Clinical Course Of Thrombotic Thrombocytopenic Purpura Research Articles

Cell Derived Microparticles (MP) in Different Phases of Thrombotic Thrombocytopenic Purpura (TTP) and Effect of Exchange Plasmapheresis (EPP) on Their Profiles

BACKGROUND: Cell derived MPs are small membrane vesicles released during cell activation or apoptosis. They express an inside-out membrane, which exposes the negatively charged phospholipid layer outside, allowing clotting factors to be anchored and generate thrombin. Among various species of MPs, RMP (red cell MPs), PMP (platelet MPs), LMP (leukocytes), EMP (endothelial cells) are of special interest. They play an important role in hemostasis, thromboses, and inflammation. MPs mirror early injury of parent cells and are sensitive early biomarkers of underlying disorders.TTP is a microangiopathy mediated by antibody-induced depletion of ADAMTS13, a von Willebrand factor-cleaving protease. Endothelial injury promotes platelet clumping and formation of platelet rich microthrombi in the microcirculation. Subsequent platelet sequestration leads to impaired microcirculation, thrombocytopenia and red cell fragmentation with a microangiopathic hemolytic anemia. Exchange plasmapheresis (EPP) is the standard therapy. It removes antibodies to ADAMTS13, replacing it with ADAMTS13 rich plasma. It is possible that EPP removes thrombogenic MPs to improve the clinical course of TTP. In this study, we investigated MP profiles in active and remission phase of TTP and the effect of EPP on MP profiles. We also aimed to determine if MP profiles may be a useful measure for monitoring clinical course and tracking progress of therapy.METHODS: A retrospective study was conducted evaluating MP assays in patients with TTP. MP profiles were reviewed in acute and remission phases of TTP. Acute phase was defined as thrombocytopenia, clinical evidence of microangiopathy and hemolytic anemia and low ADAMTS13 activity. Remission was defined as sustained normalization of laboratory parameters and no further microangiopathy for at least one month. Patients were studied longitudinally, with MP assays before and after EPP. EMP were measured by CD31+/CD42b− (EMP31), CD62E+ (EMP62); PMP by CD31+/CD42b+ (PMP42) and CD41+ (PMP41). All were measured in platelet-poor plasma by flow cytometry. All MP data are presented in units of x105/µL. The differences in MP patterns among TTP patients in active and remission phases of disease, as well as the effect of EPP on MP profiles were assessed.RESULTS: Among 20 patients with TTP, 8 (40%) were in acute phase and 12 (60%) in remission. An average of 10.7 EPP were performed. The average platelet count prior to EPP was 50.6x103/µL, which increased to 248 x103/µL following the last EPP. ADAMTS13 activity was generally <10% at the onset.For patients in the acute phase, PMPs were low: PMP41 0.32 (±0.11), PMP42 3.11 (±2.56), but consistently increased following EPP to the point of statistical significance at the last treatment: PMP41 1.26 (±0.71), p=0.005 and PMP42 7.65 (±5.03), p=0.039. Moreover, levels were higher in the remission phase for both PMPs, but only statistically significant for PMP41: 1.39, p=0.034.Conversely, EMP62E was initially elevated on presentation, but declined with successive EPP: 6.07 (±3.02) prior to initiation of EPP to 4.5 (±3.1) upon the last day of EPP. Furthermore, this pattern continued into remission, with EMP62E of 2.72 (±2.11), p=0.009. Similar to the trend in EMP62E, RMP was elevated in the acute phase (25.1 [±18.5]) before steadily declining with EPP (19.65 [±12.53]). This progressive drop in RMP persisted for those in remission phase of TTP 10 (±5.42), p=0.015.DISCUSSION/CONCLUSION: Taken collectively, cell derived MP's were found to reflect disease activity and response to therapy.There was a linear correlation between PMP levels and platelet count. Low PMPs in the acute phase reflects the initial thrombocytopenia characteristic of TTP. When EPP was initiated, the disease improved and there was a rise in PMP which mirrored the rise in platelets. This rise was sustained in the remission phase.Conversely, EMP62E and RMP are increased in acute phase TTP due to endothelial activation, microthrombi deposition and red cell fragmentation. After EPP the disease is quiescent, hence the progressive decline of EMP62E and RMP.These results show promising utility of cell derived MP profiles as a clinical tool to surveil chronic TTP patients and to predict disease relapse at an early stage. After EPP is initiated, change in MP Profiles may be used to monitor response to therapy and determine the appropriate time to wean EPP. DisclosuresNo relevant conflicts of interest to declare.

