The first mutation identified in the thrombomodulin gene in a 45-year- old man presenting with thromboembolic disease

Abstract

Thrombomodulin (TM) is the anticoagulant endothelial cell membrane- bound protein cofactor in the thrombin-mediated activation of protein C (PC). It has been clearly demonstrated that the anticoagulant and profibrinolytic functions of the PC system are important for the prevention of a thromboembolic disease. Patients with PC, protein S, or PC “‘cofactor”’ deficiency and/or dysfunction develop thromboembolic diseases. However, the molecular abnormality in at least 20% to 30% of thrombophilic patients cannot be identified by hitherto recognized defects. A putative pathologic lesion in the TM gene could be one of several candidates for these prothrombotic mutations. A directed search strategy for deletions, insertions, or point mutations in the TM gene has not been performed. Therefore, in the present study, we have analyzed the entire TM gene, including the promoter region, by polymerase chain reaction-single-strand conformation polymorphism (PCR- SSCP) in normal healthy volunteers and in patients presenting with a thromboembolic disease. We have identified a patient with a thromboembolic disease and a TM point mutation. In a 45-year-old Hispanic man with a documented pulmonary embolism, PCR-SSCP showed an aberrant band pattern and subsequent DNA sequence analysis showed a heterozygous substitution for G1456 to T. This substitution predicts an Asp468 to a Tyr change in the amino acid sequence that is located between the transmembrane domain and the sixth epidermal growth factor- like domain. The Asp468 to Tyr change would probably lead to significant structural changes not allowing the expression of the TM protein or to a conformational change that is not functional.

Mutations in the thrombomodulin gene are rare in patients with severe thrombophilia

“…Methods. The numbering of nucleotides was according to Ö hlin & Marlar (1995). The gene was divided into eight overlapping fragments as reported in Fig 1. The TM gene fragments were sequenced by an automated sequencer (ABIPrism, Applied Biosystems, Foster City, CA, USA) after amplification.…”

Prospective study of the A455V polymorphism in the thrombomodulin gene, plasma thrombomodulin, and incidence of venous thromboembolism: the LITE Study

“…It has been reported previously that the frequency of TM muta-tions among patients with thromboembolic disease is approximately 5% [7]. Thrombotic disorders have been found to result from relative deficiency of TM [8] and homozygous deficiency of TM has been shown to confer lethality in mice [9].…”

“…The human TM gene does not contain introns [10]. Several polymorphisms or mutations in the coding and promoter regions of the TM gene have been identified [7,8,[11][12][13]. The influence of these polymorphisms on the level or activity of TM is unclear.…”

Thrombomodulin with the Asp468Tyr mutation is expressed on the cell surface with normal cofactor activity for protein C activation

“…TM has been proposed to produce the soluble plasma TM through proteolytic activity generated on the cell surface (Ishii et al, 1991). The plasma TM level of the patient with Asp468Tyr was the lowest among patients with thrombotic disorders (O È hlin & Marlar, 1995), whereas no correlation was found between incidence of venous thrombosis and low plasma soluble TM levels (Faioni et al, 1998;O È hlin & Marlar, 1999). In our study, even more TM antigen was detected in the culture media of variant TM transformants than of normal TM ones.…”