Douglas R. Cary

Structure-Based Approach for the Discovery of Pyrrolo[3,2-d]pyrimidine-Based EGFR T790M/L858R Mutant Inhibitors

The epidermal growth factor receptor (EGFR) family plays a critical role in vital cellular processes and in various cancers. Known EGFR inhibitors exhibit distinct antitumor responses against the various EGFR mutants associated with nonsmall-cell lung cancer. The L858R mutation enhances clinical sensitivity to gefitinib and erlotinib as compared with wild type and reduces the relative sensitivity to lapatinib. In contrast, the T790M mutation confers drug resistance to gefitinib and erlotinib. We determined crystal structures of the wild-type and T790M/L858R double mutant EGFR kinases with reversible and irreversible pyrrolo[3,2-d]pyrimidine inhibitors based on analogues of TAK-285 and neratinib. In these structures, M790 adopts distinct conformations to accommodate different inhibitors, whereas R858 allows conformational variations of the activation loop. These results provide structural insights for understanding the structure-activity relationships that should contribute to the development of potent inhibitors against drug-sensitive or -resistant EGFR mutations.

Design, synthesis, and evaluation of novel VEGFR2 kinase inhibitors: Discovery of [1,2,4]triazolo[1,5-a]pyridine derivatives with slow dissociation kinetics

For the purpose of discovering novel type-II inhibitors of vascular endothelial growth factor receptor 2 (VEGFR2) kinase, we designed and synthesized 5,6-fused heterocyclic compounds bearing a anilide group. A co-crystal structure analysis of imidazo[1,2-b]pyridazine derivative 2 with VEGFR2 revealed that the N1-nitrogen of imidazo[1,2-b]pyridazine core interacts with the backbone NH group of Cys919. To retain this essential interaction, we designed a series of imidazo[1,2-a]pyridine, [1,2,4]triazolo[1,5-a]pyridine, thiazolo[5,4-b]pyridine, and 1,3-benzothiazole derivatives maintaining a ring nitrogen as hydrogen bond acceptor (HBA) at the corresponding position. All compounds thus designed displayed strong inhibitory activity against VEGFR2 kinase, and the [1,2,4]triazolo[1,5-a]pyridine 13d displayed favorable physicochemical properties. Furthermore, 13d inhibited VEGFR2 kinase with slow dissociation kinetics and also inhibited platelet-derived growth factor receptor (PDGFR) kinases. Oral administration of 13d showed potent anti-tumor efficacy in DU145 and A549 xenograft models in nude mice.

Discovery of Novel 1,4-Diacylpiperazines as Selective and Cell-Active eIF4A3 Inhibitors

Eukaryotic initiation factor 4A3 (eIF4A3), a member of the DEAD-box RNA helicase family, is one of the core components of the exon junction complex (EJC). The EJC is known to be involved in a variety of RNA metabolic processes typified by nonsense-mediated RNA decay (NMD). In order to identify molecular probes to investigate the functions and therapeutic relevance of eIF4A3, a search for selective eIF4A3 inhibitors was conducted. Through the chemical optimization of 1,4-diacylpiperazine derivatives identified via high-throughput screening (HTS), we discovered the first reported selective eIF4A3 inhibitor 53a exhibiting cellular NMD inhibitory activity. A surface plasmon resonance (SPR) biosensing assay ascertained the direct binding of 53a and its analog 52a to eIF4A3 and revealed that the binding occurs at a non-ATP binding site. Compounds 52a and 53a represent novel molecular probes for further study of eIF4A3, the EJC, and NMD.

TP-064, a potent and selective small molecule inhibitor of PRMT4 for multiple myeloma

Protein arginine methyltransferase (PRMT) 4 (also known as coactivator-associated arginine methyltransferase 1; CARM1) is involved in a variety of biological processes and is considered as a candidate oncogene owing to its overexpression in several types of cancer. Selective PRMT4 inhibitors are useful tools for clarifying the molecular events regulated by PRMT4 and for validating PRMT4 as a therapeutic target. Here, we report the discovery of TP-064, a potent, selective, and cell-active chemical probe of human PRMT4 and its co-crystal structure with PRMT4. TP-064 inhibited the methyltransferase activity of PRMT4 with high potency (half-maximal inhibitory concentration, IC50 < 10 nM) and selectivity over other PRMT family proteins, and reduced arginine dimethylation of the PRMT4 substrates BRG1-associated factor 155 (BAF155; IC50= 340 ± 30 nM) and Mediator complex subunit 12 (MED12; IC50 = 43 ± 10 nM). TP-064 treatment inhibited the proliferation of a subset of multiple myeloma cell lines, with affected cells arrested in G1 phase of the cell cycle. TP-064 and its negative control (TP-064N) will be valuable tools to further investigate the biology of PRMT4 and the therapeutic potential of PRMT4 inhibition.

Studies of CDK 8/19 inhibitors: Discovery of novel and selective CDK8/19 dual inhibitors and elimination of their CYP3A4 time-dependent inhibition potential

In this article, synthetic studies around a pyridylacrylamide-based hit compound (1), utilizing structure-based drug design guided by CDK8 docking models, is discussed. Modification of the pendant 4-fluorophenyl group to various heteroaromatic rings was conducted aiming an interaction with the proximal amino acids, and then replacement of the morpholine ring was targeted for decreasing potential of time-dependent CYP3A4 inhibition. These efforts led to the compound 4k, with enhanced CDK8 inhibitory activity and no apparent potential for time-dependent CYP3A4 inhibition (CDK8 IC50: 2.5nM; CYP3A4 TDI: 99% compound remaining). Compound 4k was found to possess a highly selective kinase inhibition profile, and also showed favorable pharmacokinetic profile. Oral administration of 4k (15mg/kg, bid. for 2weeks) suppressed tumor growth (T/C 29%) in an RPMI8226 mouse xenograft model.

Discovery of Novel 5-(Piperazine-1-carbonyl)pyridin-2(1H)-one Derivatives as Orally eIF4A3-Selective Inhibitors

Starting from our previous eIF4A3-selective inhibitor 1a, a novel series of (piperazine-1-carbonyl)pyridin-2(1H)-one derivatives was designed, synthesized, and evaluated for identification of orally bioavailable probe molecules. Compounds 1o and 1q showed improved physicochemical and ADMET profiles, while maintaining potent and subtype-selective eIF4A3 inhibitory potency. In accord with their promising PK profiles and results from initial in vivo PD studies, compounds 1o and 1q showed antitumor efficacy with T/C values of 54% and 29%, respectively, without severe body weight loss. Thus, our novel series of compounds represents promising probe molecules for the in vivo pharmacological study of selective eIF4A3 inhibition.