Thrombocytopenic Purpura Patients Research Articles

Safety and Tolerability of M254 in Healthy Volunteers and Immune Thrombocytopenic Purpura (ITP) Patients

Safety and Tolerability of M254 in Healthy Volunteers and Immune Thrombocytopenic Purpura (ITP) Patients

Analysis of Clinical Features, Laboratory Characteristics and Therapeutic Outcome in Patients with Thrombotic Thrombocytopenic Purpura Treated at King Fahad Medical City, Riyadh, Saudi Arabia

BackgroundThrombotic thrombocytopenic purpura (TTP) is a life-threatening disease characterized by microvascular platelet deposition and thrombus formation with resulting microangiopathic hemolytic anemia and thrombocytopenia. Deficiency of the von Willebrand factor cleavage metalloprotease, also known as ADAMTS 13, has been implicated as an important etiological factor in TTP. Few small studies have been reported on Saudi patients with TTP until now. Our aim was to analyze the clinical features, laboratory characteristics and treatment outcomes with TTP patients treated at our large tertiary care center.MethodsThis is retrospective data of 24 patients with diagnosed of TTP who were treated at King Fahad Medical City, Riyadh, Saudi Arabia between October 2006 and April 2015. Patient suspected as a case of TTP on the basis of clinical features with the evidence of microangiopathic hemolysis and thrombocytopenia were included in this study although data related to pentad of TTP was collected and wherever logistically possible ADAMTS13 levels and inhibitor titer were determined. The primary aim was outcome assessment by overall response rate (ORR) in the treated patients through Kaplan-Meier method. Paired sample t-test was applied to determine the mean significant difference among platelets (plt), hemoglobin (Hgb) & LDH on day 1 and day 7 of treatment.ResultsTwenty-four TTP patients (18 females; 6 males) admitted to our hospital from 2006 to 2015 were analyzed. The mean age was 33.5±13.9 years. Twenty-two (91%) of the patients presented with neurologic features, seven (29%) had fever, ten (42%) had renal impairment that normalized with treatment and four (20.83%) had increased troponin-T or cardiac symptoms. There were 22 patients (91.7%) with the triad of TTP, including hemolytic anemia, thrombocytopenia and neurologic abnormalities; only 2 (8.2%) had the classical pentad of TTP. Among the plausible etiology, idiopathic (51.8%) was the most common followed by acquired autoimmune abnormalities (29.2%). Plasma ADAMTS 13 activity was determined in 19 patients. Eight patients (42.1%) had severe ADAMTS 13 deficiency (activity< 5%); 5 (26.3%) had moderate decrease of ADAMTS 13 activity (activity: 5-10%); another 3 (15.8%) had low ADAMTS 13 activity and 3 (15.8%) patients had normal ADAMTS 13 (>50%) most likely due to sampling post plasma infusion in emergency situations. Median platelet count on Day 1 was 14x10^9/L, and Day 7 was 119x10^9/L (P value< 0.001), Median Hgb on Day1 was 8.25 gm/dl and Day 7 was 9.35 gm/dl (P value< 0.007), Median LDH on Day 1 was 1211 IU/L and Day7 was 278.92 IU/L (P value< 0.001) respectively. All patients received plasma exchange whereas 23 (95.8%) patients received adjunctive corticosteroids. Five patients (20.8%) were early refractory to standard treatment with therapeutic plasma exchange (TPE). Thirteen (54.2%) patients received rituximab either due to refractoriness to TPE on ~ day 7, or earlier due to cardiac or neurological manifestations at treating physician’s discretion. Average hospital stay was 27 days (range 1-131). Twenty-one out of 24 (87.5%) achieved complete remission (CR) without any subsequent relapse. On long term follow up of 22 months (median, Range 1-113), overall survival was 80%. Three patients died during acute episode because of very sever disease or delayed arrival to our center. One patient died later on because of other comorbidities while in CR.ConclusionThrombotic thrombocytopenic purpura is a life threatening condition and immediate treatment with plasma exchange along with steroids and or rituximab was very effective in preventing high risk of mortality and achieving durable CR in 87.5% of our patients. Combination of very severe CNS manifestations and delayed arrival contributed to mortality significantly. More awareness is needed for early diagnosis and early referral to higher centers. DisclosuresNo relevant conflicts of interest to declare.

Risk Factors Associated with Multiple End-Organ Damage in Idiopathic Thrombotic Thrombocytopenic Purpura (TTP) Patients

