Thrombotic Thrombocytopenic Purpura Patients Research Articles

How I treat refractory thrombotic thrombocytopenic purpura

AbstractAcquired thrombotic thrombocytopenic purpura (TTP) is characterized by thrombocytopenia and microangiopathic hemolytic anemia (MAHA) without an obvious cause, and may include fever, mild renal failure, and neurologic deficits. It is characterized by a deficiency of the von Willebrand factor (VWF) cleaving enzyme, ADAMTS13 (a disintegrin and metalloproteinase, with a thrombospondin type 1 motif, member 13), resulting in formation of microthrombi in the high sheer environment of the microvasculature. This causes microvascular occlusion, MAHA, and organ ischemia. Diagnosis is based on the presence of clinical symptoms, laboratory aberrations consistent with MAHA, decreased ADAMTS13 activity, and possibly presence of anti-ADAMTS13 autoantibodies. Upfront treatment of acute TTP includes plasma exchange and corticosteroids. A significant number of patients are refractory to this treatment and will require further interventions. There are limited data and consensus on the management of the refractory TTP patient. Management involves simultaneously ruling out other causes of thrombocytopenia and MAHA, while also considering other treatments. In this article, we describe our management of the patient with refractory TTP, and discuss use of rituximab, increased plasma exchange, splenectomy, and immunosuppressive options, including cyclophosphamide, vincristine, and cyclosporine. We also review recent evidence for the potential roles of bortezomib and N-acetylcysteine, and explore new therapeutic approaches, including recombinant ADAMTS13 and anti-VWF therapy.

Role of the skin biopsy in the diagnosis of atypical hemolytic uremic syndrome.

Atypical hemolytic uremic syndrome (aHUS) is a prototypic thrombotic microangiopathy attributable to complement dysregulation. In the absence of complement inhibition, progressive clinical deterioration occurs. The authors postulated that a biopsy of normal skin could corroborate the diagnosis of aHUS through the demonstration of vascular deposits of C5b-9. Biopsies of normal skin from 22 patients with and without aHUS were processed for routine light microscopy and immunofluorescent studies. An assessment was made for vascular C5b-9 deposition immunohistochemically and by immunofluorescence. The biopsies were obtained primarily from the forearm and/or deltoid. Patients with classic features of aHUS showed insidious microvascular changes including loose luminal platelet thrombi, except in 2 patients in whom a striking thrombogenic vasculopathy was apparent in biopsied digital ulcers. Extensive microvascular deposits of the membrane attack complex/C5b-9 were identified, excluding 1 patient in whom eculizumab was initiated before biopsy. In 5 of the 7 patients where follow-up was available, the patients exhibited an excellent treatment response to eculizumab. Patients without diagnostic clinical features of aHUS failed to show significant vascular deposits of complement, except 2 patients with thrombotic thrombocytopenic purpura including 1 in whom a Factor H mutation was identified. In a clinical setting where aHUS is an important diagnostic consideration, extensive microvascular deposition of C5b-9 supports the diagnosis of either aHUS or a subset of thrombotic thrombocytopenic purpura patients with concomitant complement dysregulation; significant vascular C5b-9 deposition predicts clinical responsiveness to eculizumab.

Platelet transfusions in platelet consumptive disorders are associated with arterial thrombosis and in-hospital mortality

AbstractWhile platelets are primary mediators of hemostasis, there is emerging evidence to show that they may also mediate pathologic thrombogenesis. Little data are available on risks and benefits associated with platelet transfusions in thrombotic thrombocytopenic purpura (TTP), heparin-induced thrombocytopenia (HIT) and immune thrombocytopenic purpura (ITP). This study utilized the Nationwide Inpatient Sample to evaluate the current in-hospital platelet transfusion practices and their association with arterial/venous thrombosis, acute myocardial infarction (AMI), stroke, and in-hospital mortality over 5 years (2007-2011). Age and gender-adjusted odds ratios (adjOR) associated with platelet transfusions were calculated. There were 10 624 hospitalizations with TTP; 6332 with HIT and 79 980 with ITP. Platelet transfusions were reported in 10.1% TTP, 7.1% HIT, and 25.8% ITP admissions. Platelet transfusions in TTP were associated with higher odds of arterial thrombosis (adjOR = 5.8, 95%CI = 1.3-26.6), AMI (adjOR = 2.0, 95%CI = 1.2-3.3) and mortality (adjOR = 2.0,95%CI = 1.3-3.0), but not venous thrombosis. Platelet transfusions in HIT were associated with higher odds of arterial thrombosis (adjOR = 3.4, 95%CI = 1.2-9.5) and mortality (adjOR = 5.2, 95%CI = 2.6-10.5) but not venous thrombosis. Except for AMI, all relationships remained significant after adjusting for clinical severity and acuity. No associations were significant for ITP. Platelet transfusions are associated with higher odds of arterial thrombosis and mortality among TTP and HIT patients.

Effect of rituximab on B cell phenotype and serum B cell-activating factor levels in patients with thrombotic thrombocytopenic purpura.

Autoantibodies inhibiting the activity of the metalloproteinase, ADAMTS13 (a disintegrin and metalloproteinase with a thrombospondin type 1 motif, member 13), underlie the pathogenesis of thrombotic thrombocytopenic purpura (TTP). Rituximab (RTX) combined with plasma-exchange (PEX) is an effective treatment in TTP. Patients can remain in remission for extended periods following PEX/RTX, and this is associated with continuing reduction in antibodies to ADAMTS13. Factors controlling B cell differentiation to autoantibody production, including stimulation through the B cell receptor and interactions with the B cell-activating factor (BAFF), may thus impact length of remission. In this cross-sectional study, we measured naive and memory B cell phenotypes [using CD19/immunoglobulin (Ig)D/CD27] following PEX/RTX treatment in TTP patients at B cell return (n=6) and in 12 patients in remission 10-68 months post-RTX. We also investigated relationships among serum BAFF, soluble CD23 (sCD23(-) a surrogate measure of acquiring B memory (CD27(+) ) phenotype) and BAFF receptor (BAFF-R) expression. At B cell return after PEX/RTX, naive B cells predominated and BAFF-R expression was reduced compared to healthy controls (P<0.001). In the remission group, despite numbers of CD19(+) B cells within normal limits in most patients, the percentage and absolute numbers of pre-switch and memory B cells remained low, with sCD23 levels at the lower end of the normal range. BAFF levels were correlated inversely with BAFF-R expression and time after therapy. In conclusion, the long-term effects of RTX therapy in patients with TTP included slow regeneration of memory B cell subsets and persistently reduced BAFF-R expression across all B cell subpopulations. This may reflect the delay in selection and differentiation of potentially autoreactive (ADAMTS13-specific) B cells, resulting in relatively long periods of low disease activity after therapy.

