Extended therapeutic window of a novel peptide inhibitor of TRPM2 channels following focal cerebral ischemia

Abstract

Introduction TRPM2 channels have been suggested to play a role in ischemic neuronal injury, specifically in males. A major hindrance to TRPM2 research has been the lack of specific TRPM2 inhibitors. The current study characterized the specificity and neuroprotective efficacy of a novel TRPM2 inhibitor. Methods Fluorescent calcium imaging (Fluo5F) was used to determine inhibitor efficacy of the TRPM2 peptide inhibitor (tat-M2NX) in HEK293 cells stably expressing hTRPM2. Adult (2–3months) and aged (18–20 months) mice were subjected to 60 min middle cerebral artery occlusion (MCAO) and injected with tat-M2NX, control scrambled peptide (tat-SCR) or clotrimazole (CTZ) either 20 min prior or 3 h after reperfusion. Infarct size was assessed using TTC staining. Results TRPM2 inhibition by tat-M2NX was observed by decreased Ca2+ influx following H2O2 exposure human TRPM2 expressing cells. Male mice pre-treated with tat-M2NX had smaller infarct volume compared to tat-SCR. No effect of tat-M2NX on infarct size was observed in female mice. Importantly, male TRPM2−/− mice were not further protected by tat-M2NX, demonstrating selectivity of tat-M2NX. Administration of tat-M2NX 3 h after reperfusion provided significant protection to males when analyzed at 24 h or 4 days after MCAO. Finally, we observed that tat-M2NX reduced ischemic injury in aged male mice. Conclusions These data demonstrate the development of a new peptide inhibitor of TRPM2 channels that provides protection from ischemic stroke in young adult and aged male animals with a clinically relevant therapeutic window.

Renoprotection: focus on TRPV1, TRPV4, TRPC6 and TRPM2

“…generating a peptidic compound by fusing a portion of the C‐terminus that corresponds >90% with the Nudix domain (M2NX) of the C‐terminus of TRPM2 channels with the TAT‐inducer of HIV (tat‐M2NX) (Shimizu et al . ). Most of the TRPM2 inhibitors lack the pharmacological profile to safely be administered to patients.…”

TRPM2 channel: A novel target for alleviating ischaemia‐reperfusion, chronic cerebral hypo‐perfusion and neonatal hypoxic‐ischaemic brain damage

“…61 MCAO-R • Neuronal death in mice was significantly reduced by administration of CTZ after reperfusion. 64 MCAO-R • Neuronal death in the neocortex in mice was substantially attenuated by TRPM2-KO. 63 CP-R • Neuronal death of CA1 pyramidal neurons in the hippocampus of mice was significantly lessened by injection of CTZ after resuscitation.…”

“…• Infarct volume in the ischaemia hemisphere in mice was also lessened by administration of tat-M2NX after reperfusion in adult and aged mice. 64…”

Redox regulation of transient receptor potential channels in the endothelium

“…Interestingly, transplantation of wild type bone marrow into TRPM2 −/− mice reversed the protective effect of TRPM2 deletion, possibly due to increased neutrophil infiltration 73 . Recently, a novel peptide inhibitor of TRPM2, tat-M2NX, was shown to reduce infarct size in male mice, even when administered 3 hours after reperfusion 74 . These findings identify TRPM2 as a possible therapeutic target to reduce ischemic damage, although direct involvement of the endothelium was not considered in these studies.…”