Introduction: Idiopathic thrombotic thrombocytopenic purpura (TTP) is a rare hematological disorder caused by deficiency of ADAMTS13 activity and subsequent development of vWF-platelets-rich microthrombi leading to tissue ischemia and organ dysfunction. Standard treatment consists of daily plasma exchange (PEX) to remove the autoantibodies and replenishes ADAMTS13 activity and immunosuppressive therapy for patients with autoimmune process. Recent publication showed that caplacizumab, an anti-vWF humanized nanobody which inhibits the interaction between vWF multimers and platelets, can be used together with PEX to reduce time to platelet normalization but not risk of relapse when compared to placebo. However, incorporate caplacizumab with PEX in high risk TTP patients maybe even more important to reduce long-term organ damage. Therefore, we performed a retrospective single center analysis of TTP patients to identify the incidence and risk factors associated with end-organ damage. Method: Sixteen adult patients with idiopathic TTP were identified from January 2005 through December 2015. Idiopathic TTP was defined as thrombotic microangiopathy with an ADAMTS13 ≤10%. Cardiac involvement was defined as an increased Troponin I (TnI) level &gt;0.1µg L-1. Renal involvement was defined as serum creatinine &gt;1.30mg/dL. Of note, all the 16 patients had normal baseline renal function. Neurological involvement was defined as documentation of confusion, headaches, seizure, aphasia and syncope. The time to a platelet response was defined as a platelet count that was ≥150,000/mm3 for at least 48 hours. All patients were treated with PEX, corticosteroids, ± rituximab. Results: The mean (range) age was 48 (23 to 78) and the female: male ratio was 12:4. All (16) patients had end-organ involvements: 16 (100%) neurological, 6 (37.5%) renal, and 4 (25%) cardiac. Seven patients (43.75%) had single organ involvement (Group one), 9 patients (56.25%) had two or three organ involvement (Group two). The mean age for Group one was 44 as compared to 51 for Group two. The mean LDH level on admission was 987 IU/L (range: 417-1,289 IU/L) for Group one as compared to 1,715 IU/L (range: 336-4,455 IU/L) for Group two (p value: 0.06). The mean platelet count on admission was 20 k/µL for Group one as compared to 18 k/µL for Group two. The mean hemoglobin level on admission was 7.5g/dL (range: 5.3-8.6g/dL) for Group one as compared to 8.78g/dL (range: 8.1-10.5g/dL) for Group two. The median time to a platelet response was 5 days in this study population. Conclusions: Our study showed that more than 50% of TTP patients presented with multi-organ involvement. Very high LDH at initial presentation prior to PEX was identified as a probable single risk factor associated with multi-organ involvement. Prompt arrest of vWF-mediated platelet aggregation with agents such as calpalacizumab in TTP patients could prevent or minimize early irreversible end-organ damage due to tissue ischemia. Identifying risk factors for severe end-organ involvement will be important to study the potential benefit of incorporating calpalacizumab into TTP treatment. Further study on the risk factors for end-organ involvements in TTP patients and their long-term outcome at a multi-center level and with a larger patient population is warranted. Disclosures No relevant conflicts of interest to declare.

Study on correlation indexes with recurrence of acquired thrombotic thrombocytopenic purpura during the first remission

Objective To study the relationship between recurrence of acquired thrombotic thrombocytopenic purpura (TTP) patients in their first remission period and expression levels of a disintegrin and metallopeptidase with thrombospondin motif (ADAMTS)13 and anti-ADAMTS13 antibody. Methods From March 2008 to June 2014, a total of 37 patients with acquired TTP who were treated in Affiliated Hospital of Yan′an University and Fuping County Hospital of Weinan City in Shaanxi Province were included in this study. According to whether relapse at the first time in follow-up period, they were divided into study group (n=15) and control group (n=22). Residual collagen binding assays, ELISA, Western blotting method were used to detect the ADAMTS13 activity, levels of ADAMTS13 antigen, anti-ADAMTS13 antibody, ADAMTS13 inhibitor, von Willebrand factor (vWF) antigen and ultralarge von Willebrand factor (ULVWF), respectively. The above indexes between two groups were compared by statistical methods. Multivariate unconditional logistic regression analysis was used to evaluate the independent influencing factors of recurrence in acquired TTP patients during their first remission. The study protocol was approved by the Ethical Review Board of Investigation in Human Being of hospitals above. Informed consent was obtained from all participants. Results ① The median plasma ADAMTS13 activity of study group was 11% (7%~124%), which was lower than that of 53%(7%~151%) in control group, and there as significance difference between two groups (u=4.018, P<0.05). Plasma ADAMTS13 activity remarkably reduced percentage was 53.3%(8/15) in study group, which was higher than that of 22.7%(5/22) in control group, the differences was statistically significant (P=0.049). ② Plasma ADAMTS13 activity level of all 37 patients with acquired TTP was positively correlated with plasma ADAMTS13 antigen level (rs=0.810, P=0.001). The median plasma ADAMTS13 antigen level of study group was 33% (3%~99%), which was lower than that of 59% (3%~128%) in control group, and there was significance difference between two groups (u=4.121, P<0.05). Remarkably reduction percentage of plasma ADAMTS13 antigen level was 13.3%(8/15) of study group, which was much higher than that of 9.1% (2/22) in control group, the differences was statistically significant between two groups (P=0.008). ③ The detection rate of plasma anti-ADAMTS13 antibody of study group was 66.7% (10/15), which was higher than that of 36.4% (8/22) in control group, and the difference was statistically significant (P=0.007). ④ The detection rate of plasma anti-ADAMTS13 inhibitors of study group was 46.7% (7/15), which was higher than that of 18.2% (4/22) in control group, and the difference was statistically significant (P=0.011). ⑤ In study group, detection rate of ULVWF polymer was 20.0% (3/15), which was much higher than that of 13.6% (3/22) in control group, and the difference was statistically significant between the groups (P=0.042). ⑥ Analyzed correlation indexes by multivariate unconditional logistic regression analysis showed that the independent risk factors affected recurrence of acquired TTP patients during the first remission including, plasma ADAMTS13 activity remarkably reduced (OR=2.95, 95%CI: 1.13-6.96, P<0.05), presence of plasma anti-ADAMTS13 antibody (OR=3.31, 95%CI: 1.08-8.19, P<0.05), and presence of plasma anti-ADAMTS13 inhibitor (OR=3.24, 95%CI: 1.24-9.03, P<0.05). Conclusions For acquired TTP patients druing first remission period, the remarkably reduced of ADAMTS13 activity level, presence of anti-ADAMTS13 antibody and anti-ADAMTS13 inhibitor could significantly increase the risk of recurrence, which could be used as important indicators for predicting recurrence of acquired TTP patients during the first remission. Key words: Purpura, thrombotic thrombocytopenic; ADAMTS13 protein, human; Von Willebrand factor; Recurrence

