Anticoagulant therapy is commonly used to prevent and treat arterial and venous blood clots in patients with cardiovascular disease, cerebrovascular disease, and cancer. Venous blood clots are the third leading cause of cardiovascular death following acute coronary artery disease and stroke. There is a significant need for effective anticoagulant therapy with minimal risk of bleeding. Variegin and its variants are a new type of antithrombin peptide that has shown promising results in preclinical studies. Variegin and its best variant, ultravariegin (UV), can more effectively inhibit blood clot formation while causing less bleeding than traditional medications such as heparin and bivalirudin. However, the short lifespan of UV remains a limitation for its use in clinical settings. PEGylation, a method of conjugating poly(ethylene glycol) (PEG) chains to peptides or drugs, may help improve the effectiveness of UV by extending its circulation time in the body. In this study, UV was PEGylated using maleimide-PEG5k and 10k. The impact of PEGylation on the antithrombin activity of UV was assessed in vitro and ex vivo in rat and rabbit plasma, showing minimal effects on the efficacy. In vivo studies in rats and rabbits revealed that PEGylated UV had a longer half-life and greater anticoagulant effects than unmodified UV did, especially when it was administered subcutaneously. PEGylation significantly extended the half-life of UV in rabbits, resulting in sustained anticoagulant effects for up to 4 days. This demonstrated that increasing the size of UV and shielding it with PEG could reduce clearance by the kidneys and prolong its circulation time. The improved half-life and antithrombin activity of PEGylated UV make it a more favorable choice for anticoagulant therapy.
Keywords: PEGylation; anticoagulant peptide; antithrombin efficacy; in vivo circulation; thrombin inhibitor; ultravariegin.