Protein C deficiency; PROC gene variants in a Danish population

Highlights

• PROC gene variants were identified in a Danish population by sequencing (N = 46).

• Variants were identified in 20/30 participants diagnosed with protein C deficiency.

• Ten different single nucleotide variants were detected; five variants were novel.

• An association between biochemical severity and the presence of a variant was found.

• Thromboembolic risk did not differ between participants with and without a variant.

Abstract

Introduction

Protein C deficiency is a heritable thrombophilia caused by numerous different genetic alterations in the protein C (PROC) gene. We aimed to identify variants causing protein C deficiency in a Danish population.

Material and methods

Sanger sequencing of the PROC gene was performed in 20 probands and 26 relatives. In total, 30participants were previously diagnosed with protein C deficiency. Protein C activity was measured by a chromogenic substrate method (N = 40) and antigen level by an enzyme-linked immunosorbent assay (N = 26).

Results

Ten different single nucleotide variants were detected in 13 probands (65%) and in seven of the relatives previously diagnosed with protein C deficiency. Five variants were novel. The median protein C activity level was lower in participants with an identified variant (50% (range: 38–75%)) than in protein C deficient participants without a variant (65% (range: 36–73%); P = 0.18). A protein C activity of 57% resulted in the highest detection rate (12/13 (92%)). Likewise, the median antigen level was lower in participants with detectable variants than in participants without (49% (range: 35–99%) vs 70% (range: 41–101%); P = 0.09). No difference was found in venous thromboembolism (VTE) prevalence comparing participants with (12/20 (60%)) and without (7/10 (70%)) a variant (P = 0.59).

Conclusion

In a Danish population, a PROC gene variant was identified in 67% of participants previously diagnosed with protein C deficiency. Five variants were novel. The study confirmed an association between biochemical severity and the presence of a PROC gene variant. The VTE risk did not seem to differ between protein C deficient participants with and without a variant.

Introduction

Protein C, a vitamin K-dependent glycoprotein, is one of the natural anticoagulants and thereby plays an important role in the inhibition of blood coagulation and thrombosis. Acquired low levels of protein C are relatively common as it can be induced by several factors including liver disease [1], treatment with K-vitamin antagonists [2], severe infections [3], use of oral contraceptives [4] and disseminated intravascular coagulation (DIC) [5]. In contrast to acquired protein C deficiency, inherited protein C deficiency is an infrequent autosomal dominant disorder with a prevalence estimated to be around 0.2–0.3% [6,7].

The gene encoding protein C (PROC) is located on chromosome 2q13-14 and includes nine exons, including one non-translated exon, and eight introns [8]. Several different genetic variants causing protein C deficiency have been identified with missense and nonsense variants being the most frequent [9]. Homozygous and compound heterozygous variants in the gene are extremely rare and cause life-threatening disorders as neonatal purpura fulminans or DIC [10]. In contrast, heterozygous protein C deficiency is more common and leads to a four to six fold increased risk for venous thromboembolism (VTE) as well as an increased risk of recurrent thrombosis [[11], [12], [13]]. Adult heterozygous carriers are therefore recommended to receive thromboprophylaxis in risk situation as immobilization, surgery and pregnancy to prevent thromboembolic disease and are recommended life-long anticoagulation after the first thrombotic episode has occurred to prevent recurrent thrombosis [14].

Protein C deficiency is diagnosed based on the protein C plasma activity and antigen level. Heterozygous protein C deficiency can be divided in two different types based on the defect in the protein C molecule. Type I is a quantitative defect of protein C causing both low antigen level and reduced activity and is the most frequent of the two types (75–80%) [7]. Type II is a qualitative defect with normal levels of antigen but with reduced protein C activity [7]. Yet, the diagnosis of protein C deficiency is challenging as there is a substantial overlap in the plasma based assays between individuals with heterozygote protein C deficiency and healthy individuals [15]. Thus, genetic analysis could be of value in the diagnosis of heterozygous protein C deficiency. However, no clear clinical consensus exists for which individuals a genetic evaluation is of clinical value or when genetic analysis is redundant.

In order to elucidate the diagnostic value of the genetic analysis, we here identified genetic variants in the PROC gene in Danish families diagnosed with protein C deficiency and investigated the association between the genotype and the protein C activity and antigen levels as well as clinical characteristics.