title: A subset of N-substituted phenothiazines inhibits NADPH oxidases.
abstract: NADPH oxidases (NOXs) constitute a family of enzymes generating reactive oxygen species (ROS) and are increasingly recognized as interesting drug targets. Here we investigated the effects of 10 phenothiazine compounds on NOX activity using an extensive panel of assays to measure production of ROS (Amplex red, WST-1, MCLA) and oxygen consumption. Striking differences between highly similar phenothiazines were observed. Two phenothiazines without N-substitution, including ML171, did not inhibit NOX enzymes, but showed assay interference. Introduction of an aliphatic amine chain on the N atom of the phenothiazine B ring (promazine) conferred inhibitory activity toward NOX2, NOX4, and NOX5 but not NOX1 and NOX3. Addition of an electron-attracting substituent in position 2 of the C ring extended the inhibitory activity to NOX1 and NOX3, with thioridazine being the most potent inhibitor. In contrast, the presence of a methylsulfoxide group at the same position (mesoridazine) entirely abolished NOX-inhibitory activity. A cell-free NOX2 assay suggested that inhibition by N-substituted phenothiazines was not due to competition with NADPH. A functional implication of NOX-inhibitory activity of thioridazine was demonstrated by its ability to block redox-dependent myofibroblast differentiation. Our results demonstrate that NOX-inhibitory activity is not a common feature of all antipsychotic phenothiazines and that substitution on the B-ring nitrogen is crucial for the activity, whereas that on the second position of the C ring modulates it. Our findings contribute to a better understanding of NOX pharmacology and might pave the path to discovery of more potent and selective NOX inhibitors.
authors.affiliation: Department of Pathology and Immunology, Medical School, School of Pharmaceutical Sciences, University of Geneva, 1211 Geneva 4, Switzerland.
authors.forename: Tamara
authors.initials: T
authors.lastname: Seredenina
authors.affiliation: Pharmaceutical Biochemistry Group, School of Pharmaceutical Sciences, University of Geneva, 1211 Geneva 4, Switzerland.
authors.forename: Gianpaolo
authors.initials: G
authors.lastname: Chiriano
authors.affiliation: Department of Pathology and Immunology, Medical School, School of Pharmaceutical Sciences, University of Geneva, 1211 Geneva 4, Switzerland.
authors.forename: Aleksandra
authors.initials: A
authors.lastname: Filippova
authors.affiliation: Department of Pathology and Immunology, Medical School, School of Pharmaceutical Sciences, University of Geneva, 1211 Geneva 4, Switzerland.
authors.forename: Zeynab
authors.initials: Z
authors.lastname: Nayernia
authors.affiliation: Department of Pathology and Immunology, Medical School, School of Pharmaceutical Sciences, University of Geneva, 1211 Geneva 4, Switzerland.
authors.forename: Zahia
authors.initials: Z
authors.lastname: Mahiout
authors.affiliation: Department of Pathology and Immunology, Medical School, School of Pharmaceutical Sciences, University of Geneva, 1211 Geneva 4, Switzerland.
authors.forename: Laetitia
authors.initials: L
authors.lastname: Fioraso-Cartier
authors.affiliation: Department of Pathology and Immunology, Medical School, School of Pharmaceutical Sciences, University of Geneva, 1211 Geneva 4, Switzerland.
authors.forename: Olivier
authors.initials: O
authors.lastname: Plastre
authors.affiliation: Pharmaceutical Biochemistry Group, School of Pharmaceutical Sciences, University of Geneva, 1211 Geneva 4, Switzerland.
authors.forename: Leonardo
authors.initials: L
authors.lastname: Scapozza
authors.affiliation: Department of Pathology and Immunology, Medical School, School of Pharmaceutical Sciences, University of Geneva, 1211 Geneva 4, Switzerland; Department of Genetic and Laboratory Medicine, Geneva University Hospitals, Geneva, Switzerland.
authors.forename: Karl-Heinz
authors.initials: KH
authors.lastname: Krause
authors.affiliation: Department of Pathology and Immunology, Medical School, School of Pharmaceutical Sciences, University of Geneva, 1211 Geneva 4, Switzerland. Electronic address: Vincent.Jaquet@unige.ch.
authors.forename: Vincent
authors.initials: V
authors.lastname: Jaquet
mesh_terms: D000818:Animals; D016466:CHO Cells; D002454:Cell Differentiation / Q000187:drug effects; D006224:Cricetinae; D003412:Cricetulus; D004353:Drug Evaluation, Preclinical; D004791:Enzyme Inhibitors / Q000737:chemistry / Q000494:pharmacology*; D057809:HEK293 Cells; D006801:Humans; D020128:Inhibitory Concentration 50; D058628:Myofibroblasts / Q000187:drug effects / Q000502:physiology; D019255:NADPH Oxidases / Q000037:antagonists & inhibitors* / Q000737:chemistry / Q000378:metabolism; D010084:Oxidation-Reduction; D018384:Oxidative Stress; D010101:Oxygen Consumption; D010640:Phenothiazines / Q000737:chemistry / Q000494:pharmacology*; D017382:Reactive Oxygen Species / Q000378:metabolism; D013329:Structure-Activity Relationship
chemical_list: D004791:Enzyme Inhibitors; D010640:Phenothiazines; D017382:Reactive Oxygen Species; D019255:NADPH Oxidases
keywords: Fibrosis; Free radicals; NADPH oxidase; NOX inhibitor; Phenothiazine; ROS; SAR; Thioridazine