Thrombotic Thrombocytopenic Purpura Associated with Bone Marrow Necrosis Complicating Metastatic Extra-Mammary Paget's Disease

As extra-mammary Paget's disease is rare and usually diagnosed at early stage when it is highly curable with surgical resection, it is much rarer to see patients with recurrent metastatic disease. Thrombotic thrombocytopenic purpura in patients with metastatic solid cancer is also a rare disease and may result from bone marrow metastasis or bone marrow necrosis. For the latter, the majority of cases are not eligible for systemic chemotherapy for rapid disease progression and poor performance status. Herein, authors report a patient with thrombotic thrombocytopenic purpura associated with bone marrow necrosis complicating extra-mammary Paget's disease who was successfully treated with docetaxel and carboplatin combination chemotherapy. Key Words: Extramammary Paget's disease; Bone marrow necrosis; Thrombotic thrombocytopenic purpura.

Anti-angiogenic effect of interferon on JAK2V617F positive myeloproliferative neoplasms and its anti-angiogenic mechanisms

To research the suppressive effect of interferon α2b (IFN-α2b) on angiogenesis of JAK2 tyrosine kinase gene point mutation (JAK2V617F) positive myeloproliferative neoplasm (MPN) and its anti-angiogenic mechanisms. (1) A total of 42 cases of JAK2V617F positive MPN patients in the No.1 Hospital of Baoding were selected between January 2012 and October 2014, including the newly diagnosed group before treatment (n=25) and the IFN-α2b treatment group (n=17). Ten cases of idiopathic immune thrombocytopenic purpura (ITP) patients were enrolled as controls. JAK2V617F/JAK2 ratio was detected by real-time fluorescent quantitative PCR to measure mutation; the expression levels of phosphorylated JAK2 (p-JAK2), vascular endothelial growth factor (VEGF), hypoxia-inducible factor-1α (HIF-1α) and CD105 marked microvascular density (MVD) in bone marrow were detected by immunohistochemistry. The correlations were analyzed among JAK2V617F mutation burden and VEGF, HIF-1α, and MVD. (2) Human erythroleukemia cell line HEL cells were treated with different concentrations of IFN-α2b. The apoptosis was detected by Hoechst staining method. The cell viability was detected by CCK-8 test. Cell migration ability was tested by transwell chambers. The protein expression levels of p-JAK2, VEGF, and HIF-1α were detected by Western blot. (1)The ratio of JAK2V617F/JAK2 in the IFN-α2b treatment group (22.69% ± 12.64%) was significantly lower than that in the newly diagnosed group (46.17% ± 19.32%) (P<0.01). The expression levels of p-JAK2, VEGF, and HIF-1α in the newly diagnosed group (82.41% ± 11.65%, 64.72% ± 25.01%, 45.12% ± 20.28%) were significantly higher than those in the IFN-α2b treatment group (60.93% ± 20.57%, 36.58% ± 15.95%, 32.15% ± 14.27%) and the control group (43.05% ± 12.59%, 25.69%± 1 3.75%, 25.07% ± 15.49%) (all P<0.01). MVD in the newly diagnosed group (26.58% ± 5.93%) was significantly higher than that in the treatment group (15.86%± 4.27%) and the control group (10.76% ± 4.01%) (P<0.01). Spearman correlation analysis showed positive correlation of JAK2V617F mutation with VEGF and MVD (r=0.589, P<0.05; r=0.577, P<0.05). (2)The apoptosis of HEL cells were increased after treated with 30 000 U/L of IFN-α2b in HEL cells after 24 h. The proliferation of HEL cell were inhibited by different concentrations of IFN-α2b in time- and dose-dependent manner. The number of membrane-permeating HEL cells after treated with 5 000 U/L of IFN-α2b in HEL cells after 24 h was significantly lower than that in the untreated cells (52.9 ± 7.5 vs 77.3 ± 6.1) (P<0.05). The expression levels of p-JAK2, VEGF, and HIF-1α were decreased in a dose-dependent manner. IFN-α2b may exert an anti-angiogenic effect by inhibiting the VEGF, HIF-1α, and MVD expression in MPN via JAK2 signal pathway.

Pulmonary Endarterectomy in a Patient with Immune Thrombocytopenic Purpura.

