Plasma-derived factor VIIa and factor X mixture agent (MC710) prophylaxis in haemophilia B patients with inhibitors.

INTRODUCTION: Haemophilia B patients with factor IX inhibitors have particularly unmet needs for conventional therapy. AIM: Phase II/III clinical trial, multicentre, open-label, prospective, self-controlled study was conducted to assess MC710 prophylaxis in haemophilia B patients with inhibitors. METHODS: We enrolled haemophilia patients who had received episodic or prophylactic treatment with bypassing agents up to that time. The participants continued their conventional therapy for 24 weeks and then MC710 was prophylactically infused intravenously every 2 or 3 days at 60 to 120 µg as FVIIa per kilogram of body weight for 24 weeks. The primary endpoint was the annual bleeding rate (ABR) requiring bypassing agents, which was compared intraindividually between the conventional therapy period and the MC710 prophylaxis period. RESULTS: A total of 11 male haemophilia B patients were enrolled. The median ABR ratio for each participant (the prophylaxis period ABR divided by the conventional therapy period ABR) was .33 (2.1/6.5), range from .00 to 3.77. ABR ratios for 9 of the 11 patients ranged from .00 to .60, and 3 of the 9 patients had zero bleeding events during the prophylaxis period. Meanwhile, ABR ratios for the remaining two patients were 2.53 and 3.77, respectively. Although a fibrinogen decrease recovered by the dose reduction was reported for only one participant as the sole adverse drug reaction in this study, no thrombotic events or other safety concerns were reported. CONCLUSION: MC710 prophylaxis is considered to be decrease the bleeding rate in haemophilia B patients with inhibitors without safety concerns.

Clinical conditions and risk factors for inhibitor-development in patients with haemophilia: A decade-long prospective cohort study in Japan, J-HIS2 (Japan Hemophilia Inhibitor Study 2).

BACKGROUND: Inhibitor-development is a serious complication in patients with haemophilia (PwH). Previous studies reported that therapeutic and genetic factors could be associated with these alloantibodies. Relevant clinical features such as genetic-background and different treatment regimens in Japan remain unclear, however. AIMS: To analyse a nation-wide Japanese registry for PwH, and to examine risk factors for inhibitor-development. METHODS AND RESULTS: Newly diagnosed patients with haemophilia A (PwHA) or haemophilia B (PwHB) without inhibitors after 2007, and with treatment records traceable from 0 to 75 exposure days (ED), were enrolled in the Japan Hemophilia Inhibitor Study 2 (J-HIS2) initiated in 2008. Of 417 patients (340 PwHA, 77 PwHB) from 46 facilities, 83 (76 PwHA, 7 PwHB) were recorded with inhibitors by July 2020. Inhibitors were observed in 31.0% of severe PwHA, 8.0% moderate and 1.6% mild and in 17.1% of severe PwHB. The majority of inhibitors (89.7% in severe PwHA and 71.4% in severe PwHB) were detected on or before 25ED (median 12ED in PwHA and 19ED in PwHB). Genotyping in these severe patients identified an association between inhibitor-development and null variants of F8 (P < .01) or F9 (P < .05). A lower incidence of inhibitors was recorded in severe PwHA treated with prophylaxis than in those treated on-demand (P < .01). A past-history of intracranial-haemorrhage appeared to be associated with inhibitor-development, while FVIII-concentrates infusion and routine vaccination on the same day was not related to inhibitor-development. CONCLUSION: The J-HIS2 study has identified significant clinical variables associated with inhibitor-development in Japanese PwH, consistent with other global studies.

[Acquired hemophilia A with reappearance of inhibitor 10 years after the completion of immunosuppressive therapy].

Acquired hemophilia A (AHA) is a disease that causes severe bleeding with the appearance of an inhibitor (INH) against blood coagulation factor VIII (FVIII). The prevalence of this condition is low; it occurs in only one in one to four million people per year; however, the number of diagnosed cases has increased in recent years owing to the greater awareness of the disease. It is noteworthy that this is a hemorrhagic disease that suddenly develops in the elderly. AHA treatment is divided into hemostatic treatment for bleeding and immunosuppressive therapy (IST) for removing FVIII-INH. As long as FVIII-INH remains, there is a risk of fatal bleeding; therefore, it is desirable to start IST soon after diagnosis. However, the use of immunosuppressive drugs for the elderly is often challenging due to concerns about adverse events, such as infectious diseases that have a considerable impact on prognosis. Ten years after the end of IST, we managed the case of a patient with AHA who had a relapse of FVIII-INH at the age of 84 years. In this case, relapse was detected early when there was no bleeding symptom, and remission was rapidly achieved with a small amount of IST without any adverse effects. There are few reports on AHA relapse; we believe that the present report will contribute meaningfully to the literature on this subject and would be useful when considering the long-term management of AHA.

