4 reference(s) found. Listing paper details in reverse chronological order. We are grateful to Keith Bradnam for improvment of this script
Title: Identification and pharmacological characterization of cholesterol-5,6-epoxide hydrolase as a target for tamoxifen and AEBS ligands de Medina P, Paillasse MR, Segala G, Poirot M, Silvente-Poirot S Ref: Proc Natl Acad Sci U S A, 107:13520, 2010 : PubMed
The microsomal antiestrogen binding site (AEBS) is a high-affinity target for the antitumor drug tamoxifen and its cognate ligands that mediate breast cancer cell differentiation and apoptosis. The AEBS, a hetero-oligomeric complex composed of 3beta-hydroxysterol-Delta8-Delta7-isomerase (D8D7I) and 3beta-hydroxysterol-Delta7-reductase (DHCR7), binds different structural classes of ligands, including ring B oxysterols. These oxysterols are inhibitors of cholesterol-5,6-epoxide hydrolase (ChEH), a microsomal epoxide hydrolase that has yet to be molecularly identified. We hypothesized that the AEBS and ChEH might be related entities. We show that the substrates of ChEH, cholestan-5alpha,6alpha-epoxy-3beta-ol (alpha-CE) and cholestan-5beta,6beta-epoxy-3beta-ol (beta-CE), and its product, cholestane-3beta,5alpha,6beta-triol (CT), are competitive ligands of tamoxifen binding to the AEBS. Conversely, we show that each AEBS ligand is an inhibitor of ChEH activity, and that there is a positive correlation between these ligands' affinity for the AEBS and their potency to inhibit ChEH (r2=0.95; n=39; P<0.0001). The single expression of D8D7I or DHCR7 in COS-7 cells slightly increased ChEH activity (1.8- and 2.6-fold), whereas their coexpression fully reconstituted ChEH, suggesting that the formation of a dimer is required for ChEH activity. Similarly, the single knockdown of D8D7I or DHCR7 using siRNA partially inhibited ChEH in MCF-7 cells, whereas the knockdown of both D8D7I and DHCR7 abolished ChEH activity by 92%. Taken together, our findings strongly suggest that the AEBS carries out ChEH activity and establish that ChEH is a new target for drugs of clinical interest, polyunsaturated fatty acids and ring B oxysterols.
Human carboxylesterase 1 (hCE1) exhibits broad substrate specificity and is involved in xenobiotic processing and endobiotic metabolism. We present and analyze crystal structures of hCE1 in complexes with the cholesterol-lowering drug mevastatin, the breast cancer drug tamoxifen, the fatty acyl ethyl ester (FAEE) analogue ethyl acetate, and the novel hCE1 inhibitor benzil. We find that mevastatin does not appear to be a substrate for hCE1, and instead acts as a partially non-competitive inhibitor of the enzyme. Similarly, we show that tamoxifen is a low micromolar, partially non-competitive inhibitor of hCE1. Further, we describe the structural basis for the inhibition of hCE1 by the nanomolar-affinity dione benzil, which acts by forming both covalent and non-covalent complexes with the enzyme. Our results provide detailed insights into the catalytic and non-catalytic processing of small molecules by hCE1, and suggest that the efficacy of clinical drugs may be modulated by targeted hCE1 inhibitors.
Title: Mammalian carboxylesterases: from drug targets to protein therapeutics Redinbo MR, Potter PM Ref: Drug Discov Today, 10:313, 2005 : PubMed
Our understanding of the detailed recognition and processing of clinically useful therapeutic agents has grown rapidly in recent years, and we are now able to begin to apply this knowledge to the rational treatment of disease. Mammalian carboxylesterases (CEs) are enzymes with broad substrate specificities that have key roles in the metabolism of a wide variety of clinical drugs, illicit narcotics and chemical nerve agents. Here, the functions, mechanism of action and structures of human CEs are reviewed, with the goal of understanding how these proteins are able to act in such a non-specific fashion, yet catalyze a remarkably specific chemical reaction. Current approaches to harness these enzymes as protein-based therapeutics for drug and chemical toxin clearance are described, as well as their uses for targeted chemotherapeutic prodrug activation. Also included is an outline of how selective CE inhibitors could be used as co-drugs to improve the efficacy of clinically approved agents.
Our quest to identify target proteins involved in the activity of tamoxifen led to the design of photoaffinity ligand analogues of tamoxifen able to cross-link such proteins. A new tritiated photoprobe, 4-(2-morpholinoethoxy)benzophenone (MBoPE), was synthesized and used to identify proteins involved in tamoxifen binding in rat liver. MBoPE, which has structural features in common with the potential antagonist of the intracellular histamine receptor (N,N-diethyl-2-[(4-phenylmethyl)phenoxy]ethanamine HCl: DPPE) is unable to bind the estrogen receptor although it does compete with tamoxifen for an antiestrogen binding site (AEBS). This tritiated benzophenone derivative was obtained by metal-catalyzed halogen-tritium replacement reaction. Because of its high specific activity, four target proteins could be photolabeled, three of which were identified with M(r) of 60,000, 49,500, and 14,000, while the fourth at 27,500 was in too low an amount and could not be sequenced. The 49.5 kDa protein corresponded by mass spectrometry to the microsomal epoxide hydrolase already identified with an aryl azide photoprobe [Mesange, F., et al. (1998) Biochem. J. 334, 107-112]. The 60 and 14 kDa proteins were identified as the carboxylesterase (ES10) and the liver fatty acid binding protein (L-FABP), respectively. The inhibitory effect of tamoxifen on carboxylesterase activity and the competitive efficacy of oleic acid on [(3)H]tamoxifen binding suggest that both proteins are AEBS subunits. Moreover, treatment of hepatocytes with antisense mRNA directed against ES10 or L-FABP abolished both tamoxifen and MBoPE binding. On the basis of previous pharmacological arguments, the 27.5 kDa protein might correspond to the sigma I receptor. Altogether, these results confirm that the microsomal epoxide hydrolase is a target for tamoxifen and provide evidence of two new target proteins implicated in cell lipid metabolism.
