Abstract 5518: Single cell RNA sequencing revealed that the mechanism of the synergistic efficacy of STA551 and immune checkpoint inhibitor results from the induction of highly activated T cells in tumor
Authors: Uchikawa Ryo , Takakuwa Kazuya , Mikami Hirofumi , Metsugi Shoichi , Taniguchi Kenji , Narita Yoshinori , Kamata-Sakurai Mika 
Published: June 2022

Abstract
Background: CD137 (4-1BB) is a co-stimulatory molecule which exerts activation signals in multiple immune cell subsets. However, agonist antibodies targeting CD137 have not been clinically successful due to severe systemic toxicity. Therefore, we generated STA551, a tumor selective CD137 agonist antibody. STA551 binds to CD137 only in the presence of ATP, which is known to be present at high levels in tumor tissue, and activates immune cells such as T cells. Previous studies have shown that STA551 exhibits antitumor efficacy without inducing a systemic immune response in a murine model. Combination studies with anti-PD-L1 antibody have also shown synergistic efficacy without inducing a systemic immune response. However, it was unclear why they exhibit strong combination effects. In this study, we elucidated the mechanism of STA551 and anti-PD-L1 antibody efficacy through comprehensive analysis of gene expression using single cell RNA sequencing.
Method & Result: In a colon 38-bearing human CD137 knock-in mouse model, combination therapy with STA551 and anti-PD-L1 antibody showed potent anti-tumor efficacy. From immunohistochemistry of tumor tissue, increased numbers of CD8+ T cells and elevated expression of PD-L1 molecules were observed. To perform a detailed mechanistic analysis, tumor tissue was collected after antibody administration and CD3 positive and CD3 negative fractions were separated and gene expression was comprehensively analyzed using single cell RNA-seq. Next, the fraction expressing Cd8a gene was extracted from CD3 positive fraction and clustered. One cluster was found to be markedly increased in STA551 and anti-PD-L1 antibody combination compared to vehicle or monotherapy treatment. Differential gene expression analysis revealed that the cluster was enriched for genes related to cytokines and chemokines. This result revealed that the combination of STA551 and anti-PD-L1 antibody induced highly activated CD8+T cells.
Conclusion: The combination of STA551 and PD-L1 showed synergistic anti-tumor efficacy in a colon38-bearing human CD137 knock-in mouse model. Detailed mechanistic analysis showed that the combination of STA551 and PD-L1 induced highly activated CD8+T cells expressing chemokines and cytokines compared to monotherapy. This cluster also express chemokines, suggesting that other immune cells have been recruited. These results suggest that even the activation of only tumor tissue, which is a characteristic of STA551, can induce a strong immune response.
Citation Format: Ryo Uchikawa, Kazuya Takakuwa, Hirofumi Mikami, Shoichi Metsugi, Kenji Taniguchi, Yoshinori Narita, Mika Kamata-Sakurai. Single cell RNA sequencing revealed that the mechanism of the synergistic efficacy of STA551 and immune checkpoint inhibitor results from the induction of highly activated T cells in tumor [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 5518.