the combination of cytokines and anti-PD-1 have been proposed (Mohamad et al, 2020). IL-12, a multifunctional cytokine, regulates both innate and adaptive immunity. The role of IL-12 in regulating immune response showed that intra-tumoral electroporation of plasmid IL-12 prime responses to anti-PD-1 treatment (Algazi et al, 2016). IL-12 produced by dendritic cells accompanied with IFN-γ play an important role in anti-tumor response induced by anti-PD-1 (Garris et al, 2018). Therefore, this study aims to explore the synergistic anti-tumor immune response of recombinant murine IL-12 (rmIL-12) and anti-PD-1.
Methods: Different doses of rmIL-12 alone or combined with anti-PD-1 were administered in different tumor models. CT26, MC38, Hepa 1-6 that generate hot tumor were used. Tumors were inoculated subcutaneously in syngeneic mouse tumor models and treated when mean tumor volume reach 60-100 mm3. Tumor resection and splenectomy were performed on tumor inoculated mice at termination. Immunocytochemistry stain was used to detect immune cells including CD4, CD8 T cells, regulatory T cells (Treg), myeloid-derived suppressor cells (MDSC) and macrophage. Serum cytokines were checked for immune response and liver enzyme for toxicity.
Results: The combination of rmIL-12 and anti-PD-1 (BIW x 3 weeks) showed better tumor control compared to anti-PD-1 alone with CT26, MC38, Hepa 1-6 cell lines. Synergistic effect was observed even with a low dose of rmIL-12 (20 ng/mice). A higher dose of rmIL-12 achieved better tumor control. In the CT26 tumor model, combination therapy suppressed tumor growth up to 73% compared to anti-PD-1 alone. These results demonstrated a clear dose-response relationship. The combination of rmIL-12 and anti-PD-1 induced higher level of IFN-γ compared to anti-PD-1 or rmIL-12 alone. A higher dose of rmIL-12 resulted in higher IFN-γ level. The IFN-γ level peaked about 12 hours after the treatment and returned near baseline in about 48 hours. The combination of rmIL-12 and anti-PD-1 showed higher percentage of CD8 T cells in tumors compared to anti-PD-1 or rmIL-12 alone. Combination group significantly decreased Treg in the tumor microenvironment, but not MDSC. Combination therapy also reprogrammed the macrophages from M2 towards M1 polarization in the tumor. Finally, no significant elevation of AST/ALT level was observed among all the groups.
Conclusion: The combination of rmIL-12 and anti-PD-1 demonstrated a synergistic immune response for tumor control. The result needs to be further confirmed in human in the near future.
Citation Format: Peiyi Tsou, Wan-Ying Lin, Meng-Na Lee, San-Chi Chen. Combination of recombinant murine IL-12 and anti-PD-1 induced synergistic anti-tumor response [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 5515.