Diverse Nrf2 Activators Coordinated to Cobalt Carbonyls Induce Heme Oxygenase‑1 and Release Carbon Monoxide in Vitro and in Vivo Aniket Nikam, †,‡ Anthony Ollivier, § Michael Rivard, § Jayne Louise Wilson, †,‡ Kevin Mebarki, § Thierry Martens, § Jean-Luc Dubois-Rande ́ , ∥ Roberto Motterlini,* ,†,‡ and Roberta Foresti* ,†,‡ † Equipe 12, Inserm U955, 8 Rue du Ge ́ ne ́ ral Sarrail, Cré teil, 94000, France ‡ Faculty of Medicine, University Paris Est Cre ́ teil, Cré teil, 94000, France § ICMPE (UMR 7182), CNRS, UPEC, University Paris Est, F-94320 Thiais, France ∥ AP-HP, Henri Mondor Hospital, Service Hospitalier, Cre ́ teil, 94000, France * S Supporting Information ABSTRACT: The Nrf2/heme oxygenase-1 (HO-1) axis affords significant protection against oxidative stress and cellular damage. We synthesized a series of cobalt-based hybrid molecules (HYCOs) that combine an Nrf2 inducer with a releaser of carbon monoxide (CO), an anti- inflammatory product of HO-1. Two HYCOs markedly increased Nrf2/HO-1 expression, liberated CO and exerted anti-inflammatory activity in vitro. HYCOs also up-regulated tissue HO-1 and delivered CO in blood after administration in vivo, supporting their potential use against inflammatory conditions. ■ INTRODUCTION Cells employ several inducible protective pathways in order to combat oxidative stress, inflammation, and other harmful conditions. The nuclear factor erythroid 2-related factor 2 (Nrf2) and its downstream genes are major players in this protective response, 1,2 and heme oxygenase-1 (HO-1), an inducible enzyme that converts the substrate heme to the biologically active molecules biliverdin, iron and carbon monoxide (CO), significantly contributes to this effect owing to its antioxidant and anti-inflammatory actions. 3,4 In particular, CO has been studied for its antiapoptotic and signaling roles as well as for preventing cardiovascular and inflammatory damage. 5−7 Biliverdin and its reduced form bilirubin are also renowned for their antioxidant and immunomodulatory activities. 8 A variety of organic scaffolds of natural origin have been investigated for their property to activate Nrf2. Typical examples include sulforaphane, curcumin, or carnosol, 9−12 which stimulate Nrf2 activation by preventing the interaction of Nrf2 and binding covalently to Kelch-like ECH-associated protein 1 (Keap 1), a cytosolic Nrf2 repressor that maintains low levels of the transcription factor in unstressed conditions. 13 In view of the importance of Nrf2, efforts are under way to discover pharmacological agents that target this system. 4,14 Indeed, interesting Nrf2 inducers such as dimethyl fumarate (DMF) and bardoxolone methyl as well as sulforaphane-rich broccoli extracts are finding their application in the clinic in diseases characterized by chronic inflammation and oxidative stress. 15−18 In addition, new approaches to activate Nrf2 are being investigated by focusing on molecules that disrupt Keap1−Nrf2 binding via noncovalent mechanisms 19 or that enhance Nrf2 expression via epigenetic modifications. 20,21 Our group is exploring an alternative strategy that aims to harness in one prototypical molecule the beneficial properties of Nrf2 activators with those of HO-1. For this endeavor, we have taken advantage of CO-releasing molecules (CO-RMs), which we identified as pharmacological compounds that deliver CO to biological tissues and mimic in many respects the salutary effects of HO-1. 4,7,22 The premise is that a new molecule, containing an Nrf2 inducer bound to a CO-RM, will provide greater tissue protection by first limiting damage through CO delivery and subsequently promoting the endogenous up- regulation of Nrf2-dependent defensive genes and proteins, a process that takes several hours due to transcription and translation processes. With this concept in mind, we recently reported the synthesis and preliminary biological character- ization of two hybrid molecules exhibiting the dual ability to activate the Nrf2/HO-1 cytoprotective pathway and to release controlled amounts of CO. 23 The design of these molecules, Received: September 29, 2015 Published: January 5, 2016 Brief Article pubs.acs.org/jmc © 2016 American Chemical Society 756 DOI: 10.1021/acs.jmedchem.5b01509 J. Med. Chem. 2016, 59, 756−762