Postmarketing safety and effectiveness of recombinant factor IX (nonacog alfa) in Japanese patients with haemophilia B.

INTRODUCTION: In 2010, nonacog alfa became the first recombinant factor IX (rFIX) available in Japan for patients with haemophilia B. AIM: To determine real-world safety (adverse events, incidence of inhibitors) and effectiveness of nonacog alfa in Japan. METHODS: This multicentre, prospective, observational, postmarketing surveillance study enrolled previously treated and untreated patients (PTPs and PUPs, respectively) who were observed for 1 and 2 years, respectively, after initiating nonacog alfa therapy. Safety and effectiveness were assessed for each treatment type. Annualized bleeding rate (ABR) and incremental recovery of rFIX were also evaluated. RESULTS: Overall, 312 of 314 patients enrolled from 173 sites were eligible for the safety analysis set (PTPs, 281; PUPs, 28; other, 3). Mean age was 25.4 (PTPs) and 14.8 (PUPs) years. Haemophilic severity ranged from mild to severe, and 133 (42.6%) patients had haemophilic arthropathy. Of 285 patients (PTPs, 257; PUPs, 28) in the effectiveness set, 112 received on-demand treatment for 1161 bleeding episodes (effectiveness rate, 93.7%) and 185 received routine prophylaxis (effectiveness rate, 95.5%). No spontaneous bleeding was observed in 52.4% of patients during prophylactic treatment. Median ABR was lower during routine prophylaxis (2.0) vs the rest of the observation period (8.3). A weak negative correlation was found between body weight and the reciprocal of rFIX recovery. Eleven adverse drug reactions occurred in 7 PTPs (2.2% [7/312]); recurrence of inhibitor was observed in 1 patient, but no new inhibitor developed in PTPs or PUPs. CONCLUSION: Nonacog alfa therapy is safe and effective in the real-world scenario in Japan.

[A Pediatric Case of Acquired Hemophilia A: The Usefulness of the Activated Partial Thromboplastin Time (APTT) Cross-Mixing Test for Early Diagnosis].

Acquired hemophilia A (AHA), a bleeding disorder caused by autoantibodies against FVIII, has the potential for life-threatening bleeding. The annual onset rate is said to be one in 4 million people, but diagnosis examples increase in adults because a disorder concept penetrated. AHA is quite rare in children, with an incidence rate of 0.045 per 1 million, but early detection is crucial because serious bleeding can happen, as in adults. We report a pediatric case who received an early diagnosis of AHA by an activated partial thromboplastin time (APTT) cross-mixing test. The 12-year-old girl had neither a past history nor a family history of bleeding episodes. She presented with intramuscular bleeding and epistaxis without trauma or medication. At diagnosis, her blood test showed prolonged APTT. Other hemostatic tests, such as the platelet count, prothrombin time and fibrinogen concentration, were within the normal range. We administered an APTT cross-mixing test that detected an inhibitor pattern and inhibitory antibodies against factors VIII. As a result, we administered prednisolone and the inhibitor disappeared after 1.5 months. In conclusion, AHA is a bleeding disorder which should be considered even in children due to the potential for life-threatening bleeding. Furthermore, the APTT cross-mixing test is useful for screening coagulation factor deficiencies and inhibitors.

ACTN1 mutations cause congenital macrothrombocytopenia.

Congenital macrothrombocytopenia (CMTP) is a heterogeneous group of rare platelet disorders characterized by a congenital reduction of platelet counts and abnormally large platelets, for which CMTP-causing mutations are only found in approximately half the cases. We herein performed whole-exome sequencing and targeted Sanger sequencing to identify mutations that cause CMTP, in which a dominant mode of transmission had been suspected but for which no known responsible mutations have been documented. In 13 Japanese CMTP-affected pedigrees, we identified six (46%) affected by ACTN1 variants cosegregating with CMTP. In the entire cohort, ACNT1 variants accounted for 5.5% of the dominant forms of CMTP cases and represented the fourth most common cause in Japanese individuals. Individuals with ACTN1 variants presented with moderate macrothrombocytopenia with anisocytosis but were either asymptomatic or had only a modest bleeding tendency. ACTN1 encodes α-actinin-1, a member of the actin-crosslinking protein superfamily that participates in the organization of the cytoskeleton. In vitro transfection experiments in Chinese hamster ovary cells demonstrated that altered α-actinin-1 disrupted the normal actin-based cytoskeletal structure. Moreover, transduction of mouse fetal liver-derived megakaryocytes with disease-associated ACTN1 variants caused a disorganized actin-based cytoskeleton in megakaryocytes, resulting in the production of abnormally large proplatelet tips, which were reduced in number. Our findings provide an insight into the pathogenesis of CMTP.

