Illuminating the mechanism of action of 4-F4t-Neuroprostane using bioorthogonal chemistry – Ima-ClickChe

Omega-3 polyunsaturated fatty acids (?3-PUFAs) and their derivatives have emerged as key compounds in many biological processes, but their mechanism of action is still obscure. Illuminating this mechanism is thus of critical interest, both for academic and drug design purposes. This is particularly true in the context of cardioprotection. Indeed, cardiovascular diseases remain a major public health problem, causing more than 17 million deaths per year, thus highlighting the urgent need to identify novel targets for pharmacological intervention. In this context, we discovered that 4(RS)-4-F4t-neuroprostane (4-F4t-NeuroP), a non-enzymatic metabolite of ?3-PUFA oxidation, reduced cellular and in vivo arrhythmias. In addition, data obtained in our consortium showed interaction of this compound with specific targets, particularly the free fatty acid receptor 4 (FFAR4), a G protein-coupled receptor that protects heart tissue from failure. This provides a putative link between this receptor and the cardioprotective effects of 4-F4t-NeuroP. However, while this link is plausible, it still needs to be established experimentally. Hence, a detailed analysis of the molecular targets and mechanisms involved is still absolutely required to improve our understanding of how this key compound triggers its biological effects, and to pave the way for developing more targeted pharmacological interventions for sudden cardiac deaths. In this context, we propose here a project that aims to validate FFAR4 as a possible signaling target for 4-F4t-NeuroP, as well as to localize and identify additional targets that may mediate its cardioprotective effects. To achieve these goals, we will develop a fully transdisciplinary strategy combining chemistry, pharmacology and pathophysiology to both delineate the signaling output of 4-F4t-NeuroP binding to FFAR4 and localize and identify any additional targets of 4-F4t-NeuroP. Specifically, our program will focus on designing and synthesizing a full range of modified 4-F4t-NeuroP probes for selective in vitro or in vivo delivery and uncaging (WP1). On the one hand, these probes will be used to decipher the impact of 4-F4t-NeuroP binding on the signaling properties and conformational features of FFAR4 using purified and cellular (recombinant and native) model systems (WP2). On the other hand, they will be used to identify additional potential 4-F4t-NeuroP targets in freshly isolated cardiomyocytes and macrophages (WP3). This research program should thus improve our understanding of the mechanisms underlying the pleiotropic effects of 4-F4t-NeuroP and its cardioprotective and anti-inflammatory actions. By doing so, it will elucidate a central biological process and pave the way for translational benefits by exploiting original 4-F4t-NeuroP targets for the clinical management of cardiac and other diseases.