title: Antithrombotic efficacy in the guinea pig of a derivative of human protein C with enhanced activation by thrombin.
abstract: Conversion by alpha-thrombin of the zymogen human protein C (HPC) to activated protein C (aPC) is an important physiologic feedback control mechanism for the coagulation cascade. Although activation of HPC by thrombomodulin-bound thrombin is relatively rapid, activation by free thrombin occurs at a significantly slower rate. Previously, we generated a "hyper-activatable" derivative of HPC (FLIN-Q3) with an increased activation rate by free alpha-thrombin in vitro. In this study, the antithrombotic efficacy of FLIN-Q3 was compared with both native zymogen and aPC in an arteriovenous shunt model of thrombosis in the guinea pig. Recombinant proteins were infused 15 minutes before and throughout a 15-minute period while blood was circulated from carotid to jugular through tubing that enclosed a thread on which fibrin was deposited. Parallel dose-dependent antithrombotic responses were observed. Under these non-steady-state conditions, the calculated infusion doses associated with a 50% reduction of thrombus mass were 2.7, 24, and 250 mg/kg/h for aPC, FLIN-Q3, and HPC, respectively. Thrombus weight correlated inversely with plasma concentration of aPC, measured amidolytically, from either direct infusion of aPC or that generated from the zymogens in the animal, and similarly correlated inversely with anticoagulant activity measured by whole blood aPTT. Neither zymogen form showed significant aPC activity before shunt circulation, suggesting a requirement for exposure to thrombin. After the infusion was discontinued for 15 minutes, a second period of thrombus formation in the shunt demonstrated the ability of zymogen forms of PC, unlike aPC, to provide "on-demand" anticoagulant responses to repeated thrombotic stimuli. Thus, a "hyper-activatable" PC molecule such as FLIN-Q3 may represent a superior form of anticoagulant therapy than either the native zymogen or aPC.
authors.affiliation: Division of Cardiovascular Pharmacology, Lilly Research Laboratories, Indianapolis, IN, USA.
authors.forename: K D
authors.initials: KD
authors.lastname: Kurz
authors.forename: T
authors.initials: T
authors.lastname: Smith
authors.forename: A
authors.initials: A
authors.lastname: Wilson
authors.forename: B
authors.initials: B
authors.lastname: Gerlitz
authors.forename: M A
authors.initials: MA
authors.lastname: Richardson
authors.forename: B W
authors.initials: BW
authors.lastname: Grinnell
mesh_terms: D000818:Animals; D000925:Anticoagulants / Q000008:administration & dosage*; D006168:Guinea Pigs; D006801:Humans; D008297:Male; D011486:Protein C / Q000008:administration & dosage* / Q000031:analogs & derivatives / Q000235:genetics; D011994:Recombinant Proteins / Q000008:administration & dosage / Q000235:genetics; D013917:Thrombin / Q000378:metabolism*; D013927:Thrombosis / Q000188:drug therapy* / Q000378:metabolism
chemical_list: D000925:Anticoagulants; D011486:Protein C; D011994:Recombinant Proteins; D013917:Thrombin