Design and Synthesis of Inhibitors Targeting DCN1/3 and Inhibitors Targeting MLL1 Methyltransferase

Abstract

The goal of my work presented in this dissertation was to fill the unmet need for compounds that can be used to interrogate the biology of the Cullin-RING ubiquitin ligases (CRLs) and the MLL1 methyltransferase. We conducted structure-based design, organic synthesis, and biochemical/biophysical assays to discover potent inhibitors targeting DCN1, DCN3 and, MLL1 methyltransferase. In DCN1 project, we describe the design, synthesis, and evaluation of peptidomimetics targeting the DCN1-UBC12 protein-protein interaction. Starting from a 12-residue UBC12 peptide, we successfully obtained a series of peptidomimetic compounds that bind to DCN1 protein with KD values of < 10 nM. Determination of a co-crystal structure of a potent peptidomimetic inhibitor complexed with DCN1 provided the structural basis for their high-affinity interaction. Cellular investigation of one potent DCN1 inhibitor, compound DI-404, reveals that it effectively and selectively inhibits the neddylation of cullin 3 over other cullin members. Further optimization of DI-404 may yield a new class of therapeutics for the treatment of human diseases in which cullin 3 CRL plays a key role. In DCN3 Project, we report the discovery of first-in-class, small-molecule inhibitors targeting the DCN3-UBE2F interaction, hereafter called DCN3 inhibitors. Our efforts have yielded potent and highly selective small-molecule DCN3 inhibitors, exemplified by TC-6304 which binds to DCN3 with a Ki value of 35 nM and fails to bind to DCN1 at concentrations as high as 30 µM. Cellular thermal shift assays showed that TC-6304 engages DCN3 in cells in a dose-dependent manner but does not affect the neddylation of any of the cullins that were examined, indicating the different roles of DCN3 when compared to DCN1. This study provides first-in-class, potent and selective small-molecule inhibitors of DCN3. In MLL project, we describe the design, synthesis and evaluation of a chemical library focused on S-adenosylmethionine-based compounds which led to the discovery of first-in-class, potent small-molecule inhibitors directly targeting the MLL1 SET domain. These are exemplified by compound TC-5115 with an IC50 value of 15 nM against MLL1 methyltransferase, which is 48 times more potent than the pan-HMT inhibitor, SAH. Determination of co-crystal structures for a number of these MLL1 inhibitors reveals that they adopt a unique binding mode that interacts with SET-I domain of MLL1 methyltransferase.