Ribosomal and immune transcripts associate with relapse in acquired ADAMTS13-deficient thrombotic thrombocytopenic purpura.

Approximately 40% of patients who survive acute episodes of thrombotic thrombocytopenic purpura (TTP) associated with severe acquired ADAMTS13 deficiency experience one or more relapses. Risk factors for relapse other than severe ADAMTS13 deficiency and ADAMTS13 autoantibodies are unknown. ADAMTS13 autoantibodies, TTP episodes following infection or type I interferon treatment and reported ensuing systemic lupus erythematosus in some patients suggest immune dysregulation. This cross-sectional study asked whether autoantibodies against RNA-binding proteins or peripheral blood gene expression profiles measured during remission are associated with history of prior relapse in acquired ADAMTS13-deficient TTP. Peripheral blood from 38 well-characterized patients with autoimmune ADAMTS13-deficient TTP in remission was examined for autoantibodies and global gene expression. A subset of TTP patients (9 patients, 24%) exhibited a peripheral blood gene signature composed of elevated ribosomal transcripts that associated with prior relapse. A non-overlapping subset of TTP patients (9 patients, 24%) displayed a peripheral blood type I interferon gene signature that associated with autoantibodies to RNA-binding proteins but not with history of relapse. Patients who had relapsed bimodally expressed higher HLA transcript levels independently of ribosomal transcripts. Presence of any one potential risk factor (ribosomal gene signature, elevated HLA-DRB1, elevated HLA-DRB5) associated with relapse (OR = 38.4; p = 0.0002) more closely than any factor alone or all factors together. Levels of immune transcripts typical of natural killer (NK) and T lymphocytes positively correlated with ribosomal gene expression and number of prior episodes but not with time since the most recent episode. Flow cytometry confirmed elevated expression of cell surface markers encoded by these transcripts on T and/or NK cell subsets of patients who had relapsed. These data associate elevated ribosomal and immune transcripts with relapse history in acquired, ADAMTS13-deficient TTP.

Cardiac troponin‐I on diagnosis predicts early death and refractoriness in acquired thrombotic thrombocytopenic purpura. Experience of the French Thrombotic Microangiopathies Reference Center

Cardiac involvement is a major cause of mortality in patients with thrombotic thrombocytopenic purpura (TTP). However, diagnosis remains underestimated and delayed, owing to subclinical injuries. Cardiac troponin-I measurement (cTnI) on admission could improve the early diagnosis of cardiac involvement and have prognostic value. To assess the predictive value of cTnI in patients with TTP for death or refractoriness. The study involved a prospective cohort of adult TTP patients with acquired severe ADAMTS-13 deficiency (< 10%) and included in the registry of the French Reference Center for Thrombotic Microangiopathies. Centralized cTnI measurements were performed on frozen serum on admission. Between January 2003 and December 2011, 133 patients with TTP (mean age, 48 ± 17 years) had available cTnI measurements on admission. Thirty-two patients (24%) had clinical and/or electrocardiogram features. Nineteen (14.3%) had cardiac symptoms, mainly congestive heart failure and myocardial infarction. Electrocardiogram changes, mainly repolarization disorders, were present in 13 cases. An increased cTnI level (> 0.1 μg L(-1) ) was present in 78 patients (59%), of whom 46 (59%) had no clinical cardiac involvement. The main outcomes were death (25%) and refractoriness (17%). Age (P = 0.02) and cTnI level (P = 0.002) showed the greatest impact on survival. A cTnI level of > 0.25 μg L(-1) was the only independent factor in predicting death (odds ratio [OR] 2.87; 95% confidence interval [CI] 1.13-7.22; P = 0.024) and/or refractoriness (OR 3.03; 95% CI 1.27-7.3; P = 0.01). A CTnI level of > 0.25 μg L(-1) at presentation in patients with TTP appears to be an independent factor associated with a three-fold increase in the risk of death or refractoriness. Therefore, cTnI level should be considered as a prognostic indicator in patients diagnosed with TTP.

Clinical and Therapeutic Studies of Acquired Thrombotic Thrombocytopenic Purpura in China

Thrombotic thrombocytopenic purpura (TTP) is a rare and severe disorder mediated by autoantibodies against ADAMTS13. In this report, we study the clinical features, laboratory aberration, and treatment effect of 55 patients with acquired TTP in China. The classic pentad occurred in only 33% of TTP patients. Severe ADAMTS13 deficiency was detected in 85% of patients. Advanced age and hyperbilirubinemia might be risk factors for poor prognosis. Early and sufficient plasma exchange is the most important approach. The addition of rituximab to plasma exchange and corticosteroids appears to be effective in inducing and sustaining long-term remission in TTP, and is suitable to be administrated during the first episode. However, more optimal therapeutic regimen warrants further investigation to treat refractory cases and to reduce relapse rate.

