Abstract 4259: PT199, a next generation anti-CD73 mAb that inhibits both membrane-bound and soluble CD73 activity to completion without “hook effect”

Abstract

Abstract CD73, also known as ecto-5’-nucleotidase, is implicated in promoting immunosuppressive tumor microenvironment (TME) through production of adenosine. The first generation of anti-CD73 therapeutic antibodies currently in clinical studies have largely exhibited subdued clinical activities mainly due to several unfavorable characteristics such as incomplete enzyme inhibition and/or “hook effect”. These weaknesses are caused by the unfavorable target binding sites and enzyme inhibition mechanisms, which explains why these molecules do not inhibit soluble CD73 enzyme activity, or why the enzyme inhibition depends on certain stochiometric ratios of CD73 and the antibody and exhibits “hook effect” in dose-response curves. Here we report a next generation humanized monoclonal antibody against human CD73 (hCD73), PT199, which binds to a unique site on human CD73 that confers a different mechanism of enzyme inhibition and consequently does not have the flaws associated with the first generation of anti-CD73 mAbs currently in clinic. By directly binding to an epitope that is in or near the CD73 catalytic domain, PT199 potently inhibits both soluble shed CD73 and membrane-bound CD73 activity to completion without “hook effect”. Enzyme kinetic studies demonstrate that PT199 inhibits CD73 enzyme activity in a noncompetitive manner, allowing complete CD73 inhibition even at high AMP concentrations reported in TME. In a cell-based functional assay, PT199 inhibition of CD73 alleviates the immunosuppression effect of AMP on T cell proliferation. Compared with the first-generation molecules, PT199 completely inhibits CD73 activity in cancer patient serum. Furthermore, when administrated to hCD73 knock-in mice, PT199 greatly elevates endogenous AMP level, demonstrating a robust pharmacodynamic effect in blocking the conversion of AMP to adenosine. Comparative mechanistic studies indicate that PT199 binds to both the “open conformation” (inactive enzyme state) and the “closed conformation” (active enzyme state) with high affinity but with shifted binding curves, suggesting that PT199 may inhibit the CD73 enzyme activity by preventing the transition of the enzyme from the “open conformation” to the “closed conformation”. PT199 did not cause B cell reduction in either acute or repeated dosing studies in NHP. In NHP toxicology studies, PT199 exhibits no clinical observations at up to 300 mg/kg weekly repeated dosing and has an excellent PK profile typical of mAbs. Taken together, our data confirms PT199 as a potential best-in-class anti-hCD73 mAb. Phase 1 clinical trial is planned to start in the first quarter of 2022. Citation Format: Haiqun (John) Jia, Jack Li, Fen Pei, Lorraine Greenwood, Lauren Pejza, Yan Long, Kelan Chen, Jessica Perer, Ming Wang, Hui Zou. PT199, a next generation anti-CD73 mAb that inhibits both membrane-bound and soluble CD73 activity to completion without “hook effect” [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 4259.