Abstract

The clinical application of peptide therapeutics is often constrained by their short plasma half-life, necessitating frequent administration and resulting in undesirable pharmacokinetic fluctuations and side effects. Here, we developed a genetically encodable multivalent fusion protein platform that combines haemadin, a selective thrombin inhibitor derived from terrestrial leeches, with an elastin-like polypeptide (ELP) partner via factor Xa (FXa)-cleavable peptide linkers. This platform enables tunable drug loading, long-term release, and stimuli-responsive activation of antithrombotic activity. After subcutaneous injection, the fusion protein undergoes temperature-triggered ELP condensation to form an in situ depot that slowly releases an inactivated prodrug, which remains inert in circulation until thrombus-associated FXa cleave the linker to liberate active haemadin on demand. We achieved high-yield expression and facile nonchromatographic purification of the fusion protein. Subcutaneous administration resulted in significant prolongation of antithrombotic protection postinjection. This approach holds strong potential to enhance the safety, efficacy, and dosing convenience of peptide therapy.