Thrombotic Thrombocytopenic Purpura (TTP) Patient Attitudes Regarding Depression Management: a Qualitative Study

▪Introduction: Recovery from an acute episode of TTP was previously thought to be complete except for a risk for relapse. However, the Oklahoma TTP-HUS Registry has documented that patients have multiple health problems that persist following recovery, including an increased prevalence of major depression (Blood122:2023;2013). Untreated major depression impacts daily functioning, quality of life and is a risk factor for suicide. Recognizing depression and establishing effective treatment should be a critical part of TTP follow-up care. The objective of our study was to identify TTP patient attitudes regarding treatments for depression.Methods: The study was based on the theoretical framework of the Theory of Planned Behavior (TPB). The theory suggests that intention is the immediate predictor of behavior. Intention is determined by 3 factors: attitude toward a behavior, subjective norms, and perceived behavioral control. This study focused on attitude; therefore, patients were primarily asked if there were any advantages or disadvantages of treating depression with either pharmacotherapy or counseling.One-on-one semi-structured interviews were conducted by the first author. Interviews were transcribed verbatim and content analysis was conducted by 3 qualitative researchers and facilitated by using the NVivo 10 software. A codebook was developed to guide the data analysis. Similar beliefs were grouped into themes and key ideas.Patients: Registry patients whose initial episode was associated with severe ADAMTS13 deficiency (<10%), 1995-2012, and who had major depression on at least 1 screening evaluation from 2004-2012. Qualitative purposeful sampling was used instead of random sampling. Patients were stratified by previous/current or no pharmacotherapy treatment regardless of counseling history.Results: Fifty-six ADAMTS13-deficient patients in clinical remission had been screened for depression; 50 patients were alive in 2013; 24 had a score of major depression at least once. Of the 24 patients, 12 (50%) had taken pharmacotherapy for their depression and 12 (50%) patients had never taken pharmacotherapy.Data saturation was achieved after interviewing 16 patients (9 pharmacotherapy; 7 no pharmacotherapy). Fourteen patients (88%) were women, 9 (56%) were black, and the median age was 49 years (range 28-69). Patients were a median of 8.5 years from their most recent TTP episode (range 2-17 years), 5 (31%) had relapsed, and 7 (44%) had been treated with rituximab.Interviews among all patients revealed that a TTP diagnosis was 'life-altering’ and 'traumatic’. Many patients felt the residual effects of TTP (a fear of relapsing, fatigue, and mild cognitive impairment) were related to their depression. A major theme across groups was that patients had a positive attitude toward counseling and believed counseling could be an effective way to treat depression. However, a history of a negative counseling experience seemed to discourage many patients from continuing counseling. Additionally, regardless of previous counseling experience, a key idea across groups was mistrust toward the mental healthcare provider’s ability. However, this mistrust was only a perception among those who had never tried counseling. Within the pharmacotherapy group, patients believed that medication was an effective way to treat depression; however, many patients reported side effects (feeling 'sluggish’ and 'not caring about anything’) as the reasons for stopping the medication. On the contrary, among the non-pharmacotherapy group patients were unsure if pharmacotherapy actually worked-'Uh, I guess it kind of would keep you steady-minded I guess’. The fear of side effects and the fear of addiction were the biggest barriers among the non-pharmacotherapy group for using pharmacotherapy.Conclusion:Hematologists need to recognize that TTP is a long-term life-altering diagnosis following recovery. Identifying and managing depression is a critical issue that can save lives. TTP patients seem to have a positive attitude toward counseling and a mixed attitude toward pharmacotherapy. Whether these fears are valid is beyond the scope of this study; however, it is imperative that hematologists discuss the issue of depression in depth with their patients and address the barriers to acceptance of treatment. This study is a first step in identifying barriers to depression treatment in TTP survivors. DisclosuresNo relevant conflicts of interest to declare.

Domain Specificity and Mechanism of Action of Anti-ADAMTS13 Antibodies at Presentation and Relapse in Acquired Thrombotic Thrombocytopenic Purpura (TTP)

Acquired thrombotic thrombocytopenic purpura (TTP) is an autoimmune disease due to autoantibodies (Auto-Ab) against ADAMTS13, resulting in a severe deficiency in enzyme activity (<10%). Little is known about the pathogenicity of anti-ADAMTS13 IgG directed against different domains of the enzyme, or how domain specificity alters between presentation, remission or TTP relapse. We developed novel assays to explore domain specificity and inhibitory potential of anti-ADAMTS13 IgG using samples from the UK TTP Registry.Full-length ADAMTS13 and ADAMTS13 fragments (MD, MDTC and MDTCS - N-terminal domains; TSR2-8 and CUB1/2 - C-terminal domains) were expressed in HEK293T cells and purified. Microtitre plates were coated with ADAMTS13 or ADAMTS13 fragments, incubated with TTP plasmas and bound IgG detected. Domain specificity was further analysed by preincubation of patient samples with ADAMTS13 fragments, assessing the ability of fragments to compete for full-length ADAMTS13 binding on the plate. From a cohort of 92 acquired TTP patients at presentation, 100% (92/92) contained auto-Ab that recognised ADAMTS13 N-terminal domains (MDTCS). Of these, 39% (36/92) had auto-Ab recognising the N-terminal domains alone, 22% (20/92) also had anti-TSR2-8, and 39% (36/92) had anti-CUB1/2 auto-Abs. There was no correlation between auto-Ab domain specificity and mortality/disease severity (anti-ADAMTS13 IgG titre, plasma exchanges to remission, platelet count and cardiac involvement).There was no evidence for auto-Ab recognising MD or MDTC domains in 25 patients tested, suggesting immunoreactivity for MDTCS was primarily mediated through the spacer domain. Consistent with this, TTP IgG inhibited proteolysis of VWF115 (a recombinant VWF A2 domain fragment), but failed to inhibit VWF106 proteolysis (VWF A2 domain fragment lacking the spacer domain binding exosite).To examine the inhibitory potency of anti-ADAMTS13 auto-Ab, IgG isolated from TTP patient samples were titrated in reactions measuring MDTCS-mediated proteolysis of FRETS-VWF73. Twenty eight out of 43 presentation samples tested had inhibitory IgG (IC50 ranging from <0.2-5μM total IgG). Addition of excess of MDTC to 19/19 of these samples did not alter the IC50, confirming inhibition in these samples is mediated solely by anti-spacer antibodies. Intriguingly, 15/43 patients had no evidence of inhibitory auto-Ab, suggesting the pathogenic mechanism in these cases may be enhanced clearance of ADAMTS13. We therefore determined ADAMTS13 antigen concentrations by ELISA. All patients at presentation exhibited markedly reduced ADAMTS13 antigen levels (n=91, median 58ng/ml (range 0-450) vs. normal n=67, median 951ng/ml (515-1829); p<0.0001). Interestingly, patients with predominantly C terminal auto-Ab and no evidence of inhibitory auto-Ab had significantly lower ADAMTS13 antigen levels at first presentation than the rest of the cohort (median 2ng/ml (0-141) vs. 57ng/ml (0-356); p=0.006).Cases where rituximab failed to normalise ADAMTS13 activity despite clinical remission were associated with the persistence of inhibitory anti-N-terminal auto-Ab. Two patients with persistently high anti-ADAMTS13 IgG but normal ADAMTS13 activity and antigen levels during remission had non-pathogenic anti-TSR2-8 antibodies that seemingly had no effect upon ADAMTS13 function or clearance. Nine out of 16 patients exhibited altered auto-Ab domain specificity upon relapse, with either loss or development of anti-C-terminal auto-Ab.In conclusion, from our functional studies ADAMTS13 inhibition appears to be mediated exclusively by anti-spacer antibodies, with no evidence for the presence of auto-Ab that recognise/inhibit other N terminal domains. Importantly, we demonstrate ADAMTS13 antigen depletion is a crucial pathogenic mechanism in acquired TTP, accounting for a large proportion of the reduced ADAMTS13 activity detected in patient samples. Our results suggest that auto-Ab directed against the C-terminal domains of ADAMTS13 may be particularly important in mediating this clearance. There is continual development of the autoimmune response during treatment of TTP patients that can result in altered domain specificity at relapse. DisclosuresNo relevant conflicts of interest to declare.

