Table_1_Characterization of a Novel Compound That Stimulates STING-Mediated Innate Immune Activity in an Allele-Specific Manner.XLS

The innate immune response to cytosolic DNA involves transcriptional activation of type I interferons (IFN-I) and proinflammatory cytokines. This represents the culmination of intracellular signaling pathways that are initiated by pattern recognition receptors that engage DNA and require the adaptor protein Stimulator of Interferon Genes (STING). These responses lead to the generation of cellular and tissue states that impair microbial replication and facilitate the establishment of long-lived, antigen-specific adaptive immunity. Ultimately this can lead to immune-mediated protection from infection but also to the cytotoxic T cell-mediated clearance of tumor cells. Intriguingly, pharmacologic activation of STING-dependent phenotypes is known to enhance both vaccine-associated immunogenicity and immune-based anti-tumor therapies. Unfortunately, the STING protein exists as multiple variant forms in the human population that exhibit differences in their reactivity to chemical stimuli and in the intensity of molecular signaling they induce. In light of this, STING-targeting drug discovery efforts require an accounting of protein variant-specific activity. Herein we describe a small molecule termed M04 that behaves as a novel agonist of human STING. Importantly, we find that the molecule exhibits a differential ability to activate STING based on the allelic variant examined. Furthermore, while M04 is inactive in mice, expression of human STING in mouse cells rescues reactivity to the compound. Using primary human cells in ex vivo assays we were also able to show that M04 is capable of simulating innate responses important for adaptive immune activation such as cytokine secretion, dendritic cell maturation, and T cell cross-priming. Collectively, this work demonstrates the conceivable utility of a novel agonist of human STING both as a research tool for exploring STING biology and as an immune potentiating molecule.

针对胞质DNA的固有免疫应答，涉及I型干扰素（IFN-I）与促炎细胞因子的转录激活。该过程由识别DNA的模式识别受体启动，并依赖接头蛋白干扰素基因刺激因子（Stimulator of Interferon Genes, STING）所介导的细胞内信号通路完成。此类应答可诱导细胞与组织形成抗微生物复制的状态，并促进长效抗原特异性适应性免疫的建立。最终，这一过程既可介导抗感染的免疫保护，也可通过细胞毒性T细胞介导肿瘤细胞的清除。值得注意的是，现有研究证实，STING依赖型表型的药理学激活，可同时增强疫苗相关免疫原性与免疫抗肿瘤治疗的效果。遗憾的是，STING蛋白在人群中存在多种变异体，这些变异体对化学刺激的反应性以及其所诱导的分子信号强度均存在差异。鉴于此，靶向STING的药物研发工作需要考量蛋白质变异体特异性的活性差异。本文中，我们报道了一种名为M04的小分子化合物，其可作为人源STING的新型激动剂。重要的是，我们发现该分子针对不同等位基因变异体STING的激活能力存在差异。此外，尽管M04对小鼠无活性，但在小鼠细胞中表达人源STING可恢复其对该化合物的反应性。通过在离体实验中使用原代人源细胞，我们还证实M04能够模拟与适应性免疫激活相关的关键固有免疫应答，包括细胞因子分泌、树突状细胞成熟以及T细胞交叉致敏。综上，本研究证实了这款新型人源STING激动剂的潜在应用价值：既可作为探究STING生物学功能的研究工具，也可作为免疫增强分子使用。