Li, LingyinYin, QianKuss, PiaMaliga, ZoltanMillán, José L.Wu, HaoMitchison, Timothy2015-07-132014Li, Lingyin, Qian Yin, Pia Kuss, Zoltan Maliga, José L. Millán, Hao Wu, and Timothy J. Mitchison. 2014. “Hydrolysis of 2′3′-cGAMP by ENPP1 and design of non-hydrolyzable analogs.” Nature chemical biology 10 (12): 1043-1048. doi:10.1038/nchembio.1661. http://dx.doi.org/10.1038/nchembio.1661.1552-4450http://nrs.harvard.edu/urn-3:HUL.InstRepos:17295791Agonists of mouse STING (TMEM173) shrink and even cure solid tumor by activating innate immunity; human STING agonists are needed to test this therapeutic hypothesis in man. The endogenous STING agonist is 2′3′-cGAMP, a 2nd messenger that signals the presence of cytosolic dsDNA. We report activity-guided partial purification and identification of ENPP1 as the dominant 2′3′-cGAMP hydrolyzing activity in cultured cells. The hydrolysis activity of ENPP1 was confirmed using recombinant protein and was depleted in tissue extracts and plasma from Enpp1-/- mice. We synthesized a hydrolysis-resistant bis-phosphothioate analog of 2′3′-cGAMP (2′3′-cGsAsMP) with similar affinity for human STING in vitro and 10 times more potent at inducing IFN-β secretion from human THP1 monocytes. Studies in mouse Enpp1-/- lung fibroblasts indicate that resistance to hydrolysis contributes significantly to its higher potency. 2′3′-cGsAsMP is therefore improved over natural 2′3′-cGAMP as a model agonist, and has potential as a vaccine adjuvant and cancer therapeutic.en-USJournal Article2015-07-1310.1038/nchembio.1661