Cyclophilin Selective Inhibitors Discovered and Developed from a DNA-Templated Library

Abstract

DNA-encoded libraries (DELs) are a powerful method for small molecule discovery, allowing for rapid, pooled screening of 105-1012 compounds in a single vessel, and have led to a multitude of novel ligands for biologically relevant targets. I and others within the Liu lab have pioneered the use of DNA-templated synthesis (DTS) to create sequence-defined DELs in a programmable manner. In this thesis, I describe work that extended DEL technology to develop novel cyclophilin-selective inhibitors that co-opt a secondary pocket near the prolyl isomerase active site. My work focused on discovering inhibitors against cyclophilin D (CypD), a regulator of the mitochondrial permeability transition pore (mPTP) and a potential therapeutic target for a variety of diseases. After an initial discovery of a weak and promiscuous inhibitor, I embarked on a thorough campaign to improve potency and selectivity for CypD, which included structure-activity relationship analyses, site-directed mutagenesis, and 10 inhibitor-CypD co-crystal structures. This pipeline yielded potent and selective CypD inhibitors, which showed bioactive CypD-mPTP inhibition in isolated mitochondria. The structural basis of CypD-selective inhibition allowed us to further establish a paradigm to create other cyclophilin selective inhibitors. I leveraged this paradigm to rationally design a cyclophilin E (CypE)-selective inhibitor through reversible covalent modification. To our knowledge, our CypD and CypE inhibitors represent the first cyclophilin selective inhibitors to date, provides new chemical tools to probe their endogenous biological functions, and establishes a first-in-class generalizable model to guide selective inhibitor development for other cyclophilins.