P16

TRPC3 inhibition reduces calcium entry in muscles of DMDmdx rats

M Giri(1) M Cotinat(1) A Mellet(1) A Bourdon(1) A Lafoux(1,2) T Girard(1) C Huchet(1,2) M Biette(1) M Ledevin(3) T Larcher(3) O Adjali(1) C Le Guiner(1) B Fraysse(1)

1:Nantes Université, CHU Nantes, INSERM, TaRGeT (Translational Research in Gene Therapy - UMR 1089) ; Nantes, France; 2:Therassay Platform, Capacités, Nantes Université ; Nantes, France; 3:INRAE Oniris, UMR 703, PAnTher, APEX ; Nantes, France

Duchenne Muscular Dystrophy (DMD) is a severe, progressive muscle-wasting disease caused by mutations in the DMD gene, resulting in dystrophin deficiency and muscle degeneration. Patients typically lose ambulation during adolescence and face premature mortality in their 20s to 40s due to respiratory or cardiac failure. Excessive calcium (Ca²⁺) entry into myocytes contributes to cellular damage and death. Previous research from our laboratory identified the TRPC3 (Transient Receptor Potential Canonical 3) channel as a key contributor to this abnormal Ca²⁺ influx. We also showed that treatment with a recombinant adeno-associated virus encoding micro-dystrophin (rAAV-MD) only partially corrects these abnormalities. The present study investigated whether inhibiting TRPC3 could reduce Ca²⁺ entry in DMDmdx rat muscles, an animal model of DMD, and could have a synergistic effect when combined with a rAAV-MD. DMDmdx rats injected systemically with a rAAV-MD or with vehicle, were treated during 6 weeks with pyrazole 10 (Pyr10), a TRPC3 inhibitor delivered via mini-osmotic pumps. rAAV-MD gene transfer was confirmed via digital PCR and micro-dystrophin expression via Western blot, while Pyr10 delivery was validated using mass-spectrometry. Finally, using Mn²⁺-quenching of fura-2 fluorescence, we found that Pyr10 and rAAV-MD treatments individually reduced Ca²⁺ entry across DMDmdx muscle cell membrane, which is an encouraging outcome. However, no synergistic effect was observed, and neither treatment improved muscle strength or DMD histological features. As calcium dysregulation is an early and critical event in DMD progression, these results underscore the need for further studies to fully evaluate the therapeutic potential of TRPC3 inhibition for DMD.