RT Journal Article SR Electronic T1 A phenyl pyrrolidine derivative reveals a dual inhibition mechanism of myocardial mitochondrial permeability transition pore, which is limited by its myocardial distribution. JF Journal of Pharmacology and Experimental Therapeutics JO J Pharmacol Exp Ther FD American Society for Pharmacology and Experimental Therapeutics SP JPET-AR-2020-000359 DO 10.1124/jpet.120.000359 A1 mathieu panel A1 Abdelhakim Ahmed-Belkacem A1 Isaac Ruiz A1 jean-françois Guichou A1 jean-michel pawlotsky A1 bijan ghaleh A1 didier morin YR 2020 UL http://jpet.aspetjournals.org/content/early/2020/12/15/jpet.120.000359.abstract AB Mitochondrial permeability transition pore (mPTP) opening is a key event in cell death during myocardial ischemia-reperfusion. Inhibition of its modulator cyclophilin D (CypD) by cyclosporine A (CsA) reduces ischemia-reperfusion injury. The use of cyclosporine A in this indication is debated, however, targeting mPTP remains a major goal to achieve. We investigated the protective effects of a new original small-molecule cyclophilin inhibitor, C31, specifically designed to target CypD. CypD peptidylprolyl cis-trans isomerase PPIase activity was assessed by the standard chemotrypsin coupled assay. The effects of C31 on mPTP opening was investigated in isolated mouse cardiac mitochondria by measuring mitochondrial swelling and calcium retention capacity (CRC), in rat H9C2 cardiomyoblasts and in adult mouse cardiomyocytes by fluorescence microscopy, in isolated perfused mouse hearts and ex vivo after drug infusion in mice. C31 potently inhibited CypD PPIase activity and mitochondrial swelling. C31 was more effective at increasing mitochondrial CRC than CsA and was still able to increase CRC in Ppif-/- (CypD-inactivated) cardiac mitochondria. C31 delayed both mPTP opening and cell death in cardiomyocytes subjected to hypoxia-reoxygenation. However, high concentrations of both drugs were necessary to reduce mPTP opening in isolated perfused hearts and neither CsA nor C31 inhibited mPTP opening in heart after in vivo infusion, underlying the importance of myocardial drug distribution for cardioprotection. C31 is an original inhibitor of mPTP opening involving both CypD-dependent and -independent mechanisms. It constitutes a promising new cytoprotective agent. Optimization of its pharmacokinetic properties is now required prior to its use against cardiac ischemia-reperfusion injury. Significance Statement This study demonstrates that the new cyclophilin inhibitor C31 potently inhibits cardiac mPTP opening in vitro and ex vivo. The dual mechanism of action of C31 allows to prevent mPTP opening beyond CypD inhibition. Further development of the compound might bring promising drug candidates for cardioprotection. However, the lack of effect of both C31 and cyclosporine A following systemic administration demonstrates the difficulties of targeting myocardial mitochondria in vivo and should be taken into account in cardioprotective strategies.