Complement Activation May Trigger the Onset of Thrombotic Thrombocytopenic Purpura in Patients with Severe ADAMTS13 Deficiency

Thrombotic thrombocytopenic purpura (TTP), a potential fatal syndrome, is often associated with severe deficiency of plasma ADAMTS13 activity, either resulting from ADAMTS13 mutations or acquired anti-ADAMTS13 autoantibodies that inhibit plasma ADAMTS13 activity. Patients with severe ADAMTS13 do not always have TTP signs and symptoms, which often occur following infections or inflammatory responses. The mechanism of TTP flare is not fully understood. In the present study, complement activation markers (iC3b, C5b, Bb, and C4b) were determined by enzyme-linked absorbent assays (ELISA) in the initial plasmas (prior to plasma exchange) of 20 patients with acquired TTP with severe ADAMTS13 deficiency (less than 20% of normal) and plasmas from 20 healthy controls. Of 20 TTP patients, 19 exhibited positive inhibitor in the 50:50 mixing study. Plasma levels of iC3b (1,000 ± 1,062 ng/ml), sC5b-9 (1,342±867 ng/ml), and Bb (38.2±47.7 ng/ml), as well as C4b (74.3±49.5 ng/ml) in acquired TTP patients were significantly higher than those in healthy controls (p value less than 0.01) These results indicate that complement activation in both classic and alternative pathways is a common phenomenon in patients with acquired autoimmune TTP. To demonstrate the causative effect of complement activation in TTP, we turned to our Adamts13 null mice. C57BL/6 (Adamts13-/-) mice are resistant to the development of spontaneous and Shigatoxin-induced TTP syndrome. When injected with a murine specific monoclonal antibody against complement factor H (CFH) (800 micro grams/mouse), which inhibits binding of circulating CFH to endothelial cells and C3b, Adamts13-/- mice (C57BL/6) developed more severe thrombocytopenia and anemia than wild type mice did within 6 days without additional challenge. However, renal insufficiency manifested by the increase of plasma BUN concentration was similar in both groups (Fig. 1). These results indicate that complement activation through an alternative pathway, following antibody-mediated inhibition of CFH or other complement regulatory components, may trigger the onset of TTP in light of severe ADAMTS13 deficiency. [Display omitted] DisclosuresNo relevant conflicts of interest to declare.

A Phase II Study Evaluating the Efficacy of Rituximab in the Management of Patients with Relapsed or Refractory TTP

Background: As many as 30-50% of patients diagnosed with Thrombotic Thrombocytopenic Purpura (TTP) either relapse or become refractory to therapeutic plasma exchange (TPE). There remains a pressing need to evaluate novel treatments for these patients. Rituximab is a chimeric anti-CD20 monoclonal antibody that has demonstrated efficacy potentially by eliminating active B-lymphocytes and production of ADAMTS 13 inhibitor considered central to the pathogenesis of TTP.Intervention: We conducted a prospective multi-center phase II open-label trial in Canadian hospital-based apheresis units to evaluate the efficacy of rituximab in the management of adult patients with relapsed or refractory TTP. Functional and antigenic ADAMTS13 enzyme, inhibitor levels and anti-CD 20 were measured at initiation, 8, 12, 24 and 52-weeks as well as routine hematologic parameters, complete remission, rate of relapse, overall mortality and adverse events. Rituximab was administered intravenously once weekly for a total of 4 doses at 375 mg/m2 in eligible consenting patients.Results: Twenty refractory and 20 relapsing TTP patients were enrolled (N=40). A complete response (CR) was defined by a platelet count >150 x 109/L, an LDH <1.5 times the upper limit of normal, absence of progressive neurologic symptoms and no further need for TPE. At week 8, 77% of refractory patients and 90% of relapsed patients were in CR. At week 52, 15/20 refractory patients (3 deaths, 1 lost to follow-up) and 18/20 of relapsed patients (1 withdrew, 1 lost to follow-up) were alive and in CR. Of the 40 patients enrolled a total of 3 patients died; all in the refractory arm: two with functional ADAMTS13 <10% and detectable inhibitor. Two deaths were related to fungal infection (week 2) and active TTP (week 8). A third death was attributed to thrombotic complications at week 29. All refractory patients with normal ADAMTS13 enzyme at baseline had a complete response at week 8 and remained in remission for the remainder of follow-up (52 weeks). Despite being in complete hematological and clinical remission, relapsing TTP patients still had a positive inhibitor at 24 (50% of patients) and 52 weeks (10% of patients). All patients had < 3% CD 20+ B-lymphocytes at 8 and 12 weeks post rituximab. Median platelet counts in the refractory group recovered more slowly than in the relapsing TTP patients. By week 8 refractory TTP patients received a median of 26 plasma exchanges and relapsed patients received a median of 11(P<0.001).Conclusion: At the conclusion of this study the overall mortality rate was 8%, 15% (95% CI: 5% to 36%) for the refractory group and 0% (95% CI: 0% to 16%) for the relapsed group. Nine refractory patients were dialysis-dependent at baseline and only 2 had a serum creatinine >250 µmol/L and remained on dialysis at week 52. These patients had normal ADAMTS13 enzyme measurements throughout the time period of the study. Results of this study indicate that TPE and rituximab is an effective combination in patients with relapsed or refractory TTP. DisclosuresFoley:Novartis: Consultancy, Honoraria; Pfizer: Consultancy, Honoraria; Celgene: Consultancy, Honoraria; Sanofi: Consultancy, Honoraria; Lundbeck: Consultancy, Honoraria; Hoffman-LaRoche: Consultancy, Honoraria, Research Funding; Jansen: Consultancy, Honoraria; Alexion: Consultancy, Honoraria. Off Label Use: Rituxan is a CD20-directed cytolytic antibody indicated for the treatment of patients with: -Non-Hodgkin’s Lymphoma (NHL) (1.1) -Chronic Lymphocytic Leukemia (CLL) (1.2) - Rheumatoid Arthritis (RA) in combination with methotrexate in adult patients with moderately-to severely-active RA who have inadequate response to one or more TNF antagonist therapies (1.3) . Arnold:GSK: Honoraria, Research Funding; Hoffman-LaRoche: Research Funding; Bristol Myers Squibb: Consultancy; Amgen: Consultancy, Honoraria, Research Funding. Clark:Alexion: Honoraria; CSN: Honoraria; ASN: Honoraria; Octapharm: Honoraria; Danone Research: Honoraria.

