Background From 20 to 50% of patients who survive an acute episode of the acquired form of thrombotic thrombocytopenic purpura relapse but clinical and laboratory markers of recurrence are not well established. Design and MethodsIn 109 patients enrolled in an international registry we evaluated, in the frame of a retrospective cohort study, the predictive role of the metalloprotease ADAMTS13 as measured in plasma during remission. Anti-ADAMTS13 antibodies and von Willebrand factor were also evaluated in a smaller number of the same patients. ResultsMedian values of ADAMTS13 activity and antigen were significantly lower in patients with recurrent thrombotic thrombocytopenic purpura than in those with no recurrence (activity: 12% vs. 41%; p=0.007; antigen: 36% vs. 58%; p=0.003). A severe deficiency of ADAMTS13 activity (10% or less) was associated with a higher likelihood of recurrence (odds ratio 2.9; 95% confidence interval 1.3 to 6.8; p=0.01). Anti-ADAMTS13 antibodies were also more prevalent in patients with recurrent thrombotic thrombocytopenic purpura (odds ratio 3.1; 95% confidence interval 1.4 to 7.3; p=0.006). The presence during remission of both severe ADAMTS13 deficiency and anti-ADAMTS13 antibodies increased the likelihood of recurrence 3.6 times (95% confidence interval 1.4 to 9.0; p=0.006). The presence of ultralarge von Willebrand factor multimers and of associated diseases or conditions did not increase recurrence. ConclusionsSurvivors of an acute episode of acquired thrombotic thrombocytopenic purpura with severely reduced levels of ADAMTS13 and/or with anti-ADAMTS13 antibodies during remission have an approximately three-fold greater likelihood of developing another episode of thrombotic thrombocytopenic purpura than patients with higher protease activity and no antibody.Key words: thrombotic thrombocytopenic purpura, ADAMTS13, von Willebrand factor, risk factors, recurrence.Citation: Peyvandi F, Lavoretano S, Palla R, Feys H B., Vanhoorelbeke K, Battaglioli T, Valsecchi C, Canciani MT, Fabris F, Zver S, Réti M, Mikovic D, Karimi M, Giuffrida G, Laurenti L, antibodies as markers for recurrence of acquired thrombotic thrombocytopenic purpura during remission. Haematologica 2008 Feb; 93(2):232-239. DOI: 10.3324/haematol.11739 ©2008 Ferrata Storti Foundation. This is an open-access paper. ADAMTS13 and anti-ADAMTS13 antibodies as markers for recurrence of acquired thrombotic thrombocytopenic purpura during remission IntroductionAcquired thrombotic thrombocytopenic purpura (TTP) is a rare but severe disease characterized by microangiopathic hemolytic anemia and consumptive thrombocytopenia leading to disseminated microvascular thrombosis that causes variable signs and symptoms of organ ischemia and damage.1 The pathophysiology of TTP has become clearer after it was demonstrated that the plasma metalloprotease ADAMTS13 (a disintegrin and metalloprotease with thrombospondin 1 repeats) cleaves the platelet-adhesive plasma protein von Willebrand factor (VWF) at the peptide bond Tyr1605-Met1...

SummaryThe congenital or acquired deficiency of the von Willebrand factor (VWF) cleaving protease, ADAMTS-13 has been specifically associated with a diagnosis of thrombotic thrombocytopenic purpura (TTP), a microangiopathy characterized by the formation of occlusive platelet thrombi. The mechanisms of TTP were investigated in 100 patients diagnosed on the basis of the presence of at least three of the following: thrombocytopenia, haemolytic anaemia, elevated serum levels of lactate dehydrogenase and neurological symptoms. Plasma levels of ADAMTS-13 were severely reduced (<10% of normal) in 48%, moderately reduced (between 10% and 46%) in 24% and normal (>46%) in 28%. A neutralizing antibody was the cause of the deficiency in 38% of the cases, with a higher prevalence of this mechanism (87%) in the 48 patients with severely reduced ADAMTS-13. Double heterozygosity for a 29 base pair (bp) deletion and a nucleotide insertion and homozygosity for a 6 bp deletion in the ADAMTS13 gene were identified only in two patients born from consanguineous marriages. In conclusion, this study indicated that ADAMTS-13 was normal in nearly one-third of patients with TTP and that ADAMTS-13 deficiency was not associated with the presence of neutralizing antibodies in more than half of the patients.

Inherited deficiencies of plasma proteins involved in blood coagulation generally lead to lifelong bleeding disorders, whose severity is inversely proportional to the degree of factor deficiency. Haemophilia A and B, inherited as X-linked recessive traits, are the most common hereditary hemorrhagic disorders caused by a deficiency or dysfunction of blood coagulation factor VIII (FVIII) and factor IX (FIX). Together with von Willebrand's disease, a defect of primary haemostasis, these X-linked disorders include 95% to 97% of all the inherited deficiencies of coagulation factors. The remaining defects, generally transmitted as autosomal recessive traits, are rare with prevalence of the presumably homozygous forms in the general population of 1:500,000 for FVII deficiency and 1 in 2 million for prothrombin (FII) and factor XIII (FXIII) deficiency. Molecular characterization, carrier detection and prenatal diagnosis remain the key steps for the prevention of the birth of children affected by coagulation disorders in developing countries, where patients with these deficiencies rarely live beyond childhood and where management is still largely inadequate. These characterizations are possible by direct or indirect genetic analysis of genes involved in these diseases, and the choice of the strategy depends on the effective available budget and facilities to achieve a large benefit. In countries with more advanced molecular facilities and higher budget resources, the most appropriate choice in general is a direct strategy for mutation detection. However, in countries with limited facilities and low budget resources, carrier detection and prenatal diagnosis are usually performed by linkage analysis with genetic markers. This article reviews the genetic diagnosis of haemophilia, genetics and inhibitor development, genetics of von Willebrand's disease and of rare bleeding disorders.

The degradation of von Willebrand factor (VWF) depends on the activity of a zinc protease (referred to as ADAMTS-13), which cleaves VWF at the Tyr 1605 -Met 1606 peptide bond. Little information is available on the physiological mechanisms involved in regulation of AD-AMTS-13 activity. In this study, the role of ions on the ADAMTS-13/VWF interaction was investigated. In the presence of 1.5 M urea, the protease cleaved multimeric VWF in the absence of NaCl at pH 8.00 and 37°C, with an apparent k cat /K m Х 3.4 ؋ 10 4 M ؊1 s ؊1 , but this value decreased by ϳ10-fold in the presence of 0.15 M NaCl. Using several monovalent salts, the inhibitory effect was attributed mostly to anions, whose potency was inversely related to the corresponding Jones-Dole viscosity B coefficients (ClO 4 ؊ > Cl ؊ > F ؊ ). The specific inhibitory effect of anions was due to their binding to VWF, which caused a conformational change responsible for quenching the intrinsic fluorescence of the protein and reducing tyrosine exposition to bulk solvent. Ristocetin binding to VWF could reduce the apparent affinity and reverse the inhibitory effect of chloride. We hypothesize that, after secretion into the extracellular compartment, VWF is bound by chloride ions abundantly present in this milieu, becoming unavailable to proteolysis by AD-AMTS-13. Shear forces, which facilitate GpIb␣ binding (this effect being artificially obtained by ristocetin), can reverse the inhibitory effect of chloride, whose concentration gradient across the cell membrane may represent a simple but efficient strategy to regulate the enzymatic activity of ADAMTS-13.