The use of therapeutic apheresis in cardiovascular disease

In the last few years, therapeutic apheresis (TA) has emerged as a valuable treatment option for certain life-threatening cardiovascular diseases (CVDs) and for all the cardiac dysfunctions caused by autoimmune or metabolic disorders. Besides the well-established indications for apheresis treatment, such as familial hypercholesterolaemia, hyperviscosity syndrome and thrombotic thrombocytopenic purpura (TTP), we discuss the novel approaches in the therapy of dilated cardiomyopathy, cardiac failure and some specific syndromes of severe dysfunction occurring after heart transplantation. The rationale for using apheresis in such patients is the contribution in immune modulation that this procedure can undoubtedly provide. The clinical course of TTP has dramatically changed, thanks to the introduction of therapeutic plasma exchange. Low-density lipoprotein apheresis has been extremely efficacious, safe and suitable for lowering cholesterol levels and can be used even for long-term treatment. In Waldenström macroglobulinaemia and other hyperviscosity syndromes, plasma exchange has demonstrated to be an efficient tool for reducing blood viscosity and the risk of a consequent cardiac dysfunction. However, TA may induce oxidative injury to erythrocytes, making these cells more prone to haemolysis and causing a significant reduction in their half-life. Ascorbate administration can be useful to lower the levels of hydrogen peroxide and proinflammatory mediators in patients undergoing apheresis. Further data are needed to support this benefit and to test other potential antioxidant therapies. Many of these therapeutic indications need further studies to be definitively approved, but preliminary data are encouraging.

A case of thrombotic thrombocytopenic purpura associated with bupropion

To report a case of bupropion-associated thrombotic thrombocytopenic purpura (TTP) syndrome. A 55-year-old man was admitted with complaints of diarrhea, acute renal failure, and confusion ~ 54 days after bupropion initiation for smoking cessation. Subsequently he had a tonic-clonic seizure and had to be intubated because of altered consciousness. Laboratory findings were compatible with microangiopathic hemolytic anemia. He was diagnosed as TTP for which the Naranjo adverse drug reaction probability scale indicated a probable relationship with bupropion (a score of 5). He was treated with plasma exchange, systemic corticosteroids, hemodialysis and recovered fully. Bupropion is an anti-depressant drug also indicated for smoking cessation. It has widely reported neuropsychiatric and allergic adverse effects; however, TTP associated with bupropion has only been reported once. The clinical course of TTP in this case was compatible with TTP related to acute, immune mediated drug toxicity, which suggests that auto-antibodies might have been responsible. Given the fact that the clinical condition is compatible with acute, immune-mediated TTP syndrome, we suggest bupropion deserves evaluation for auto-antibody induction. Prescribers should be aware of the possible risk of thrombocytopenia and TTP.

Quantitative Analysis of the Domain-Specific Autoantibodies to ADAMTS13 in Patients with Acquired Thrombotic Thrombocytopenic Purpura

