Protein C deficiency with recurrent systemic thrombosis associated with compound heterozygous PROC missense variants

Abstract

Herein, we identified compound heterozygous PROC missense variants in a protein C deficient patient with recurrent thrombotic events, including intestinal necrosis, extrahepatic portal vein obstruction, and lower limb venous thrombosis. The patient's protein C activity and antigen levels were extremely low (<10 % and 5 %, respectively). Exome sequencing analysis revealed two rare missense variants (c.76G>A:p.Val26Met in exon 3 and c.1000G>A:p.Gly334Ser in exon 9), both confirmed to be associated with protein C deficiency and one synonymous variant (c.423G>T:p.Ser141Ser in exon 6) in PROC. PCR amplification of genomic DNA spanning these exons followed by Sanger sequencing analysis revealed that the c.76G>A and the synonymous c.423G>T variants were in the same allele, whereas the c.1000G>A variant was on the opposite allele, indicating compound heterozygosity. Western blot analysis of Huh-7 and HEK293T cells transfected with expression vectors encoding PROC with or without these variants demonstrated that Gly334Ser-PROC expression levels were significantly decreased in culture media collected from HEK293T cells, while the expression levels of protein C with these variants were not significantly altered in cell lysates. This suggests that these variants may affect both protein activity and the secretory process of protein C.