A NEW ADAMATS13 MISSENSE MUTATION (A4085T) IN THROMBOTIC THROMBOCYTOPENIC PURPURA (TTP) DIAGNOSED IN PREGNANCY

CALDERAZZO JC; KEMPFER AC; POWAZNIAK Y; LOPEZ IR; SÁNCHEZ LUCEROS A; LAZZARI MA

KYOTO

Congreso; XXIII CONGRESS OF INTERNATIONAL SOCIETY ON THROMBOSIS AND HAEMOSTASIS; 2011

INTERNATIONAL SOCIETY ON THROMBOSIS AND HAEMOSTASIS

TTP is a microvascular thrombotic disease, associated with ADAMTS13 deficiency which may be caused by mutations. Currently the relevance of the CUB domains related to ADAMTS13 activity is not clear. Objective: To identify ADAMTS13 mutations in an adult woman that had TTP precipitated only by pregnancies. We studied the effect of the mutation in biosynthesis and enzymatic activity by means of expression studies in mammalian cells. Materials and Methods: Patient with TTP episodes during three pregnancies. Plasma samples were measured for ADAMTS13 activity (ac) and antigen (ag), IgG anti-ADAMTS13 antibody and VWF multimers. Mutation was carried out by screening sequencing of the 29 exons of ADAMTS13 gene. The wild type (WT) and the ADAMTS13 mutant (site-directed mutagenesis) construct were transiently expressed in HEK293 cells. Culture medium and cell lysate were evaluated by ELISA kit for ADAMTS13 ac and ag levels and by SDS/PAGE assay. Results: One year after the first pregnancy, she had 20% of ADAMTS13 ac [normal value (nv) = 46–184%] and 31% ULVWF multimers (nv < 15%), whereas no IgG anti-ADAMTS13 antibody was detectable. The ADAMTS13 ag was 2% (nv = 40–130%) at the 24 weeks of the third pregnant. To exclude that the A4085T mutation found in our patient did not represent disease-related polymorphisms, we screened 50 healthy donors. In vitro expression of ADAMTS13 mutant led to a defect of secretion (ag = 8 SD3%) causing intracellular accumulation (ag =128 SD 13%, ac = 133 SD 5%) which was fully active compared to WT (ag/ac = 100%). Conclusions: In a moderate deficiency of ADAMTS13, pregnancy appears to be crucial in triggering TTP due to the ADAMTS13 physiological fall which started in the second trimester. The results show that A4085T induces a plasmatic ADAMTS13 deficiency that is likely caused by reduced secretion of the protease to circulation. The data confirmed that the second CUB domains are critical for ADAMTS13 secretion. DEVELOPING WORLD SCIENTISTS AWARD