October 10, 2017

Blocking cGAS in Autoimmunity 

Cells of the innate immune system have developed multiple sensors to detect the presence of infectious agents and certain forms of cellular damage. One such sensor is the enzyme cyclic GMP-AMP synthase (cGAS), which responds to the presence of cytosolic double-stranded DNA (dsDNA) by catalyzing the formation of a cyclic nucleotide (cGAMP) from GTP and ATP. cGAMP acts as a second messenger to activate a signaling pathway that leads to the secretion of a number of cytokines, most importantly, interferon-β1 (IFNB1). The cGAS pathway plays a role in the immune response to intracellular pathogens (viruses and bacteria), by responding to the presence of the invading organisms’ dsDNA. However, for patients bearing an inactivating mutation of TREX1, the enzyme primarily responsible for degrading cytosolic dsDNA, cGAS promotes excessive IFNB1 production, leading to an autoimmune disease similar to systemic lupus erythematosus. This led Vanderbilt Basic Sciences investigator Manuel Ascano, his collaborators Dinshaw Patel (Memorial Sloan-Kettering Cancer Center) and Fraser Glickman (The Rockefeller University), and their laboratories to search for a small molecule inhibitor of cGAS. They developed a mass spectrometry-based high-throughput screen to evaluate a library of 123,306 compounds in search of cGAS inhibitors. Then using the four most promising hits, they embarked on a structure-guided medicinal chemistry effort that produced the lead molecule RU.521. A crystal structure of RU.521 in complex with cGAS and dsDNA indicated that the inhibitor binds in the same region of the enzyme’s active site cavity as the substrates, intermediate, and product of the reaction. Kinetic studies suggested a noncompetitive inhibitory mechanism. Studies using a murine macrophage-like cell line demonstrated that the compound is active in intact cells, and that it does not interfere, directly or indirectly, with a wide range of unrelated inflammatory signaling pathways. The ability of RU.521 to reduce IFNB1 production by bone marrow-derived macrophages from Trex1 knockout mice suggested that it may be a valuable probe to investigate the role of the cGAS pathway in autoimmunity. The work is published in the journal Nature Communications [J. Vincent et al. Nat. Commun., 2013, 8:750 DOI: 10.1038/s41467-017-00833-9].