Predicting the Temporal Course of Laboratory Abnormality Resolution in Patients with Thrombotic Microangiopathy

Background:Screening for deficiency of ADAMTS- 13, a von Willebrand factor-cleaving protease, is commonly performed when there exists a clinical suspicion for thrombotic thrombocytopenic purpura (TTP). The clinical utility of ADAMTS13 testing at presentation has been questioned, as initial treatment decisions are variably influenced by results. Furthermore, current literature remains inconclusive about the differences in response to plasma exchange therapy in patients with severe (<5%) and non-severe (>5%) ADAMTS13 deficiency.Methods:With IRB approval, we performed a single center, retrospective review of patients undergoing plasma exchange therapy at Duke University Medical Center for indication of thrombotic microangiopathy from 2007 through June 2013. We retrospectively identified patients with “severe” deficiency (ADAMTS13² 5%, n=22) and those without severe deficiency (ADAMTS13 >5% or “non-severe” n=22). Cases of TTP associated with bone marrow transplantation, and chemotherapeutic agents were excluded. We trended daily laboratory values for creatinine, platelets, and LDH through completion of TPE . Time to recovery of laboratory abnormalities to normal was noted. We compared the time to resolution of laboratory abnormalities between patients with ADAMTS13² 5% (“severe”) to those with level >5% (“non-severe”).Results:Demographic and clinical information for TTP patients with ADAMTS13<5% and >5% are shown in the Table below. All patients underwent plasma exchange therapy plus steroids after diagnosis with TTP. Patients in both groups were comparable with respect to age and gender, but there were more episodes of recurrent TTP (n=9) in the ADAMTS13 <5% group compared to >5% (n=4). Patients with severe ADAMTS13 deficiency required more TPE (n=19 ) than patients with non-severe ADAMTS13 deficiency (n=13). There were 5 deaths in the non-severe ADAMTS13 cohort as compared to no deaths in the severe ADAMTS13 deficiency.Recovery of platelet count, LDH and Cr level to within normal range are shown for each group within 7, 14 and 21 days of presentation. Recovery of platelet counts for both groups were comparable. No recovery of platelet counts was seen in 3 patients for each group. The majority of patientÕs with and without severe ADAMTS13 deficiency appeared to recover platelet count by day 14 (81% for severe v. 75% for non-severe ADAMTS13 deficiency). Recovery of LDH differed in the two groups. Patients with non-severe ADAMTS13 deficiency were more likely to show no recovery in LDH (38% v. 4% in severe deficiency) or delayed LDH recovery (62% normalized LDH by D21 in non-severe v 95% in severe deficiency). As previously reported, patients with non-severe ADAMTS13 deficiency were more likely to have renal disease and less likely to recover renal function. Few patients in the severe deficiency group presented with abnormal renal function, but in those who had renal dysfunction, the majority (71%) recovered renal function by day 14.Conclusions:Our studies show the temporal resolution of laboratory parameters in patients with severe and non-severe ADAMTS13 deficiency. We show that both groups have similar platelet recovery patterns in response to TPE. However, patients with non-severe ADAMTS13 deficiency have delayed normalization of LDH, higher rates of ESRD and higher mortality. Although additional prospective analysis will need to be performed, this data provides preliminary data showing differing disease pathophysiology of the two groups and that patients with non-severe ADAMTS13 may benefit from alternative/adjunctive therapies to reduce mortality. TableClinical FeaturesADAMTS13 <5%ADAMTS13>5%Total number2222Males (M)/Females (F)7(M)/15(F)9(M)/13(F)Age (mean)4950Recurrent episodes94Mean # TPE1913Deaths05Platelet Count RecoveryTotal number with abnormal plts2221No recovery33*Plts >150K by day ² 713 (59%)9 (45%)Plts>150K by day ² 1418 (82%)15 (75%)Plts >150K by day ² 2121 (86%)15 (75%)LDH normalizationTotal number with elevated LDH2221No recovery1 (5%)8 (38 %)Normal LDH by day ²715(68 %)6 (29%)Normal LDH by day ²1420 (91%)10 (48%)Normal LDH by day ²2121 (95%)13 (62%)Renal function recoveryTotal number with abnormal Cr values at presentation712No recovery+29*Normal Cr by day 73 (42.85%)0Normal Cr by day 145 (71.42%)1 (8.33%)Normal Cr by day 215 (71.42%)3 (16.66%) DisclosuresArepally:TEVA Pharma: Consultancy.