Abstract 1167 Background:Cleavage of ultra-large von Willebrand factor (UL-VWF) multimer by ADAMTS13 is an essential process to maintain the appropriate VWF response to platelet in blood flow. The production of autoantibodies to ADAMTS13 causes the accumulation of UL-VWF multimer, resulting in the formation of platelet-rich thrombi in microcirculation, which is considered as a pivotal pathophysiology of acquired thrombotic thrombocytopenic purpura (TTP). Previous studies showed that the major recognition sites of the autoantibodies resided in the cysteine-rich and spacer domains, which include substrate binding exosites. To date, an ELISA kit to measure the total IgG autoantibodies to whole ADAMTS13 molecule is available, and the domain-specific autoantibodies were detected with recombinant truncated proteins using western blot or immunoprecipitation methods. Objective:The aim of this study was to perform a radioimmunoprecipitation assay to quantify the domain-specific autoantibodies to ADAMTS13 in acquired TTP. This approach was expected to allow us the sensitive detection and the precise quantification of these autoantibodies, leading to the understanding of pathophysiology and clinical course of TTP. Materials and methods:Two kinds of 35S-methionine labeled antigens, MDTCS and T2-8/CUB, were prepared using in vitro transcription/translation kit (TNT Quick Coupled Transcription/Translation system, Promega). MDTCS was a peptide from amino acid residue-1 to 687, including metalloprotease, disintegrin-like, TSP1-1, cysteine-rich and spacer domains, containing the substrate binding exosites and the catalytic site. T2-8/CUB was a peptide from residue-686 to 1427, including seven TSP1 repeats and two CUB domains, which are associated with the shear-related function in blood flow. The correct synthesis and molecular size of the two radiolabeled antigens were confirmed with SDS-PAGE. First, to establish the quantitative assay, each of the antigens, MDTCS or T2-8/CUB, was mixed with mouse anti-ADAMTS13 monoclonal antibodies, whose epitopes were already well examined in our previous study (ASH Annual Meeting 2009 114:3182). The immune complex was precipitated with protein G beads and the beads were washed, and then applied in a liquid scintillation counter. Second, each of the antigens was similarly immunoprecipitated and quantified with IgG samples purified from 12 acquired TTP patients. As a control, IgG samples from healthy subjects with no histories of autoimmune disease were used. Results:Each of the radiolabeled antigens was expressed as a single band with expected molecular size with SDS-PAGE and successfully immunoprecipitated with monoclonal anti-ADAMTS13 antibodies corresponding to each epitopes, indicating the intact conformation of the antigens. Furthermore, this assay system showed the appropriate dose-dependent escalation curve according to the addition of the monoclonal antibodies, verifying its quantitative analysis. The titration of IgG sample from TTP patients using each antigen, MDTCS or T2-8/CUB, revealed that all of the samples exhibited significantly higher titers to both of the antigens than the control IgG samples, indicating the reliable sensitivity of this assay. Conclusion:We performed a quantitative analysis of the domain-specific autoantibodies to ADAMTS13 in TTP using the radioimmunoprecipitation assay. This sensitive approach may enable us to clarify the relationship between the epitopes of autoantibodies to ADAMTS13 and the clinical characteristics of TTP. Disclosures:Matsumoto:Alexion Pharma: Membership on an entity's Board of Directors or advisory committees. Fujimura:Baxter BioScience: Membership on an entity's Board of Directors or advisory committees; Alexion Pharma: Membership on an entity's Board of Directors or advisory committees.

TTP disease course is independent of myeloma treatment and response

TTP disease course is independent of myeloma treatment and response

Leukocyte proteases cleave von Willebrand factor at or near the ADAMTS13 cleavage site

AbstractThe function of von Willebrand factor (VWF) is regulated by proteolysis, which limits its multimeric size and ability to tether platelets. The importance of ADAMTS13 metalloprotease in VWF regulation is demonstrated by the association between severe deficiency of ADAMTS13 and thrombotic thrombocytopenic purpura (TTP). However, ADAMTS13 activity levels do not always correlate with the clinical course of TTP, suggesting that other proteases could be important in regulating VWF. We identified 4 leukocyte proteases that cleave the synthetic VWF substrate FRETS-VWF73 and multimeric VWF. Elastase and proteinase 3 (PR3) cleave multimeric VWF and FRETS-VWF73 at the V1607-T1608 peptide bond; cathepsin G and matrix metalloprotease 9 cleave VWF substrates at the Y1605-M1606 and M1606-V1607 bonds, respectively. Isolated intact human neutrophils cleave FRETS-VWF73 at the V1607-T1608 peptide bond, suggesting that elastase or PR3 expressed on leukocyte surfaces might cleave VWF. In the presence of normal or ADAMTS13-deficient plasma, cleavage of FRETS-VWF73 by resting neutrophils is abolished. However, activated neutrophils retain proteolytic activity toward FRETS-VWF73 in the presence of plasma. Although the in vivo relevance remains to be established, these studies suggest the existence of a “hot spot” of VWF proteolysis in the VWF A2 domain, and support the possibility that activated leukocytes may participate in the proteolytic regulation of VWF.