Immune thrombocytopenic purpura (ITP) patients are at high risk for bleeding complications regarding surgeries involving cardiopulmonary bypass. We report an ITP patient with chronic thromboembolic pulmonary hypertension who underwent uncomplicated pulmonary endarterectomy with receiving postoperative intravenous immunoglobulin (IVIG) therapy. The positive outcome of this case may suggest that pulmonary endarterectomy surgery is performed safely for ITP patients.

Cardiac Complications in Thrombotic Thrombocytopenic Purpura: Data from Nationwide Inpatient Sample

BackgroundThrombotic thrombocytopenic Purpura (TTP) has been reported to be associated with serious cardiac complications including arrhythmia, sudden cardiac death, myocardial infarction, cardiogenic shock, and heart failure. These complications are believed to be the sequelae of diffuse platelet thrombi leading to infarction in cardiac tissue. However, the true burden of cardiac complications in TTP in clinical practice remains unclear.MethodsWe used the 2009-2011 Nationwide Inpatient Sample database to identify hospitalizations in patients ≥18 years with a diagnosis of TTP (International Classification of Diseases, 9th revision, Clinical Modification [ICD-9-CM] code 446.6. Nationwide Inpatient Sample is the largest all-payer publicly available inpatient care database in the US. It contains data from five to eight million hospital stays from about 1,000 hospitals across the country and approximates a 20% sample of all US hospitals. ICD-9-CM codes were used to identify any of the following coded cardiac complications- 1) arrhythmias, 2) acute myocardial infarction (MI), 3) cardiogenic shock, 4) acute heart failure, and 5) conduction abnormalities. Univariate and multivariate logistic regression was used to determine the odds of various cardiac complications in patients with TTP compared to patients without TTP. Univariate analysis was done to compare baseline demographic and hospital characteristics between patients among TTP patients with and without cardiac complications. Data analysis was done using STATA version 13.0 (College Station, TX).ResultsOf the estimated total 4,367 TTP hospitalizations, 22.2 % (n=969) developed at least 1 of the above cardiac complications. Mean age of our cohort was 56.2±17.3, with the majority of patients being white (55.1%). Compared to those with TTP and no reported cardiac events, dyslipidemia (p=0.01) hypertension (p=0.001), peripheral vascular disease (p=0.03), acute kidney injury (p<0.001), chronic kidney disease (p=0.003), stroke (p=0.01), sepsis (p=0.01) and coronary artery disease (p<0.001) were more likely in patients with cardiac complications (Table 1). On multivariate analysis, patients with TTP were found to develop acute MI more commonly than patients without TTP (OR 5.22, 95% CI 3.81-7.15; p<0.001). Interestingly, acute heart failure was found to be less common in TTP patients (OR 0.7, 95% CI 0.55-0.90; p=0.002). There was no significant difference in the incidence of arrhythmias, cardiogenic shock and conduction disorders among patients with TTP compared to patient without TTP (Figure 1).ConclusionIn this study of large national database, acute MI was found to be the most common cardiac complication in patients with TTP. Although reported, arrhythmias, cardiogenic shock and conduction disorders were not significantly associated with TTP.TableCharacteristicTTP without Cardiac Complications (n=3,398)TTP with Cardiac Complications (n=969)pAge, mean ± SD44.5±15.256.2±17.3<0.001Sex0.094Male31.338.7Female68.761.3Race0.009White41.855.1Black45.133.5Hispanic8.34Asian or Pacific Islander1.13.5Native American0.70.6Other33.3Primary Payer<0.001Medicare18.937.8Medicaid23.914.9Private insurance42.437.4Self-pay10.15No charge0.72Other42.9Region0.520Northeast15.618.2Midwest26.727.9South45.639.5West12.114.4ComorbiditiesSmoking26.826.20.871Obesity12.19.10.210Dyslipidemia13.221.90.009Hypertension50.362.90.001DM23.527.90.227PVD1.14.40.029CAD5.115.4<0.001AKI35.561.1<0.001CKD16.527.30.003Stroke4.911.80.007Sepsis3.99.70.01AKI = Acute Kidney Injury; CAD = Coronary Artery Disease; CKD = Chronic Kidney Disease; DM = Diabetes Mellitus; PVD = Peripheral Vascular Disease [Display omitted] 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.

Evaluation of a chromogenic commercial assay using VWF-73 peptide for ADAMTS13 activity measurement

IntroductionThrombotic thrombocytopenic purpura (TTP) is a thrombotic microangiopathy (TMA), related to a severe functional deficiency of ADAMTS13 activity (<10% of normal). ADAMTS13 activity is thus crucial to confirm the clinical suspicion of TTP, to distinguish it from other TMAs, and to perform the follow-up of TTP patients. Material and methodsWe compared the performance of the commercial chromogenic assay Technozym® ADAMTS13 Activity ELISA (chromogenic VWF73 substrate, Chr-VWF73, Technoclone, Vienna, Austria), to that of our in-house FRETS-VWF73 used as reference method. A large group of 247 subjects (30 healthy volunteers and 217 patients with miscellaneaous TMAs) was studied. ResultsThe lower limit of detection of the Chr-VWF73 was 3%, which is well adapted to the clinically relevant threshold for TTP diagnosis (10%). Our results showed a reasonable agreement between FRETS-VWF73 and Chr-VWF73 assays to distinguish samples with an ADAMTS13 activity <10% from those with an ADAMTS13 activity >10%. However, Chr-VWF73 assay provided false negative results in ~12% of acute TTP patients. Inversely, the Chr-VWF73 assay globally underestimated ADAMTS13 activity in detectable values ranging from 11 to 100% (with a great variability compared to FRETS-VWF73), which may be a concern for the follow-up of TTP patients in remission. ConclusionIn-house assays developed and performed by expert laboratories remain the reference methods that should be used without limitation to control values provided by commercial assays when needed. Also, the development of an international reference preparation will be crucial to improve standardization.

