Session Type: ACR Plenary Session
Session Time: 11:00AM-12:30PM
Background/Purpose: Type I interferon, (IFN-I) is strongly implicated in the pathogenesis of SLE as well as rare monogenic ‘interferonopathies’ such as Aicardi-Goutieres Syndrome (AGS) caused by mutations in the DNA exonuclease, TREX1. A new DNA activated IFN-I pathway, cyclic GMP-AMP (cGAMP) synthase (cGAS), was recently discovered and linked to mouse models of AGS and Lupus. To identify potential inhibitors of the DNA-cGAS interaction, we performed in silico screening of chemical and drug libraries and tested the lead candidate, X6, in the mouse model of AGS.
Methods: In silico structure-based drug screening and ligand binding studies were performed by the CANDO platform followed by docking analysis via Autodock Vina and Chimera. In vitro cGAS activity/cGAMP production was analyzed by Thin Layer Chromatography (TLC). Following DNA cell transfections, cytokines were quantified by qPCR, ELISA or an ISRE-luciferase reporter assay. Trex1-/- mouse were treated orally with 25mg/kg/day drug X6 (n=8) for 8 weeks from birth. Multiple Reaction Monitoring by Ultra-Performance Liquid Chromatogram coupled with tandem Mass Spectrometer (UPLC-MS/MS) was used to quantify cGAMP. Heart pathology on Hematoxylin and Eosin stained slides was blindly scored by a pathologist.
Results: In silico analysis of drug libraries identified several antimalarial drugs (AMD) which could potentially inhibit cGAS activity by interacting with the cGAS/DNA dimer complex. TLC revealed that AMDs attenuated cGAS activity and inhibited cGAMP production in a dose dependent manner. These AMD also inhibited IFN-b expression in THP1 cells transfected with dsDNA and in 293T cells transfected with cGAS/STING plasmids validating that cGAS is a target of AMD. Based on the relative potencies of AMD and additional modeling, we synthesized several new AMD derivatives. One of these compounds, X6, had excellent water solubility and cell penetration. X6 localized to the cytosol and had a lower toxicity profile compared to quinacrine. Biochemical and cellular assays revealed that X6 was a potent inhibitor of IFN-I production. Since deficiency of the TREX1 in mice (TREX1-/-) leads to an autoimmune myocarditis and lupus-like systemic autoimmunity with increased Interferon Signature Genes (ISGs) expression and cGAMP production, we treated these mice with X6. When compared to vehicle control treated TREX1-/- mice, drug X6 reduced ISGs ISG15 (p<0.01) and ISG20 (p<0.01) expression in the spleen and CXCL10 (p<0.01) and ISG15 (p<0.05) expression in the heart. In addition, treatment with X6 resulted in a statistically significant reduction in cGAMP in the heart (p<0.05) as well as a reduction in endocardial fibrosis severity scores (p<0.05) compared to control.
Conclusion: Our studies indicate that, in addition to reducing activation of intracellular TLRs, AMD attenuate cGAS activity. Since disease in TREX1-/- mice is absolutely dependent on the cGAS pathway as indicated by rescue of disease in double knockouts, we show here that the novel AMD derivative, X6, attenuates disease by inhibiting cGAS activity and IFN-b expression in vivo. This class of drugs could be beneficial for the treatment of AGS and /or Lupus.
To cite this abstract in AMA style:An J, Woodward J, Minie M, Sun X, Tanaka L, Peng Y, Snyder J, Sasaki T, Elkon KB. Novel Anti-Malarial Drug Derivative Inhibited Type I Interferon Production and Autoimmune Inflammation through Inhibition of CGAS-Sting Pathway in Trex1-/- Mouse [abstract]. Arthritis Rheumatol. 2016; 68 (suppl 10). https://acrabstracts.org/abstract/novel-anti-malarial-drug-derivative-inhibited-type-i-interferon-production-and-autoimmune-inflammation-through-inhibition-of-cgas-sting-pathway-in-trex1-mouse/. Accessed March 20, 2019.
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