ABSTRACT
Oxetane-containing ring systems are increasingly used in medicinal chemistry programs to modulate druglike properties. We have shown previously that oxetanes are hydrolyzed to diols by human microsomal epoxide hydrolase (mEH). Mapping the enzymes that contribute to drug metabolism is important since an exaggerated dependence on one specific isoenzyme increases the risk of drug-drug interactions with co-administered drugs. Herein, we illustrate that mEH-catalyzed hydrolysis is an important metabolic pathway for a set of more structurally diverse oxetanes and the degree of hydrolysis is modulated by minor structural modifications. A homology model based on the Bombyx mori EH crystal structure was used to rationalize substrate binding. This study shows that oxetanes can be used as drug design elements for directing metabolic clearance via mEH, thus potentially decreasing the dependence on cytochromes P450. Metabolism by mEH should be assessed early in the design process to understand the complete metabolic fate of oxetane-containing compounds, and further study is required to allow accurate pharmacokinetic predictions of its substrates.
Subject(s)
Drug Design , Epoxide Hydrolases/chemistry , Ethers, Cyclic/chemistry , Insect Proteins/chemistry , Animals , Bombyx/enzymology , Cytochrome P-450 Enzyme System/chemistry , Cytochrome P-450 Enzyme System/metabolism , Drug Interactions , Epoxide Hydrolases/metabolism , Ethers, Cyclic/metabolism , Humans , Hydrolysis , Insect Proteins/metabolism , Metabolic Networks and Pathways , Models, Chemical , Molecular Structure , Protein Binding , Protein Domains , Substrate SpecificityABSTRACT
Oxetanyl building blocks are increasingly used in drug discovery because of the improved drug-like properties they confer on drug candidates, yet little is currently known about their biotransformation. A series of oxetane-containing analogs was studied and we provide the first direct evidence of oxetane hydrolysis by human recombinant microsomal epoxide hydrolase (mEH). Incubations with human liver fractions and hepatocytes were performed with and without inhibitors of cytochrome P450 (P450), mEH and soluble epoxide hydrolase (sEH). Reaction dependence on NADPH was investigated in subcellular fractions. A full kinetic characterization of oxetane hydrolysis is presented, in both human liver microsomes and human recombinant mEH. In human liver fractions and hepatocytes, hydrolysis by mEH was the only oxetane ring-opening metabolic route, with no contribution from sEH or from cytochrome P450-catalyzed oxidation. Minimally altering the structural elements in the immediate vicinity of the oxetane can greatly modulate the efficiency of hydrolytic ring cleavage. In particular, higher pKa in the vicinity of the oxetane and an increased distance between the oxetane ring and the benzylic nitrogen improve reaction rate, which is further enhanced by the presence of methyl groups near or on the oxetane. This work defines oxetanes as the first nonepoxide class of substrates for human mEH, which was previously known to catalyze the hydrolytic ring opening of electrophilic and potentially toxic epoxide-containing drugs, drug metabolites, and exogenous organochemicals. These findings will be of value for the development of biologically active oxetanes and may be exploited for the biocatalytic generation of enantiomerically pure oxetanes and diols.
Subject(s)
Epoxide Hydrolases/metabolism , Ethers, Cyclic/metabolism , Microsomes, Liver/metabolism , Cytochrome P-450 Enzyme System/metabolism , Hepatocytes/metabolism , Humans , Kinetics , Liver/metabolism , Oxidation-ReductionABSTRACT
The mineralocorticoid receptor (MR) is a nuclear hormone receptor involved in the regulation of body fluid and electrolyte homeostasis. In this study we explore selectivity triggers for a series of nonsteroidal MR antagonists to improve selectivity over other members of the oxosteroid receptor family. A biaryl sulfonamide compound was identified in a high-throughput screening (HTS) campaign. The compound bound to MR with pKi =6.6, but displayed poor selectivity over the glucocorticoid receptor (GR) and the progesterone receptor (PR). Following X-ray crystallography of MR in complex with the HTS hit, a compound library was designed that explored an induced-fit hypothesis that required movement of the Met852 side chain. An improvement in MR selectivity of 11- to 79-fold over PR and 23- to 234-fold over GR was obtained. Given the U-shaped binding conformation, macrocyclizations were explored, yielding a macrocycle that bound to MR with pKi =7.3. Two protein-ligand X-ray structures were determined, confirming the hypothesized binding mode for the designed compounds.
Subject(s)
Drug Design , Mineralocorticoid Receptor Antagonists/chemistry , Mineralocorticoid Receptor Antagonists/pharmacology , Receptors, Glucocorticoid/chemistry , Receptors, Glucocorticoid/metabolism , Receptors, Mineralocorticoid/metabolism , Dose-Response Relationship, Drug , High-Throughput Screening Assays , Humans , Mineralocorticoid Receptor Antagonists/chemical synthesis , Models, Molecular , Molecular Structure , Structure-Activity Relationship , Substrate SpecificityABSTRACT
A novel series of melanin concentrating hormone receptor 1 (MCHr1) antagonists were the starting point for a drug discovery program that culminated in the discovery of 103 (AZD1979). The lead optimization program was conducted with a focus on reducing lipophilicity and understanding the physicochemical properties governing CNS exposure and undesired off-target pharmacology such as hERG interactions. An integrated approach was taken where the key assay was ex vivo receptor occupancy in mice. The candidate compound 103 displayed appropriate lipophilicity for a CNS indication and showed excellent permeability with no efflux. Preclinical GLP toxicology and safety pharmacology studies were without major findings and 103 was taken into clinical trials.