Low dose Rituximab for Preemptive Management of Thrombotic Thrombocytopenic Purpura ( TTP )

Low dose rituximab has been successful in treating some patients with autoimmune cytopenias. Recently reported data indicate preemptive rituximab may postpone hematologic relapses of TTP in patients with chronic severe ADAMTS13 deficiency.A now 22-year-old woman had multi-relapsing TTP with five episodes over the previous 10 years, three of which were successfully treated with four doses of rituximab 375 mg/m2. The most recent course in 2009 was given preemptively because intense ADAMTS13 monitoring had shown that previous hematologic relapses followed shortly after developing severe ADAMTS13 deficiency. Asymptomatic chronic IgG deficiency less than 300 mg% had followed this most recent rituximab course given in 2009.In 2014, continued intense ADAMTS13 monitering disclosed a new progressive decrease in ADAMTS13 levels from normal to severely deficient levels with a weak inhibitor, but no hematologic or clinical changes. Just two 100 mg rituximab doses have been followed by a progressive rise of ADAMTS13 levels to solidly normal values with significant improvement by day 13 post initiation and normalization by day 34.Low dose rituximab could be a viable alternative for preemptively managing TTP patients with persistently severly low ADAMTS13 levels. DisclosuresOff Label Use: Off label use of rituximab to treat TTP.

The Role of ADAMTS13-Specific Circulating Immune Complexes in Prediction of Recurrence of Acquired Thrombotic Thrombocytopenic Purpura

Introduction. Acquired thrombotic thrombocytopenic purpura (TTP) is a rare thrombotic microangiopathy due to the development of autoantibodies against the VWF-cleaving protease ADAMTS13. We recently developed and validated a new ELISA method for the detection of ADAMTS13-specific circulating immune complexes (CICs) in acquired TTP patients. This study aims at investigating the clinical relevance of ADAMTS13-specific CICs at disease presentation in a large cohort of acquired TTP patients.Methods. We measured ADAMTS13-specific CICs by ELISA in 51 patients from the Milan TTP Registry, at the first episode of acquired TTP. All patients presented anti-ADAMTS13 autoantibodies by western blotting and severe ADAMTS13 deficiency (i.e., <10% of normal) by FRETS-VWF73 or CBA assays. We studied the associations between ADAMTS13-specific CICs levels and (i) ADAMTS13-related measurements (i.e., ADAMTS13 antigen, anti-ADAMTS13 IgG), (ii) clinical and laboratory markers of disease severity (i.e., muco-cutaneous bleeding, neurological, renal and cardiovascular symptoms, number of platelets, LDH, hemoglobin, creatinine), (iii) short- and long-term clinical outcomes (i.e., number of plasma exchange procedures required to attain remission, recurrence). Statistical analyses were performed using linear, logistic and Cox regression models.Results. The prevalence of ADAMTS13-specific CICs in patients experiencing a first episode of acquired TTP was 39% (95% confidence intervals [CI]: 26-52%). ADAMTS13-specific CICs were only associated with ADAMTS13 antigen levels at regression analyses (beta per 10% increase in ADAMTS13 antigen, 95% CI: 0.05, 0.02-0.08). The presence of ADAMTS13-specific CICs was associated with more than a four-fold increase in the risk of recurrence at 2 years after the first TTP episode (hazard ratio, 95%CI: 4.2, 1.1-16.4).Conclusions. ADAMTS13-specific CICs are neither a biomarker of disease severity, nor a predictor of clinical outcome during acute phase. Conversely, ADAMTS13-specific CICs seem to have relevance in predicting the recurrence of acute TTP episodes. DisclosuresPeyvandi:NovoNordisk: Research Funding; Kedrion : Research Funding; NovoNordisk: Speakers Bureau; CSL Behring : Speakers Bureau; Baxter: Speakers Bureau; Bayer: Speakers Bureau.

Introduction of a Quick Assay for ADAMTS13 Activity Improved a Survival of Acquired TTP Patients Who Received Platelet Transfusions

Introduction: Daily plasma exchange (PE) has become a definitive first-line treatment for acquired thrombotic thrombocytopenic purpura (TTP). However, it remains to be controversial whether platelet transfusion is harmful or not for patients with acquired TTP during initial treatment. Some articles showed that platelet transfusion was considered as hazardous because platelet transfusion might generate widespread fresh platelet aggregates in circulation, but others reported that the mortality rate was not different between patients with and without platelet transfusion. Herein, we conducted a retrospective analysis of a large cohort of patients with acquired idiopathic TTP (ai-TTP) in Japan evaluating whether platelet transfusion was associated with unfavorable outcomes.Patients and Methods: Our laboratory has been functioning as a nationwide referral center for thrombotic microangiopathies (TMAs) in Japan. We collected a large dataset of medical information on 1211 patients with TMA from March 2000 to December 2013. Among them, 263 were ai-TTP patients with severe deficiency (<10% of normal controls) of ADAMTS13 activity (ADAMTS13:AC), with positive anti-ADAMTS13 inhibitors. These ai-TTP patients were retrospectively analyzed in detail using reported medical records that contained detailed clinical data and outcome information. ADAMTS13:AC was determined by classic von Willebrand factor multimer (VWF) assay. These results were available within 4 to 7 days in the initial period (March 2000 to March 2005, n=91). Subsequently, with use of a chromogenic ADAMTS13-act-ELISA, reporting occurred within 2 days of obtaining a plasma sample (April 2005 to December 2013, n=172). All plasma samples collected before March 2005 were re-examined by act-ELISA.Results: In 263 ai-TTP patients, 48 patients received platelet transfusions during the initial treatment and 215 did not. Factors associated with increased mortality included age greater than 60 years and presentation with central nervous system (CNS) dysfunction (p<0.05 for each), but not use of platelet transfusions [22.9% (11/48) versus 17.7% (38/215), respectively].Next, we analyzed the mortality rate in 2 groups categorized by method of ADAMTS13 activity assay. In the initial period using classic VWF assay, mortality of patients with platelet transfusion tended to be higher versus those without platelet transfusions (p=0.051, Log-rank test) as shown in Figure left (Kaplan-Meier survival curve). In contrast, in the latter period using the more rapid act-ELISA, no differences in mortality of patients with versus without platelet transfusions was noted (p=0.52, Figure right). To evaluate whether platelet transfusion is a significant hazard for patients with ai-TTP, Cox-proportional-hazards regression analysis was performed using 5 variables (age, sex, rituximab use, presence of CNS dysfunction, and platelet transfusion). Older age and use of platelet transfusion were independently associated with mortality in the initial period, and age and presence of CNS dysfunction were identified as unfavorable factors in the latter period (Table).Conclusions: Our results clearly indicated that platelet transfusion was harmful in ai-TTP patients with severe deficiency of ADAMTS13:AC before 2005 (when a quick ADAMTS13:AC assay was unavailable). After 2005 (when a quick ADAMTS13:AC assay available), the survival rate of ai-TTP patients with platelet transfusions became almost indistinguishable from those without. This result may indicate that persistent PE after platelet transfusion together with confirmation of severe ADAMTS13:AC reduces risk for aggravated platelet thrombi formation in microvasculatures of patients. However, use of platelet transfusion before PE may lead to serious thrombotic complications in heart and brain, a complication that should be avoided. [Display omitted] [Display omitted] DisclosuresMatsumoto:Alfresa Pharma Corporation: Patents & Royalties. Fujimura:Alfresa Pharma Corporation: Patents & Royalties.