Enhanced phagocytosis of auto antibodies-opsonized blood cells by macrophages from virally infected mice. (INM3P.440)

Abstract Autoimmune haemolytic anemia and idiopathic thrombocytopenic purpura are common immune disorders resulting from the phagocytosis of autoantibodies-opsonized blood cells. It has been reported that viral infection may be associated with autoimmune haemolytic anemia and idiopathic thrombocytopenic purpura in patients and may incriminate IgG and IgM antibody isotypes. Lactate Dehydrogenase-elevating Virus triggers the activation of immune cells although it does not induce usually any pathology in mice. Here we report that LDV infection increases the pathogenicity of IgM anti-platelet mAbs which is mediated by FcαμR. Interestingly, we show that this mechanism results from an enhanced Fc-dependent phagocytosis by macrophages rather than an increased expression of FcαμR. Our results also indicate that the pathogenicity of IgG anti-red blood cells is increased in virally infected mice. To date, three activating FcγR have been identified in mice, binding IgGs with different affinity : FcγRI, FcγRIII and FcγRIV. Since FcγRI has a high affinity for monomeric IgG2a, this receptor is poorly available for the binding of circulating-opsonized IgG2a. However we report an increased expression of each receptor by macrophages in contrast to FcamR whose expression is not regulated. Furthermore, we show the functional involment of these three receptors in the exacerbation of IgG-mediated anemia.

A8.14 Effect of rituximab on B-cell phenotypes and on B-cell activating factor receptor expression in patients with thrombotic thrombocytopenic purpura

Background and AimsThrombotic Thrombocytopenic Purpura (TTP) is due to deficiency or dysfunction of the metalloproteinase ADAMTS13, which cleaves von Willebrand factor. This results in increased platelet adhesion and thrombus formation...

Utilization Of ADAMTS13 Testing In Suspected Thrombotic Thrombocytopenic Purpura (TTP)

IntroductionThrombotic thrombocytopenic purpura (TTP) is a rare but life-threatening condition, characterized by thrombotic microangiopathy (TMA) that results in microangiopathic hemolytic anemia (MAHA), thrombocytopenia and clinical symptoms including altered mental status, renal impairment and fever. Diagnosing TTP in acute setting continues to be a challenge, since the clinical pentad is not consistently present in TTP, making it difficult to differentiate from other conditions with TMA. The initial clinical decision for plasma exchange (PEX) is supported if there are MAHA and thrombocytopenia without a clear alternative cause. Acquired autoimmune deficiency of ADAMTS13 is known to cause idiopathic TTP and forms a basis for success of PEX in many patients by replenishing ADAMTS13 in addition to immunosuppressive therapy. However, low levels of ADAMTS13 alone are known to be neither sufficiently sensitive nor specific for diagnosis of TTP in acute setting. The aims of this retrospective study were to evaluate: (i) the utilization of ADAMTS13 testing in suspected TTP; (ii) the utilization of PEX in patients with TTP diagnosis; (iii) the association of low ADAMTS13 activity with early mortality in TTP; and (iv) the incidence of ADAMTS13 deficiency in non-TTP diagnoses. MethodsAfter IRB approval, we identified 49 adult patients at a single tertiary medical center, who had ADAMTS13 testing between 2005 and June 2013 for suspected TTP. We searched the final diagnosis, either TTP or non-TTP. The results of ADAMTS13 testing and use of PEX were reviewed in the TTP and non-TTP patients. We identified the final diagnosis of the non-TTP patients. We assessed association of severely low ADAMTS13 activity (<10%) with early mortality defined as death within 3 months of diagnosis in patients with TTP diagnosis. ResultsOf the 49 patients who had ADAMTS13 testing 18 (37%) were male. The median age was 51years (range 20-81). All patients with TTP had PEX except one who could not initiate PEX due to early death. Twenty-five of the 30 patients (83%) with TTP had low ADAMTS13 activity and 17 of the 30 patients (57%) had severe deficiency (<10%). Five of the 30 patients with TTP suffered early mortality. The relative risk of death among TTP patients with severely low ADAMTS13 activity (<10%) was 1.15 (95% CI 0.22-5.90) compared to TTP patients with low to normal ADAMTS13 activity. Nineteen of the 49 patients who had ADAMTS13 testing subsequently had non-TTP diagnosis. Eight of these 19 patients with non-TTP diagnoses were found to have low ADAMTS13 activity: two patients with DIC secondary to sepsis; two patients with thrombocytopenia associated with primary presentation of hemophagocytic lymphohistiocytosis (HLH); one patient each with malignancy associated with TMA, vasculitis due to autoimmune disease, hematopoietic stem cell transplantation associated TMA and sepsis with renal failure; None of the eight patients with non-TTP diagnosis had severe deficiency. Four patients with non-TTP diagnosis underwent PEX and all had normal ADAMTS13 activity. Three of the 4 patients were subsequently diagnosed with drug induced TMA after oral opioid injection and one with hemolytic uremic syndrome. ConclusionsPatients with a strong suspicion of TTP should be treated with PEX. ADAMTS13 activity is found to be relatively specific for TTP, being supportive of the clinical diagnosis in majority of cases. There was no statistically significant correlation between death and severely low ADAMTS13 activity in the cohort of TTP patients studied. ADAMTS13 activity can be low in a variety of non-TTP conditions. Its utility in diagnosis and prognosis of non-TTP conditions, especially HLH, needs further evaluation. Disclosures:No relevant conflicts of interest to declare.

