Abstract and Introduction
Abstract
The diagnosis of thrombophilia caused by protein S deficiency remains difficult. From 2005 to 2010, we documented 135 patients with suspected hereditary protein S deficiency for whom mutational analysis of the PROS1 gene had been performed by direct double-stranded sequencing of the amplified 15 exons including splice sites. Multiplex ligation-dependent probe amplification was performed on 12 of 15 exons in cases with no mutation found but a large deletion in the PROS1 gene was suspected. Mutations were identified in 49 patients, 9 by familial screening. Altogether, 17 new and 11 previously described mutations of PROS1 were identified among the 49 patients. After the exclusion of acquired protein S deficiency due to pregnancy or hormonal contraceptives, there remained only 1 case with protein S activity levels less than 40% that could not be explained by sequence variations or deletions in the examined regions of the PROS1 gene. After the exclusion of conditions associated with acquired protein S deficiency, persistently low protein S activity levels are highly indicative of a genetic alteration in PROS1. We observed a clear correlation between the laboratory phenotype and the type of mutation.
Introduction
Protein S (gene symbol, PROS1; GeneID, 5627; MIM No. 176880) was first described in 1979 by DiScipio and Davie as a glycoprotein containing γ-carboxylated glutamic acid residues similar to other proteins involved in coagulation processes. In contrast with other vitamin K–dependent clotting factors, protein S, which was named after the place of its first isolation, Seattle, WA, lacks serine protease activity. Further studies demonstrated that it has cofactor function for activated protein C. Under normal conditions, more than 60% of protein S is bound to the C4b binding protein, and unbound protein S is available to form a complex with activated protein C in the presence of phospholipids and calcium ions. This complex irreversibly inactivates factors Va and VIIIa by proteolysis, thus interfering with the formation of the prothrombinase and tenase complexes on procoagulant surfaces. Thrombin inactivates protein S by proteolytic cleavage.
Two highly homologous genes are located near the centromeric region of chromosome 3, at 3p11.1–3q11.2: one is the active, encoding gene PROS1, and the other, PROS2 or PROSP, is, in all likelihood, a pseudogene with no open reading frame. PROS2 lacks exon 1, which contains the translational start site and encodes for a signal peptide. Furthermore, there are several destructive mutations contained in PROS2.
Owing to the cofactor function of protein S, the laboratory phenotype is difficult to assess. Typically, measurement is performed using a latex ligand immunoassay for the determination of free antigen and using clotting assays that detect protein S–mediated enhancement of the anticoagulant function of activated protein C. In addition to congenital deficiencies, there are numerous physiologic and pathologic conditions that lead to decreased protein S levels. Inflammation leads to increased C4b binding protein levels and concomitantly decreases the levels of unbound and functional protein S. Pregnancy and hepatic disorders, such as hepatitis and cirrhosis, and treatment with hormonal contraceptives and the lingering effect of previous anticoagulation similarly lead to decreased levels.