Weekend Versus Weekday Mortality for Thrombotic Thrombocytopenic Purpura (TTP) Related Hospitalizations in the United States: Data from the National Inpatient Sample

Introduction: Previous research has shown that weekend hospital admissions are associated with an increased mortality in comparison to weekday admissions for a number of emergent conditions including myocardial infarction [Relative Risk (RR) 1.048; 95% confidence interval [CI], 1.022 to 1.076; P value <0.001], pulmonary embolism (RR 1.17, 95% CI 1.03 to 1.34, P value <0.01) and gastrointestinal hemorrhage (RR 1.17, 95% CI 1.03 to 1.34, P value <0.01) . Thrombotic Thrombocytopenic Purpura (TTP) is a hematological emergency with a significant morbidity and mortality if not recognized early. However, studies evaluating if a similar "weekend" effect exists in TTP are lacking.Methods: We used the Nationwide Inpatient Sample database to identify patients admitted with TTP in the United States using ICD 9 CM code 446.6 from 2009 to 2011. Baseline data for demographic variables, age, gender, race, hospital characteristics- region, hospital type (rural versus urban, teaching versus non-teaching), bed-size, insurance payer and comorbidities were derived for weekend and weekday admissions. Logistic regression analysis was used to calculate the adjusted relative risk of in-hospital mortality of weekend versus weekday admissions. Data analysis was done using STATA 13.0 (College Station, TX: StataCorp LP)Results: Of the 6634, estimated TTP related hospitalizations, 19.5 % were admitted on the weekends and 80.5 % admitted on the weekdays. The mean age was 48±0.5 years and 66.4 % were females. A higher in-hospital mortality rate was seen among weekend admissions as compared to weekday admissions (RR 1.32, 95% CI 1.30-1.33, p value <0.01). On multivariate analysis (table 1), weekend admission remained as an independent predictor of increased mortality (adjusted RR 1.16, 95% CI 1.15-1.17, P value <0.01) after adjusting for other confounders including age, gender, comorbidities, hospital type and size. Similarly, acute kidney injury (adjusted RR 3.41, 95% CI 3.34-3.43, P value <0.001), stroke (adjusted RR 5.46, 95% CI 5.31-5.62, P value <0.001), and sepsis (adjusted RR 6.57, 95% CI 6.40-6.75, Pvalue <0.001) were associated with significantly increased risk of mortality among patients with TTP (table 1).Conclusions: A significantly higher in-hospital mortality occurs among TTP patients admitted on the weekends as compared to weekdays. Future research should focus on identifying the underlying factors for this difference so that quality improvement measures could be taken to mitigate this difference.Table 1:Logistic Regression Analysis showing the adjusted relative risk (RR) of various patient and hospital characteristics in predicting in-hospital mortality for patients with TTP.VariableAdjusted RR95% CI of Adjusted RRP valueWeekend admission1.161.15-1.17<0.001Pay - Medicare - Medicaid - Private including HMO - self-pay - no charge - other1.0 1.33 1.19 1.63 1.36 2.02.. 1.28-1.38 1.14-1.25 1.50-1.77 1.11-1.67 1.73-2.36<0.001 <0.001 <0.001 <0.001 <0.001Race - white - black - hispanic - asian or pacific islander - native american - other1.0 1.01 0.93 1.13 1.05 1.070.98-1.03 0.89-0.97 1.07-1.19 0.94-1.16 1.02-1.130.47 0.003 <0.001 0.34 0.003Region - Northeast -Midwest - South - West1 0.92 1.05 0.970.86-0.98 0.99-1.11 0.91-1.040.01 0.06 0.48Co-morbidities - smoking - obesity - dyslipidemia - hypertension - diabetes mellitus - peripheral vascular disease - coronary artery disease - acute kidney injury - chronic kidney disease - stroke - sepsis0.90 0.78 0.60 0.68 0.99 1.32 1.06 3.41 1.10 5.46 6.570.88-0.92 0.76-0.79 0.59-0.61 0.67-0.69 0.97-1.00 1.29-1.34 1.05-1.07 3.34-3.43 1.08-1.11 5.31-5.62 6.40-6.75<0.001 <0.001 <0.001 <0.001 0.12 <0.001 <0.001 <0.001 <0.001 <0.001 <0.001Age1.041.043-1.046<0.001Female0.780.78-0.79<0.001Hospital Type - rural - urban non teaching - urban teaching1.0 0.92 1.050.88-0.97 0.99-1.110.002 0.061Bed size - small - medium - large0.95 1.010.89-1.01 0.96-1.070.11 0.51 DisclosuresNo relevant conflicts of interest to declare.

