Antithrombin III Deficiency in Indian Patients with Deep Vein Thrombosis: Identification of First India Based AT Variants Including a Novel Point Mutation (T280A) that Leads to Aggregation

Abstract

Antithrombin III (AT) is the main inhibitor of blood coagulation proteases like thrombin and factor Xa. In this study we report the identification and characterization of several variants of AT for the first time in Indian population. We screened 1950 deep vein thrombosis (DVT) patients for AT activity and antigen levels. DNA sequencing was further carried out in patients with low AT activity and/or antigen levels to identify variations in the AT gene. Two families, one with type I and the other with type II AT deficiency were identified. Three members of family I showed an increase in the coagulation rates and recurrent thrombosis in this family was solely attributed to the rs2227589 polymorphism. Four members of family II spanning two generations had normal antigen levels and decreased AT activity. A novel single nucleotide insertion, g.13362_13363insA in this family in addition to g.2603T>C (p.R47C) mutation were identified. AT purified from patient’s plasma on hi-trap heparin column showed a marked decrease in heparin affinity and thrombin inhibition rates. Western blot analysis showed the presence of aggregated AT. We also report a novel point mutation at position g.7549 A>G (p.T280A), that is highly conserved in serpin family. Variant protein isolated from patient plasma indicated loss of regulatory function due to in-vivo polymerization. In conclusion this is the first report of AT mutations in SERPINC1 gene in Indo-Aryan population where a novel point mutation p.T280A and a novel single nucleotide insertion g.13362_13363insA are reported in addition to known variants like p.R47C, p.C4-X and polymorphisms of rs2227598, PstI and DdeI.

Association of SERPINC1 Gene Polymorphism (rs2227589) With Pulmonary Embolism Risk in a Chinese Population

“…Moreover, rs2227589 with haplotype in the SERPINC1 gene of FV Leiden carriers showed significantly decreased antithrombin levels (Segers et al, 2014). However, another familial study showed that both rs2227589 carriers and noncarriers have low antithrombin levels (Bhakuni et al, 2015). Thus, variations located in the coding regions of SERPINC1 but not the rs2227589 (C > T) located in the intron may cause antithrombin deficiency.…”

“…Previous study of normal Spanish Caucasian showed a functional effect of the rs2227589 on antithrombin levels (Anton et al, 2009). The findings regarding the association between rs2227589 and antithrombin levels were inconsistent among different studies (Segers et al, 2014; Bhakuni et al, 2015). Furthermore, the association between rs2227589 and the risk of VTE in Northern European, American, and Norwegian populations is inconsistent (Tregouet et al, 2009; Austin et al, 2011; Dahm et al, 2012; Jiang et al, 2017; Kajuna et al, 2018).…”

The relationship between gene polymorphism of MTRR A66G and lower extremity deep venous thrombosis

“…The gene contains 15 exons, ranging from 43 bp to 1213 bp in size and 14 introns ranging from 180 bp to 5 kb in size. The human coding sequence contains 2094 base pairs encoding 698 amino acid peptides and methionine synthase (MS) plays an important role in maintaining intracellular levels of methionine, tetrahydrofolate and homocysteine at appropriate levels [26]. If the MTRR gene mutates, the level of homocysteine in plasma may be elevated.…”

Anticoagulant SERPINs: Endogenous Regulators of Hemostasis and Thrombosis

“…Type II AT deficiency can be further divided into additional subtypes: type IIa caused by mutations in the thrombin binding site, type IIb caused by mutations in the heparin binding site or type IIc in which mutations near the reactive center loop have pleiotropic effects on both heparin and thrombin interactions ( 56 ). Interestingly, a SERPINC1 variant causing type II AT deficiency has been shown to induce polymerization of AT in the plasma ( 57 ). This mechanism is distinct from other SERPIN family variants described that destabilize protein structure and cause intracellular aggregation and retention.…”