Abstract 6644: A comprehensive in vitro and in vivo system to evaluate STING agonist efficacy in cancer therapeutics

Abstract

A number of stimulator of interferon genes protein (STING) agonists has recently been developed to treat cancer and infectious diseases. To investigate the binding of ligand to STING and subsequent activation of the STING pathway, we used the protein thermal shift assay, a high-throughput screening method for intermolecular interaction involving a protein of interest, to identify previously unknown compounds that bind to STING. To determine the functional consequence of ligand binding, we constructed IFN-β-Luc THP1 reporter cell line, to evaluate the pathway activation together with IFNβ secretion. In addition, we incubated human peripheral blood mononuclear cells (PBMCs) with STING agonist and measured phosphorylation of STING, phosphorylation of IRF3 and the secretion of cytokines such as IFN-β by ELISA or Luminex. To determine the in vivo therapeutic effects of STING agonists, we performed efficacy studies in syngeneic mouse models of melanoma (B16F10 cell line), colorectal cancer (CT-26 cell line) or breast cancer (4T1 cell line) with intratumoral delivery of test compounds. Test Treatment with STING agonists resulted in significant tumor growth inhibition as measured by tumor volume and significantly improved survival. Tumor-reactive IFN-γ secreting CD8+ T cells were harvested from spleens, purified by immunomagnetic sorting, and immediately used in ELISPOT assays using the mouse IFN-γ ELISPOT Kit. Tumor-infiltrating lymphocytes (TILs) have also been assessed in harvested melanoma (B16F10) tumors. We found that treatment with STING agonists resulted in increased CD69 percentage in CD8+ T cells in TILs, and decreased MHCII levels in macrophages and DCs in TILs.

In summary, we have established reporter-based in vitro assays and in vivo efficacy validation platforms, which can be used for STING agonists efficacy evaluation for cancer therapy and combination IO target therapy.

Citation Format: Xiaohe Shi, Feifei Fan, Ruicheng Wei, Xiangyang Zuo, Xiutao Lu, Quanhong Sun. A comprehensive in vitro and in vivo system to evaluate STING agonist efficacy in cancer therapeutics [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 6644.