An inquiry into the relationship between ABO blood group and thrombotic thrombocytopenic purpura

ABO blood group accounts for up to 40% of the variability in plasma von Willebrand factor (VWF) levels, which vary in the rank order AB > B > A > O > Bombay. This may be due in part to the influence of ABO-associated oligosaccharides on the proteolysis of VWF by the metalloprotease ADAMTS13, which is markedly deficient in thrombotic thrombocytopenic purpura (TTP). Using ABO blood group as a surrogate for baseline VWF levels as well as susceptibility to proteolysis by ADAMST13, we set out to determine whether ABO blood group influences the clinical course of TTP. We conducted a retrospective analysis of the clinical course of 76 patients with primary, sporadic TTP treated at two institutions over the past 10 years. We found no significant differences between group O and non-O patients with respect to presenting platelet count and lactate dehydrogenase concentration, maximum serum creatinine concentration, and total number of therapeutic plasma exchanges per episode. Substrate-related contributors to the highly variable phenotype and clinical course of TTP warrant further investigation.

An inquiry into the relationship between ABO blood group and thrombotic thrombocytopenic purpura

Background and Objectives ABO blood group accounts for up to 40% of the variability in plasma von Willebrand factor (VWF) levels, which vary in the rank order AB > B > A > O > Bombay. This may be due in part to the influence of ABO-associated oligosaccharides on the proteolysis of VWF by the metalloprotease ADAMTS13, which is markedly deficient in thrombotic thrombocytopenic purpura (TTP). Using ABO blood group as a surrogate for baseline VWF levels as well as susceptibility to proteolysis by ADAMST13, we set out to determine whether ABO blood group influences the clinical course of TTP. Methods We conducted a retrospective analysis of the clinical course of 76 patients with primary, sporadic TTP treated at two institutions over the past 10 years. Results We found no significant differences between group O and non-O patients with respect to presenting platelet count and lactate dehydrogenase concentration, maximum serum creatinine concentration, and total number of therapeutic plasma exchanges per episode. Conclusions Substrate-related contributors to the highly variable phenotype and clinical course of TTP warrant further investigation.

The course of ADAMTS‐13 activity and inhibitor titre in the treatment of thrombotic thrombocytopenic purpura with plasma exchange and vincristine

The therapeutic efficacy of plasma exchange (PE) in thrombotic thrombocytopenic purpura (TTP) is attributed to the restoration in ADAMTS-13 (a disintegrin and metalloproteinase with thrombospondin motif-13) activity by substitution of the enzyme and removal of ADAMTS-13-neutralizing autoantibodies. We explored this rationale by analysing ADAMTS-13 activity and corresponding inhibitor levels during PE-treatment in 27 episodes from 23 adults with TTP. All patients with an initial episode of TTP (n = 14) and nine of 11 patients with a relapse showed severe ADAMTS-13 deficiency. ADAMTS-13 inhibitors were detected in 81% of these patients. Twenty-one patients responded to PE-therapy and two patients died. For patients with severe ADAMTS-13 deficiency, 15 patients (71%) showed a PE-induced recovery in ADAMTS-13 activity and six patients (29%) had persistent severe ADAMTS-13 deficiency despite clinical response. Three patients with recurrent TTP demonstrated a permanent increase in inhibitor titre during therapy. Six patients (43%) with an initial episode of TTP displayed a transient increase in inhibitor titre during PE-therapy, which was associated with deterioration in clinical and haematological symptoms of TTP. Treatment with vincristine induced an immediate increase in platelet count and ADAMTS-13 activity in seven of eight patients. We conclude that ADAMTS-13 activity and inhibitor levels, as measured using current methodology, do not solely determine the clinical course of TTP.

Incidence and clinical course of thrombotic thrombocytopenic purpura due to ticlopidine following coronary stenting. EPISTENT Investigators. Evaluation of Platelet IIb/IIIa Inhibitor for Stenting.