The splenic autoimmune response to ADAMTS13 in thrombotic thrombocytopenic purpura contains recurrent antigen-binding CDR3 motifs

AbstractAcquired thrombotic thrombocytopenic purpura (TTP) is the consequence of a severe ADAMTS13 deficiency resulting from autoantibodies inhibiting ADAMTS13 or accelerating its clearance. Despite the success of plasma exchange the risk of relapse is high. From 2 patients (A and B), splenectomized for recurrent episodes of acquired TTP, the splenic B-cell response against ADAMTS13 was characterized through generation of human monoclonal anti-ADAMTS13 autoantibodies (mAbs) by cloning an immunoglobulin G (IgG)4κ- and IgG4λ-Fab library using phage display technology and by Epstein-Barr virus transformation of switched memory B cells (CD19+/CD27+/IgG+). Sequence analysis of the anti-ADAMTS13 IgGs of both patients revealed that the VH gene use was limited in our patients to VH1-3 (55%), VH1-69 (17%), VH3-30 (7%), and VH4-28 (21%) and contained 8 unique and thus far not reported heavy-chain complementarity determining region 3 motifs, of which 4 were shared by the 2 patients. The discovery of several highly similar anti-ADAMTS13 autoantibodies in 2 unrelated TTP patients suggests that the autoimmune response is antigen driven, because the probability that such similar immunoglobulin rearrangements happen by chance is very low (<10−9).

Cardiac injury is a common postmortem finding in thrombotic thrombocytopenic purpura patients: is empiric cardiac monitoring and protection needed?

Thrombotic thrombocytopenic purpura (TTP) is a rare and potentially fatal disease. Early implementation of therapeutic plasma exchange (TPE) has decreased the mortality rate from >90% to <10%. However, fatalities still occur in these patients. The goal of this study was to characterize the causes of death and related postmortem findings in patients with TTP in the current era of emergent TPE to identify possible areas for improvement in the care of these patients. We analyzed clinical history, laboratory and histopathologic findings, and causes of death of patients with active TTP or TTP in clinical remission autopsied at our institution over 22 years. Of 18 patients, 15 had TTP judged to be a cause of death: it was an underlying cause of death in five cases, intermediate in three, and contributing in seven. The most common immediate causes of death were cardiac arrest and myocardial infarction. The most common TTP-related findings at autopsy were thrombi/emboli in heart (9), lung (11), brain (3), kidney (7), followed by hemorrhages in heart (7), lung (8), brain (2), kidney (7), and infarcts in heart (5), lung (4), brain (6) and kidney (3). Analysis of the cases with TTP as a cause of death suggests that the mechanism of death is commonly cardiac in origin. Proactive measures to monitor and protect the heart may be beneficial in these patients.

Clinical characteristics of systemic lupus erythematosus associated thrombotic thrombocytopenic purpura

We retrospectively reviewed the clinical records of 12 systemic lupus erythematosus (SLE) patients with thrombotic thrombocytopenic purpura (TTP) admitted into Peking Union Medical College Hospital (PUMCH) during October 2008 to September 2013. TTP occurred in 0.45% (12/2 693) of patients with SLE. The median age (2 males, 10 females) was 23 years. Their SLEDAI scores ranged from 12 to 30. All patients had significant thrombocytopenia and renal abnormalities. Ten of them had acute renal failure. Cardiac involvement and neurological deficits were observed in 9 and 8 of them respectively. Eight of them had injuries of more than 4 organs. Renal biopsy of 3 patients revealed lupus nephritis (type Ⅳ, n=2; type Ⅴ, n=1) and thrombotic microangiopathy. Eleven TTP patients received steroid pulse therapy (1 g/d, 3 days) and six of them survived. Six patients were rescued by therapeutic plasma exchange and 4 of them recovered. Among another 6 patients not on plasma exchange therapy, only 3 recovered. Finally 2 died, 3 gave up with exacerbated symptoms after therapy and 7 survived. Key words: Lupus erythematosus, systemic; Purpura, thrombotic thrombocytopenic

A novel flow‐based assay reveals discrepancies in ADAMTS‐13 inhibitor assessment as compared with a conventional clinical static assay

Several static Bethesda-type assays are routinely used to determine ADAMTS-13-neutralizing autoantibodies in acquired thrombotic thrombocytopenic purpura (TTP), but the inhibitory activity of these antibodies has not been thoroughly evaluated under the more physiologic condition of flow. We investigated whether ADAMTS-13 inhibitor assessment with the FRETS-VWF73 assay is predictive for evaluation under flow. Anti-ADAMTS-13 autoantibodies were purified from patients with acquired TTP by chromatography involving an ADAMTS-13 affinity matrix and/or proteinG. ADAMTS-13 activity was measured with the FRETS-VWF73 assay and a novel flow assay determining the ADAMTS-13-mediated decrease in platelet aggregate surface coverage, caused by perfusion of a suspension containing platelets, erythrocytes and von Willebrand factor (VWF) over a surface coated with extracellular matrix components. The neutralizing activities of ADAMTS-13 inhibitors were compared under static conditions and under flow by use of the two assays. The suitability of the flow-based ADAMTS-13 activity assay for quantification of ADAMTS-13 inhibitors could be demonstrated by reversibility of the ADAMTS-13-dependent decrease in surface coverage upon addition of goat ADAMTS-13 antiserum. Testing the neutralizing activity of purified autoantibodies from six patients in the flow assay according to their FRETS-VWF73-based inhibitor titers gave rise to vastly different inhibitory effects, indicating a discrepancy in inhibitor assessment between static and flow conditions. Anti-ADAMTS-13 autoantibodies may show inhibitory properties invivo that are not consistent with the ADAMTS-13 inhibitor levels determined in routine static assays, possibly because certain epitopes are selectively exposed under shear. Consequently, the course of disease and treatment efficacy may vary among TTP patients, despite common inhibitor titers.