Background/Purpose: STING plays a critical role at the intersection of innate and adaptive immunity and has been linked to the pathogenesis of SSc and SLE. While initially characterized as the adaptor protein leading to type 1 IFN induction downstream of the cytosolic DNA sensor cyclic guanosine-monophosphate adenosine-monophosphate synthase (cGAS), STING can be activated independently of cGAS by several other mechanisms. Beyond its role in IFN induction, STING directly activates other pathways, e.g. NF-kB-driven inflammatory cytokine expression and the PERK-eIF2a arm of the unfolded protein response. We developed SP2H – a targeted STING degrader, preventing all STING driven activation mechanisms – as a new treatment strategy for SSc, SLE and other diseases characterized by excessive STING activation.

Methods: SP2H’s effects on STING protein levels, and on inflammatory cytokine expression induced by STING agonists, were assessed in vitro using established cell lines and primary human fibroblasts. Inhibition of STING activation by single sub-cutaneous (sc) doses of SP2H in vivo was assessed in C57Bl6 mice following a single 1mg/kg dose of the STING agonist diABZI. Survival, serum cytokine levels and inflammatory gene expression were assessed in Trex1-/- mice following treatment with 0.3mg/kg and 3.0 mg/kg or vehicle control once daily (qd) sc SP2H for 56 days (n=10/group).

Results: SP2H selectively degraded STING in vitro with DC50 values of 40-100nM, and inhibited IFN-b expression induced by STING agonists with EC50 values of 20-100nM. When given 16h prior to diAZBI, single sc doses of SP2H (either 0.3mg/kg, 1mg/kg or 3mg/kg) resulted in a >1.2log10 reduction in diABZI-induced serum interferon-b levels at all three doses, and dose-dependent reductions in serum CXCL10 (0.6-1.6log10), CCL2 (1.3-2.3log10), all p< 0.0001, and IL-6 (1.6-2.7log10), all p< 0.01 (ANOVA). Treatment of Trex1-/- mice with SP2H for 8 weeks was well tolerated and improved overall survival (Kaplan-Meier analysis, log rank analysis for trend p=0.0494), with 100% survival at the higher SP2H dose as shown in Figure 1. The 3mg/kg dose of SP2H resulted in >4.5-fold lower serum levels of both CXCL10 and TNF-a (both p< 0.0001, ANOVA) compared to vehicle controls at the end of treatment, and significantly reduced expression of inflammatory genes (CXCL10, TNF-a, IFN-b, CCL2, IFIT1, ISG15, IFI44, MX1) in the heart.

Conclusion: Targeted degradation by SP2H leads to selective STING depletion and inhibition of STING-driven inflammation in vitro and in vivo. Treatment of Trex1-/- mice with SP2H was well-tolerated, improved survival and reduced inflammation, supporting further clinical development of SP2H as a potential treatment for diseases driven by the pathological overactivation of STING.

Figure 1: Kaplan Meier analysis of the survival of Trex1-/- mice treated with vehicle control, 0.3mg/kg SP2H or 3mg/kg SP2H sc qd. Each group comprised 5 male and 5 female animals. Animals were randomized by body weight, anti-dsDNA antibody titer and serum lactate dehydrogenase titer at age 5 weeks. Treatment was initiated at age 6 weeks and continued daily for 8 weeks.

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ACR Meeting Abstracts - https://acrabstracts.org/abstract/sp2h-a-targeted-degrader-of-stimulator-of-interferon-genes-sting-selectively-inhibits-sting-driven-inflammation-in-vitro-and-in-vivo-and-improves-survival-in-trex1-mice/