Snake venom defensin peptide mCadam10_MYO-2 attenuates collagen-driven platelet activation and arterial thrombosis via GPVI-associated signaling

Arterial thrombosis is a principal driver of ischemic cardiovascular and cerebrovascular disease, yet clinical antiplatelet therapy remains constrained by hemorrhagic risk. Glycoprotein VI (GPVI), a platelet collagen receptor, offers a promising hemostasis-sparing target for antithrombotic intervention. However, peptide-based scaffolds capable of modulating GPVI-dependent platelet activation remain insufficiently explored. Given the diverse hemostasis-regulatory repertoire of snake venoms, this study identified three snake venom defensin and defensin-like peptides, mCadam10_MYO-2 (from Crotalus adamanteus), DACU-1 (from Deinagkistrodon acutus), and DACU-2 (from Deinagkistrodon acutus), and further examined their antiplatelet activities. Comparative sequence and structural analyses revealed a conserved cysteine framework and a β-defensin-like scaffold. Among the three peptides, mCadam10_MYO-2 produced the most potent inhibitory effect on collagen-induced platelet aggregation. Furthermore, mCadam10_MYO-2 reduced platelet spreading on collagen and delayed clot retraction, indicating broad suppression of collagen-driven adhesive and contractile platelet responses. Mechanistic analyses linked these effects to attenuated GPVI-associated signaling, including altered activation-related RNA signatures, reduced collagen-induced Syk and PLCγ2 phosphorylation, and direct binding to recombinant GPVI. Additionally, mCadam10_MYO-2 markedly prolonged carotid artery occlusion time in an electrical injury-induced thrombosis model, but did not significantly prolong tail bleeding time or alter platelet indices under the tested conditions. Collectively, these findings show that mCadam10_MYO-2 suppresses GPVI-associated platelet activation and thrombosis, supporting its potential as a molecular scaffold for hemostasis-sparing antithrombotic development.