The interferon gene-stimulating protein (STING) is a pivotal molecular target for regulating intrinsic immune function [1]. Upon activation, STING promotes the synthesis of type I interferons (IFNs) and pro-inflammatory cytokines, which can be leveraged to treat infections, inflammation, and tumors. The endogenous ligand for STING is 2′,3′-cyclic GMP-AMP (cGAMP), which interacts with the STING protein in the endoplasmic reticulum to facilitate its dimerization. STING translocates to the Golgi apparatus following activation, recruiting the TBK1 enzyme to mediate its phosphorylation. Subsequently, this process recruits Interferon Regulatory Factor 3 (IRF3), which TBK1 phosphorylates. Another pathway activated by STING triggers NF-κB activation that collaborates with IRF3 to induce the expression of type I interferons and other cytokines [[2], [3], [4]]. However, overactivation of STING and specific mutations in STING proteins often leads to developing inflammatory and autoimmune disorders. These disorders include gouty arthritis syndrome, STING-associated vasculopathy in infancy, and COPA syndrome, which may result in encephalopathies, pneumonitis, arthritis, and interstitial pneumonia among other severe conditions [[5], [6], [7], [8], [9]]. Furthermore, STING has been implicated in systemic lupus erythematosus development as well as non-alcoholic fatty liver disease (NAFLD), both potentially leading to cirrhosis and liver fibrosis [10,11]. Given the limited current range of therapeutic options available, there exists an urgent need for developing inhibitors targeting STING.

Several STING inhibitors and degraders have recently been identified, including the covalent inhibitors C170 and H151 (Fig. 1), which inhibit STING activation by disrupting its palmitoylation [12]. However, these compounds exhibit off-target effects leading to increased toxicity. The non-covalent inhibitor SN011 (Fig. 1) inhibits STING activation by entering the binding pocket as a dimer [13], however, its binding site remains unidentified, limiting further structural optimization. Additionally, the degraders SP23 and P8 (Fig. 1) target STING for degradation via the proteasomal and lysosomal pathways, respectively; however, their poor pharmacokinetic properties have hindered their progression into clinical trials [14,15]. As a result, there is an urgent need to develop more effective and safer STING inhibitors or degraders. Covalent inhibitors offer advantages over non-covalent ones such as increased potency, extended target binding duration, enhanced selectivity, and the capacity to target 'non-druggable' targets, thereby addressing drug resistance [[16], [17], [18]]. Therefore, it is essential to structurally optimize the covalent inhibitors C170 and H151 targeting STING. In C170, the nitrofuran ring is critical as it forms a covalent bond with Cys91 of STING, occupying the palmitoylation site. As a result, nitrofuran is unsuitable for modification [12]. The discovery of degraders SP23 and P8 suggests that structural optimization of C170 could be achieved by introducing substituents on its benzene ring [14,15]. The recently reported H151 analog compound 42 [19] suggested that introducing electron-withdrawing substituents on the phenyl ring may enhance inhibitory potency and oral bioavailability.

In this study, the benzene ring of C170 was functionalized with a hydroxyl group. Subsequently, 34 compounds were designed and synthesized by incorporating various substituents into the hydroxyl group. The inhibition of IFNβ gene expression induced by SR717 resulted in IC50 values of 0.75 μM and 0.62 μM for Y2 and Z5, respectively, which are superior to that of C170 (1.41 μM). Furthermore, the cytotoxicity of compound Y2 was significantly lower than that of C170 and Z5 across four different cell types. Y2 significantly inhibited STING pathway activation induced by the STING agonist SR717 in THP1 cells and MSA-2-induced STING pathway activation in vivo in mice. Moreover, Y2 effectively alleviated cisplatin-induced acute kidney injury (AKI) in mice. Notably, the inhibitory effect of Y2 was superior to that of C170, both in vivo and in vitro. Given that H151 shares the same mechanism of action as C170, we applied the discovery strategy used for Y2 to H151, resulting in the synthesis of the compound HY2. To further investigate the SAR, additional H151 analogs were synthesized, culminating in the optimized compound HY2. This compound inhibited SR717-induced upregulation of inflammatory factors downstream of STING in THP1 and RAW264.7 cells. In the AKI mouse model, HY2 significantly enhanced survival and effectively mitigated the deleterious effects of cisplatin on the kidneys and spleen.