ADAMTS13 activity and genetic mutations in Japan

Thrombotic thrombocytopenic purpura (TTP), a life threatening disease, can be induced by congenital or acquired deficiency of plasma metalloprotease ADAMTS13. Since the publication of the first genetic analysis in patients with congenital ADAMTS13 deficiency in 2001, more than 100 genetic defects in the ADAMTS13 gene have been reported worldwide. Genetic analysis in patients with ADAMTS13 deficiency has greatly contributed to the understanding of the etiology of TTP. A rapid and quantitative assay method for the plasma ADAMTS13 activity was developed recently in 2005 and opened a new area of TTP research - namely genetic research using a general population to evaluate age and gender differences of ADAMTS13 activity as well as phenotype - genotype correlations of genetic polymorphisms and estimation of a homozygote or a compound heterozygote ADAMTS13 deficiencies. The Japanese general population study included 3616 individuals with an age between 30 - 80 years confirming other studies that while ADAMTS13 activity decreased with age, VWF antigen increased and VWF antigen levels are lowest in blood group O indviduals, whereas ADAMTS13 activity levels were not associated with the AB0 blood group. 25 polymorphisms with a minor allele frequency of more than 0.01 were found, among them 6 missense mutations and 19 synonymous mutations, except P475S missense polymorphisms that was only idenitified in an East Asian population, characterized by reduced ADAMTS13 activity. Prevalence of congenital ADAMTS13 deficiency in the Japanese population was estimated about one individual in 1.1 x 106 to be homozygote or compound heterozygote for ADAMTS13 deficiency. So far more than 40 mutations in Japanese congenital TTP patients were found, but R193W, Q449*, C754Afs*24 (c.2259delA) and C908Y were identified in more than four patients suggesting the precipitaion of these mutations in the Japanese population.

The value of anti-von Willebrand factor antibody in the mechanism of idiopathic thrombotic thrombocytopenic purpura

Objective This study is aimed at determining whether anti-von Willebrand factor (VWF) autoantibodies are present in the plasma of idiopathic thrombotic thrombocytopenic purpura (TTP) patients with normal ADAMTS13 activity and undetectable anti-ADAMTS13 antibodies, and at examining whether murine monoclonal antibodies (mAbs) against human VWF decrease the susceptibility of VWF to ADAMTS13 in vitro. Methods Anti-VWF autoantibodies and ultralarge VWF (UL-VWF) multimers were measured in plasma samples of 53 adult patients with idiopathic TTP by enzyme-linked immunosorbent assay and sodium dodecylsulphate-agarose gel electrophoresis, respectively.Moreover, the effects of eight murine mAbs to different human VWF domains on VWF cleavage by ADAMTS13 were evaluated under fluid shear stress and static/denaturing conditions, respectively. Results Anti-VWF antibodies and UL-VWF multimers were detected in two TTP patients with normal ADAMTS13 activity and undetectable anti-ADAMTS13 antibodies.The SZ34, an anti-VWF mAb, inhibited VWF proteolysis mediated by ADAMTS13 under flow, but not static conditions. Conclusion Anti-VWF antibody may be one of the causes of idiopathic TTP with normal ADAMTS13 activity and undetectable anti-ADAMTS13 antibodies.（Chin J Lab Med,2012,35:1118-1124） Key words: Purpura; thrombocytopenic; idiopathic; von Willebrand factor; Autoantibodies; ADAM proteins

Delayed visual recovery in pregnancy-associated thrombotic thrombocytopenic purpura with bilateral serous retinal detachment

To present a pregnancy-associated thrombotic thrombocytopenic purpura (TTP) patient with bilateral serous retinal detachment (SRD). Case report. A 28-year-old nulliparous woman with 31 weeks gestation was presented to the local hospital with preeclampsia, hemolytic anemia, thrombocytopenia and bilateral blurry vision. Funduscopic examination showed bilateral macular SRD. Within the first month after delivery of a live female baby via cesarean section (at 32 weeks gestation), the patient experienced a recurrent course of hemolytic anemia and thrombocytopenia, and was then transferred to our hospital. On admission, her best corrected visual acuity (BCVA) was 0.1 OU; optical coherence tomography (OCT) confirmed the presence of bilateral macular SRD; electroretinography (ERG) examination showed diminished rod responses with reduced a and b waves in cone and mixed rod-cone responses. She was ultimately diagnosed with TTP and was treated systemically with fresh frozen plasma, rituximab, prednisone and cyclophosphamide. Despite persistent visual disturbances, she was discharged 1 month after admission with stabilization of systemic manifestations. At her first follow-up visit 6 months after discharge, surprisingly, her BCVA had improved to 1.0 OU; fundus examination and OCT confirmed the complete resolution of bilateral macular SRD and ERG revealed subnormal (right) and normal (left) electrophysiological responses. We believe that in this case, the clinical context (pregnancy) in which TTP developed, the unreported ERG characteristics and the unexpected delayed visual recovery are worth reporting. TTP should be considered as a potential differential diagnosis in patients with pregnancy-associated SRD. Appropriate systemic treatment might be mandatory for visual recovery.

