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(Arteriosclerosis, Thrombosis, and Vascular Biology. 1995;15:2181-2187.)
© 1995 American Heart Association, Inc.

Articles

Biochemical Prototype for Familial Thrombosis

A Study Combining a Functional Protein C Mutation and Factor V Leiden

Michael Kalafatis; Deshun Lu; Rogier M. Bertina; George L. Long; Kenneth G. Mann

From the Department of Biochemistry, University of Vermont, College of Medicine, Burlington (M.K., D.L., G.L.L., K.G.M.), and Hemostasis and Thrombosis Research Center, University Hospital, Leiden, Netherlands (R.M.B.).

Correspondence to Kenneth G. Mann, PhD, Department of Biochemistry, Given Building, Health Science Complex, University of Vermont, College of Medicine, Burlington, VT 05405-0068.

Abstract Resistance to activated protein C (APC) is associated with a single amino acid substitution in factor V (Arg⁵⁰⁶Gln, factor V Leiden) that results in delayed inactivation of the molecule by APC. The mutation is present in 20% of patients with a first episode of deep venous thrombosis. Arterial and venous thromboses are also associated with the type II protein C deficiency (protein C_Vermont). In protein C_Vermont, the substitution Glu²⁰Ala alone (rPC^20A) is responsible for the defective anticoagulant properties of PC_Vermont. It was recently established that a thrombotic episode occurred in 73% of family members who are heterozygous for both a functional protein C gene mutation and the factor V Leiden mutation. We evaluated the molecular defect that would accrue in the combined deficiency state of factor V^R506Q/Va^R506Q and rAPC^20A using recombinant APC and natural purified factor V^R506Q from patients homozygous for the Arg⁵⁰⁶Gln substitution. While wild-type recombinant APC (rAPC) slowly cleaves and inactivates factor V^R506Q and factor Va^R506Q, minimal cleavage of membrane-bound factor V^R506Q and Va^R506Q by rAPC^20A at Arg³⁰⁶ and Arg⁶⁷⁹ occurs, and no loss in cofactor activity is observed. Our data demonstrate that rAPC^20A cannot inactivate either factor V^R506Q or factor Va^R506Q at biologically relevant rates because of impaired cleavage at Arg³⁰⁶ and Arg⁶⁷⁹. The result is a stable procofactor and stabilization of an active cofactor in patients possessing both mutations. Our data provide a prototype of familial thrombosis that most likely would be manifested in vivo by the occurrence of massive thrombosis.

Key Words: thrombosis • factor V Leiden • protein C_Vermont • blood coagulation • APC resistance

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