In vitro and in vivo studies of 6,8-(diaryl)imidazo[1,2-a]pyrazin-3(7H)-ones as new antioxidants Frederic De Wael a , Paul Jeanjot a,z , Cédric Moens a,b , Tony Verbeuren c , Alex Cordi c , Eliete Bouskela d , Jean-François Rees b , Jacqueline Marchand-Brynaert a, * a Unité de Chimie Organique et Médicinale, Université catholique de Louvain, Bâtiment Lavoisier, Place Louis Pasteur 1, B-1348 Louvain-la-Neuve, Belgium b Institut des Sciences de la vie, Université catholique de Louvain, Bâtiment Carnoy, Place Croix du Sud 5, B-1348 Louvain-la-Neuve, Belgium c Institut de Recherches Servier, Rue des Moulineaux 11, F-92150 Suresnes, France d Laboratorio de Pesquisas em Microcirculaçao, Universidade do estado do Rio de Janeiro, Rua Sao Francisco Xavier 524, 20550-013 Rio de Janeiro, Brazil article info Article history: Received 10 January 2009 Revised 6 May 2009 Accepted 11 May 2009 Available online 15 May 2009 Keywords: Imidazopyrazinone Aminopyrazine Antioxidant Ischemia/reperfusion abstract A series of 5-aryl and 3,5-diaryl-2-amino-1,4-pyrazines and the derived imidazopyrazinones has been synthesized to study the chemical oxidative degradation of the bicyclic systems in vitro. Imidazopyraz- inones mainly degraded following two independent pathways producing their precursors, namely aminopyrazines, and the corresponding amidopyrazines, respectively. Despite the fact that there is no influence of the substituent of the 3-aryl group on the ratio of the products aminopyrazine/amidopyr- azine, diarylimidazopyrazinones and diarylaminopyrazines are good antioxidants in vivo. They protected against microvascular damages in ischemia/reperfusion with similar efficiencies. Ó 2009 Elsevier Ltd. All rights reserved. 1. Introduction Reactive oxygen species (ROS) and reactive nitrogen species (RNS) are produced in aerobic organisms as part of the normal physiological and metabolic processes. They are very important mediators of cell injury or death due to the damages they can in- flict if they are produced in excessive concentrations or in wrong locations. The damages that ROS/RNS cause, essentially on biolog- ical macromolecules (membrane lipids, proteins, nucleic acids, ...), are directly or indirectly implicated in the pathogenesis of various disorders such as cardiovascular diseases, reperfusion injury, Alz- heimer’s and other neurodegenerative diseases, cancer develop- ment and progression, inflammation as well as in the aging process. 1–9 Therefore the interest for the protective role of antioxidants in medicine has been growing over the last 15 years. Antioxidants are considered as potential drugs due to their ability to reduce or inhibit the free radical reactions initiated by ROS/RNS. Currently available radical scavengers are structurally related to natural anti- oxidants (vitamin A/b-carotene, vitamin C, vitamin E, green tea extracts, flavonoids, ...) and to industrial compounds such as highly hindered ortho-substituted phenols. 10–14 Coelenterazine 1a (Scheme 1) and more generally imidazo[1,2- a]pyrazin-3(7H)-ones (imidazopyrazinones) are bioluminescent substrates of luciferases, 15–17 naturally designed to react with oxy- gen in light-producing reactions. Numerous studies have been devoted to the oxidation mechanism of coelenterazine and related derivatives 1, essentially in the context of colored light production via an efficient biochemical process and the development of ana- lytical tools in biochemistry. 18–23 A few years ago, our research group considered coelenterazine and other imidazopyrazinones 1 as potential leads in the discovery of novel antioxidants for thera- peutic use. 24 We showed that imidazopyrazinones have good anti- oxidant properties as they are highly reactive with radical anion superoxide, lipid radicals, nitrofurantoin-derived radicals and per- oxynitrite; LDL protection by compounds 1 has been further dem- onstrated in vitro. Finally, cellular protection against oxidative stress and UV-irradiation damages has been observed, in the pres- ence of compounds 1, on various cell lines (human keratinocytes, rat hepatocytes, rat neuronal cells, fish erythrocytes). 24 During these previous studies, we found that the chemical precursors of imidazopyrazinones synthesis, namely the 2-amino-1,4-pyrazine derivatives 3, are also endowed with remarkable antioxidant prop- erties providing R 3 is a para-phenol (or a catechol) moiety. 25,26 The main metabolite of luciferase-catalyzed oxidation of coelenterazine (1a) is coelenteramide (2a) which is initially pro- duced in the excited state; this species emits light during decay (Scheme 1). 27,28 Similarly, numerous synthetic analogs 1 display 0968-0896/$ - see front matter Ó 2009 Elsevier Ltd. All rights reserved. doi:10.1016/j.bmc.2009.05.025 * Corresponding author. Tel.: +32 10 472740; fax: +32 10 474168. E-mail address: jacqueline.marchand@uclouvain.be (J. Marchand-Brynaert). z Accidentally deceased on January 19, 2004. Bioorganic & Medicinal Chemistry 17 (2009) 4336–4344 Contents lists available at ScienceDirect Bioorganic & Medicinal Chemistry journal homepage: www.elsevier.com/locate/bmc