Changes and Clinical Significances of Th17 Cells and Their Secretive IL-17 in Patients With Idiopathic Thrombocytopenic Purpura

Objective To investigate the changes and clinical significances of T cells helper (Th) 17 cells and interleukin (IL)-17 of idiopathic thrombocytopenic purpura (ITP) patients before and after glucocorticoid treatment.Methods From November 2010 to January 2012,a total of 38 cases of ITP patients who received treatments in First Affiliated Hospital of Dalian Medical University were included into this study,as study group.A total of 30 healthy adults who received medical examination at the same period were included into control group.There was no significant difference between two groups among age,gender and other general information (P＞0.05).Flow cytometry (FCM) was used to detect the rate of Th17 cells in ITP patients.The levels of serum IL 17 in ITP patients were measured by enzyme-linked immunosorbent assays (ELISA) (The study protocol was approved by the Ethical Review Board of First Affiliated Hospital of Dalian Medical University).Results Before treatment,the rate of Th17 cells and the levels of serum IL-17 in study group were significantly higher than that in control group [(1.38±0.42) ％,(25.52±6.83) ng/L vs.(0.71±0.29) ％,(8.23±3.15) ng/L] (P＜0.05).The percentage of Th17 cells [(1.05±0.31)％] was significantly decreased after treatment (P＜0.05) and the same tendency was also found in the levels of serum IL-17[(17.46±5.14) ng/L](P＜0.05).Conclusions The rate of Th17 cells and the levels of serum IL-17 significantly elevate in ITP patients.Glucocorticoid might play a role in ITP treatment by down regulating Th17 cells number and concentrations of IL-17. Key words: idiopathic thrombocytopenic purpura; Th17 cells; interleukin-17

Complications of Pregnancy in Women with Thrombotic Thrombocytopenic Purpura

AbstractAbstract 3322 BackgroundThrombotic Thrombocytopenic Purpura (TTP) occurring in association with pregnancy or puerperium accounts for 12–25% of all TTP acute episodes. Pregnancy leads to acute TTP in women affected by congenital TTP in the absence of periodic prophylactic plasma infusions, while the risk of acute TTP during pregnancy for women with the acquired form is not well known. Moreover, it is not known whether the presence of anti-ADAMTS13 antibodies that characterize acquired TTP affect the outcome of subsequent pregnancies. The aim of this study was to evaluate maternal-foetal outcome of pregnancies started after the diagnosis of TTP. MethodsWe analyzed clinical and laboratory features of 25 pregnancies of 22 women with TTP (all acquired TTP) out of 320 TTP patients in our cohort, all referred to the Milan TTP Registry, Milan (Italy), from 1994 to 2012. We tested the available biological samples for ADAMTS13 activity using FRET method, anti-ADAMTS13 autoantibodies by Western Blotting and ultra-large von Willebrand Factor (ULVWF) multimers ratio. ResultsWe found that 18 out of 25 pregnancies (72%) were complicated by either TTP recurrence (11/25, 44%) or spontaneous abortion in the first trimester (7/25, 28%). The incidence of TTP recurrence was 0.02 cases/week gestation (median duration of pregnancy at event: 32 weeks). The incidence of spontaneous abortion was 0.01 cases/week gestation (median duration of pregnancy at event: 6 weeks). Women’s parity was associated with spontaneous abortion, with a relative rate of 2.8 (95% confidence interval: 0.5–14.2) for multigravidae versus primigravidae.Interestingly, almost all miscarriages (6/7, 86%) occurred in women who experienced a pregnancy-related TTP episode during a previous pregnancy. To understand if this high rate of spontaneous abortion could be related to TTP, we analyzed ADAMTS13 activity levels, anti-ADAMTS13 antibodies and ULVWF multimers pattern. In the pregnancies complicated by TTP relapse, ADAMTS13 activity was severely reduced in the acute phase, in association with the presence of antiADAMTS13 antibodies and reduction of ULVWF multimers (ULVWF ratio < 0.85); in the cases of pregnancies complicated by spontaneous abortion, the mean ADAMTS13 activity level in the first trimester was 31%, with the presence of antiADAMTS13 antibodies and excess of ULVWF multimers (ULVWF ratio > 1.21); in the group of uncomplicated pregnancies, the mean ADAMTS13 activity levels was 97% in the first trimester and remained > 35% until delivery, with absence of antiADAMTS13 antibodies and normal ULVWF multimers. ConclusionsObstetric complications are frequent during pregnancies in women affected with acquired TTP. ADAMTS13 activity levels > 35% in the absence of antiADAMTS13 antibodies seem to confer little or no risk, while lower ADAMTS13 activity levels and the presence of antiADAMTS13 antibodies during pregnancy are predictive of poor gravidic outcome, either with acute TTP or spontaneous abortion in the first trimester. Surprisingly, although confidence intervals were wide, miscarriage rates were highest in multigravidae. Pre-gravidic and gravidic monitoring of ADAMTS13 activity levels and anti-ADAMTS13 autoantibodies is crucial in the management of pregnancies in TTP patients. Disclosures:No relevant conflicts of interest to declare.