Thrombotic thrombocytopenic purpura (TTP) is a rare and often fatal disorder characterized by thrombocytopenia, microangiopathic hemolytic anemia, mental status changes, and renal dysfunction. Ticlopidine hydrochloride is 1 of several drugs that have been associated with this disorder and is currently used routinely in the approximately 500000 patients per year in the United States who undergo a percutaneous coronary intervention involving a stent. To determine the incidence and describe the clinical course of TTP due to ticlopidine therapy following stenting. Retrospective analysis of cohort of all patients undergoing coronary stenting at the Evaluation of Platelet IIb/IIIa Inhibitor for Stenting (EPISTENT) study sites. Sixty-three centers throughout the United States and Canada. A total of 43322 patients who underwent a percutaneous coronary intervention and received a coronary stent during a 1-year period from 1996 to 1997. Cases of TTP following stenting during the 1-year period to determine the incidence of TTP due to ticlopidine therapy following coronary stenting. Additional cases were collected from these and other centers across North America to further describe the clinical presentation and course of TTP due to ticlopidine therapy following stenting. Nine cases of TTP following stenting were recognized at the 63 centers during the specified period, giving an incidence of 1 case per 4814 patients treated (0.02%; 95% confidence interval, 1 case per 2533 to 1 case per 10 541 patients treated). Ten additional cases of TTP related to ticlopidine therapy following stenting were identified from other centers, were identified from the primary centers outside the pre-defined period, or involved a noncoronary stent. Four patients (21%) received ticlopidine for 2 weeks or fewer, 14 patients (74%) for 2 to 4 weeks, and 1 patient (5%) for 8 weeks. The mean time of ticlopidine treatment prior to TTP diagnosis was 22 days (range, 5-60 days). The overall mortality rate was 21% (4/19), with all 4 deaths occurring in patients not treated with plasmapheresis, whereas there were no deaths among the 13 patients who received plasmapheresis. The findings of a TTP incidence of 0.02% in our cohort of ticlopidine-treated patients following coronary stenting suggests that TTP occurs much more commonly in this population than the estimated incidence of 0.0004% in the general population. The mortality rate for this rare complication exceeds 20%. Limiting ticlopidine therapy to 2 weeks after stenting does not prevent the development of TTP. Rapid diagnosis and treatment that includes plasmapheresis are critical for improved survival.

United States Thrombotic Thrombocytopenic Purpura Apheresis Study Group (US TTP ASG): multicenter survey and retrospective analysis of current efficacy of therapeutic plasma exchange.

Thrombotic thrombocytopenic purpura (TTP) remains enigmatic from the perspective of its etiology, pathophysiology, and treatment. Once recognized, the accepted standard of care for TTP is daily therapeutic plasma exchange (TPE). However, the diversity in TPE treatment protocols has made comparisons of clinical research between institutions difficult. This study strived to assess the current practice of TPE in order to provide direction for prospective controlled clinical trials. Twenty large apheresis centers within the United States comprising the US TTP ASG responded to a survey to establish the current status of TPE in TTP. A retrospective analysis from data provided by 14 of 20 centers included 115 initial presentations of primary TTP with an overall mortality rate of 10% and relapse rate of 37%. The majority of deaths (58%) occurred within 48 hours of presentation. Variation in therapeutic targets (platelet count [plt] and serum LDH) and the number of plasma volumes exchanged per procedure did not affect the relapse rate. Initial plt and LDH were not predictive of mortality. Response, relapse, and mortality rates with the combination of 5% albumin for the initial 50% of TPE followed by plasma for the final 50% of TPE as replacement were comparable or possibly better than plasma-only replacement strategies. Forty percent of centers routinely used a TPE taper; however, there was no statistical difference in relapse rates comparing the taper and non-taper sub-groups. By controlling for adjunctive modalities such as steroids and anti-platelet agents, it is hoped that future prospective clinical trials may optimize the role of TPE in TTP, minimize patient exposure to blood products and procedures, shorten the clinical course of TTP, and reduce mortality.