title: Squalene synthase as a chemotherapeutic target in Trypanosoma cruzi and Leishmania mexicana.
abstract: Trypanosoma cruzi and Leishmania parasites have a strict requirement for specific endogenous sterols (ergosterol and analogs) for survival and growth and cannot use the abundant supply of cholesterol present in their mammalian hosts. Squalene synthase (SQS, E.C. 2.5.1.21) catalyzes the first committed step in sterol biosynthesis and is currently under intense study as a possible target for cholesterol-lowering agents in humans, but it has not been investigated as a target for anti-parasitic chemotherapy. SQS is a membrane-bound enzyme in both T. cruzi epimastigotes and Leishmania mexicana promastigotes with a dual subcellular localization, being almost evenly distributed between glycosomes and mitochondrial/microsomal vesicles. Kinetic studies showed that the parasite enzymes display normal Michaelis-Menten kinetics and the values of the kinetic constants are comparable to those of the mammalian enzyme. We synthesized and purified 3-(biphenyl-4-yl)-3-hydroxyquinuclidine (BPQ-OH), a potent and specific inhibitor of mammalian SQS and found that it is also a powerful non-competitive inhibitor of T. cruzi and L. mexicana SQS, with K(i)'s in the range of 12-62 nM. BPQ-OH induced a dose-dependent reduction of proliferation the extracellular stages of these parasites with minimal growth inhibitory concentrations (MIC) of 10-30 microM. Growth inhibition and cell lysis induced by BPQ-OH in both parasites was associated with complete depletion of endogenous squalene and sterols, consistent with a blockade of de novo sterol synthesis at the level of SQS. BPQ-OH was able to eradicate intracellular T. cruzi amastigotes from Vero cells cultured at 37 degrees C, with a MIC of 30 microM with no deleterious effects on host cells. Taken together, these results support the notion that SQS inhibitors could be developed as selective anti-trypanosomatid agents.
authors.affiliation: Laboratorio de Quimica Biológica, Centro de Bioquimica y Biofisica, Instituto Venezolano de Investigaciones Cientificas, Caracas 1020, Venezuela. jaurbina@ivic.ve
authors.forename: Julio A
authors.initials: JA
authors.lastname: Urbina
authors.forename: Juan Luis
authors.initials: JL
authors.lastname: Concepcion
authors.forename: Salomé
authors.initials: S
authors.lastname: Rangel
authors.forename: Gonzalo
authors.initials: G
authors.lastname: Visbal
authors.forename: Renee
authors.initials: R
authors.lastname: Lira
mesh_terms: D000818:Animals; D000981:Antiprotozoal Agents / Q000378:metabolism / Q000494:pharmacology* / Q000627:therapeutic use; D002522:Chlorocebus aethiops; D016503:Drug Delivery Systems; D004791:Enzyme Inhibitors / Q000494:pharmacology / Q000627:therapeutic use; D013186:Farnesyl-Diphosphate Farnesyltransferase / Q000037:antagonists & inhibitors* / Q000737:chemistry / Q000302:isolation & purification; D007700:Kinetics; D007894:Leishmania mexicana / Q000187:drug effects* / Q000201:enzymology / Q000254:growth & development; D008018:Life Cycle Stages; D008958:Models, Molecular; D011528:Protozoan Infections / Q000188:drug therapy; D011812:Quinuclidines / Q000494:pharmacology*; D014344:Trypanocidal Agents / Q000494:pharmacology; D014349:Trypanosoma cruzi / Q000187:drug effects* / Q000201:enzymology / Q000254:growth & development; D014709:Vero Cells
chemical_list: C099381:3-(biphenyl-4-yl)-3-hydroxyquinuclidine; D000981:Antiprotozoal Agents; D004791:Enzyme Inhibitors; D011812:Quinuclidines; D014344:Trypanocidal Agents; D013186:Farnesyl-Diphosphate Farnesyltransferase