Characterization of two novel mutations of the antithrombin gene observed in Japanese thrombophilic patients.

We investigated the molecular basis of reduced functional levels of antithrombin (AT) in two individuals suffering from thromboembolic events. In each case direct sequencing of amplified DNA revealed 13,260-13,262 del in one patient and 2511C>A in the other patient, predicting a heterozygous E381del and P16H, respectively. Both patients had no 20210A allele and factor V Leiden mutation. To understand the molecular mechanism responsible for antithrombin deficiency, stable expression experiments were performed using HEK293 cells transfected with the expression vector containing the wild-type or the mutated recombinant cDNA. In these experiments, the media levels of the two mutated antithrombins were the same as that of wild type, but the specific activity of the E381del mutant decreased significantly compared with that of wild type. These results showed that the E381del mutation was responsible for type II deficiency, whereas the other mutation, P16H, did not produce any definite abnormality which could contribute to antithrombin deficiency.