The Influence of Race and ADAMTS13 Status On Outcomes in Thrombotic Thrombocytopenic Purpura

Abstract 4638Thrombotic thrombocytopenic purpura (TTP), a syndrome characterized by thrombocytopenia and microangiopathic hemolytic anemia due to deficiency of the von Willebrand factor cleaving protease ADAMTS13, was once a highly fatal condition, but therapeutic plasma exchange (PEX) has dramatically improved outcomes. We report the results of a retrospective chart review of TTP patients treated with PEX at our institution from September 2006 through December 2011. Complete demographic, clinical, laboratory, treatment, and response data were collected to examine the influence of race on clinical outcomes.A total of 49 patients were treated for TTP. This cohort was predominantly female (71%) and African American (61%). Three patients were of Hispanic ethnicity. The median age was 50 years (14–81), and the median number of comorbidities at presentation was 4 (range 0–14). Assays for ADAMTS13 activity with reflex to ADAMTS13 inhibitor were performed by the Blood Center of Wisconsin. All patients were treated with PEX (1–1.5 plasma volume using fresh frozen plasma as replacement fluid) once daily until plateau in the platelet count, and then PEX frequency was tapered over 2 weeks in remitting patients.The median number of PEX procedures for the first episode was 8 (1–22). The median total PEX for all episodes was 9 (1–56). The majority of patients, 37 (75.5%), received corticosteroids, and rituximab was administered to 19 (38.8%) of patients at some point during their treatment, usually for slow response or relapse.An initial response to PEX, defined by normalization of platelet count (above 140×109/L) and resolution of signs of hemolysis, was achieved in 36/49 (73.5%) patients. The response rate was not significantly different between African Americans (73.3%) and Caucasians (68.8%, p=0.742), and all 3 Hispanic patients responded to initial treatment. Pre-treatment ADAMTS13 activity was severely deficient (<10% of normal) in 20/39 (51.3%) patients. An inhibitor of ADAMTS 13 was detected (>0.4 Inhibitor Units) in 19/23 (82.6%) patients tested. African Americans were more likely to have severely deficient ADAMTS13 activity (16/25, 64%) than Caucasians (2/13, 15.4%; p=0.004).The initial response rate was higher in patients with severely deficient ADAMTS 13 activity (90.0%) in comparison to those with >10% activity (68.4%), however the difference did not reach statistical significance (p=0.067). Of the patients with severely deficient ADAMTS 13 activity and detectable inhibitor levels, 89.5% had an initial response to PEX. All four patients with severely deficient ADAMTS 13 activity but undetectable inhibitor levels also responded to PEX.The total number of relapse events was 16 (32.7%), and the median time to relapse was 19.4 months. All three Hispanic patients, 4 (25%) Caucasians, and 9 (30%) African Americans relapsed. One-year relapse free survival was similar among Caucasians and African Americans at 72% and 75% respectively (p=0.852). All three Hispanic patients relapsed within the first year. One-year overall survival was similar for Caucasians and African Americans at 81.3% and 83.3% respectively (p=0.968). A total of 10 deaths were documented. The causes of death were relapse of TTP (3/10), sepsis (4/10), GI bleeding (2/10), and unknown etiology (1/10).In this unusual TTP cohort with 61% African Americans, we found that African Americans were more likely to have severely deficient ADAMTS13 activity than Caucasians, but response to PEX, relapse, and survival for Caucasian and African Americans were very similar. This study also supports pre-treatment assays for ADAMTS13 activity and its inhibitor, not only for the diagnosis of TTP, but also for predicting response to PEX therapy. Disclosures:No relevant conflicts of interest to declare.

C0128 Decrease of microparticles and hemostasis improvement after the treatment in hemophilia A. coincidence or causation?

thrombocytopenic purpura or cardiovascular disease patients and from healthy subjects. MPswere also obtained from purified human blood cell subpopulations. Identification of plasminogen activators on MPs was performed by zymography and their capacity to generate plasmin by chromogenic assay. Results: Circulating MPs isolated from patientsgenerate a range of plasmin activity at their surface. This property was related to a variable content in uPA and/or tPA. Using distinct MP subpopulations derived from endothelial cells, platelets, leukocytes and erythrocytes, we demonstrated that plasmin is generated on endothelialand leukocytederived MPs, but is absent on MPs from platelet or erythrocyte origins. Leukocyte-derived MPs bear uPA and its receptor uPAR whereas endothelial-derived MPs carry tPA and tPA/inhibitor complexes. Comment: Endothelialand leukocyte-derivedMPs, bearing respectively tPA or uPA, support at least a part of fibrinolytic activity in the circulation that is modulated in pathological settings. This blood-borne fibrinolytic activity conveyed by MPs provides a more comprehensive view on the role of MPs in the haemostatic equilibrium and put forward the basis for a potential new biomarker.