Discovery and characterization of a novel cGAS covalent inhibitor for the treatment of inflammatory bowel disease

Jia Song1,2, Rui-rui Yang2,3,4, Jie Chang2, Ya-dan Liu5, Cheng-hao Lu5, Li-fan Chen2,3, Hao Guo2,3, Ying-hui Zhang2,3, Zi-sheng Fan2,5, Jing-yi Zhou2,5, Gui-zhen Zhou2,5, Ke-ke Zhang2,5, Xiao-min Luo2,3,5, Kai-xian Chen2,3,5, Hua-liang Jiang1,2,3,4,5, Su-lin Zhang2,3, Ming-yue Zheng2,3,5
1 The First Affiliated Hospital of USTC, Division of Life Sciences and Medicine, University of Science and Technology of China, Hefei 230001, China
2 Drug Discovery and Design Center, State Key Laboratory of Drug Research, Shanghai Institute of Materia Medica, Chinese Academy of Sciences, Shanghai 201203, China
3 University of Chinese Academy of Sciences, Beijing 100049, China
4 Shanghai Institute for Advanced Immunochemical Studies, and School of Life Science and Technology, Shanghai Tech University, Shanghai 200031, China
5 School of Chinese Materia Medica, Nanjing University of Chinese Medicine, Nanjing 210023, China
Correspondence to: Hua-liang Jiang:, Su-lin Zhang:, Ming-yue Zheng:,
DOI: 10.1038/s41401-022-01002-5
Received: 20 June 2022
Accepted: 19 September 2022
Advance online: 13 October 2022


Cyclic GMP-AMP synthase (cGAS), a cytosolic DNA sensor, acts as a nucleotidyl transferase that catalyzes ATP and GTP to form cyclic GMP-AMP (cGAMP) and plays a critical role in innate immunity. Hyperactivation of cGAS-STING signaling contributes to hyperinflammatory responses. Therefore, cGAS is considered a promising target for the treatment of inflammatory diseases. Herein, we report the discovery and identification of several novel types of cGAS inhibitors by pyrophosphatase (PPiase)-coupled activity assays. Among these inhibitors, 1-(1-phenyl-3,4-dihydro-1H-pyrrolo[1,2-a]pyrazin-2-yl)prop-2-yn-1-one (compound 3) displayed the highest potency and selectivity at the cellular level. Compound 3 exhibited better inhibitory activity and pathway selectivity than RU.521, which is a selective cGAS inhibitor with anti-inflammatory effects in vitro and in vivo. Thermostability analysis, nuclear magnetic resonance and isothermal titration calorimetry assays confirmed that compound 3 directly binds to the cGAS protein. Mass spectrometry and mutation analysis revealed that compound 3 covalently binds to Cys419 of cGAS. Notably, compound 3 demonstrated promising therapeutic efficacy in a dextran sulfate sodium (DSS)-induced mouse colitis model. These results collectively suggest that compound 3 will be useful for understanding the biological function of cGAS and has the potential to be further developed for inflammatory disease therapies.
Keywords: cyclic GMP-AMP synthase; covalent inhibitor; inflammatory bowel disease; high-throughput screening

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