Thrombosis from a prothrombin mutation conveying antithrombin resistance.

We identified a novel mechanism of hereditary thrombosis associated with antithrombin resistance, with a substitution of arginine for leucine at position 596 (p.Arg596Leu) in the gene encoding prothrombin (called prothrombin Yukuhashi). The mutant prothrombin had moderately lower activity than wild-type prothrombin in clotting assays, but the formation of thrombin-antithrombin complex was substantially impaired. A thrombin-generation assay revealed that the peak activity of the mutant prothrombin was fairly low, but its inactivation was extremely slow in reconstituted plasma. The Leu596 substitution caused a gain-of-function mutation in the prothrombin gene, resulting in resistance to antithrombin and susceptibility to thrombosis.

The prevalence of neutralizing antibodies against adeno-associated virus capsids is reduced in young Japanese individuals.

Pre-existing antibodies against adeno-associated virus (AAV), caused by natural AAV infections, interfere with recombinant AAV vector-mediated gene transfer. We studied the prevalence of neutralizing antibodies against AAV serotypes 1, 2, 5, 8, and 9 in healthy subjects (n = 85) and hemophilia patients (n = 59) in a Japanese population. For healthy subjects, the prevalence of neutralizing antibodies against AAV serotypes 1, 2, 5, 8, and 9 was 36.5%, 35.3%, 37.6%, 32.9%, and 36.5%, respectively, while that in hemophilia patients was 39.7%, 28.8%, 35.6%, 32.9%, and 27.4%, respectively. There was no difference in the prevalence of neutralizing antibody against each AAV serotype between the healthy subjects and the hemophilia patients. The prevalence of neutralizing antibodies against all AAV serotypes increased with age in both healthy subjects and hemophilia patients. High titers of neutralizing antibodies against AAV2 (≥1:224) and AAV8 (≥1:224) were more evident in older individuals (≥42 years old). Approximately 50% of all screened individuals were seronegative for neutralizing antibodies against each AAV tested, while approximately 25% of individuals were seropositive for each AAV serotype tested. The prevalence of seronegativity for all AAV serotypes was 67.0% (healthy subjects, 68.6%; hemophilia patients, 65.0%) and 18.6% (healthy subjects, 20.5%; hemophilia patients, 15.7%) in young (<42 years old) and older subjects (≥42 years old), respectively. The findings from this study suggested that young subjects are more likely to be eligible for gene therapy based on AAV vectors delivered via an intravascular route because of the low prevalence of antibodies to AAV capsids.

Burden of congenital hemophilia A requiring treatment in Japan: The HIKOBOSHI study.

Background: Treatment of congenital hemophilia A (HA) in Japan has greatly improved with the widespread adoption of prophylactic factor (F)VIII concentrates. However, it is unknown if this has translated into a real-world reduction in disease and treatment burden. Objectives: To describe HA disease burden in Japan based on information from two medical information databases, JMDC and Real World Data Co., Ltd. (RWD). Methods: Eligible individuals were diagnosed with congenital HA and prescribed FVIII concentrates, bypassing agents, or emicizumab. Treatment patterns and disease burden data were derived from health insurance claims and electronic medical records. Results: Data on 459 people with HA were retrospectively collected from 2005 to 2020 in the JMDC database (median [min, max] of 37 [2, 186] months of available records), and 229 people with HA from 1985 to 2020 in the RWD database (median [min, max] of 154 [0, 409] months of available records). Mean (standard deviation) ages at the time of the first record were 25.0 (16.8) years (JMDC) and 19.2 (20.3) years (RWD). In the JMDC database, mean monthly FVIII dose increased from 2201 IU in 2005 to 8239 IU in 2013 to 11,377 IU in 2019; HA-related drug costs increased accordingly. Mean (95% confidence interval) annual outpatient and out-of-hours visits decreased slightly between 2013 and 2019 (outpatient visits: from 22.9 [16.8-29.0] to 14.3 [12.6-16.1] per person; out-of-hours visits: from 1.3 [0.2-2.5] to 0.6 [0-1.4]). There was no change in mean number of hospitalizations. Conclusions: Challenges remain in HA, including treatment burden, outpatient visits, and hospitalizations.

The seroprevalence of neutralizing antibodies against the adeno-associated virus capsids in Japanese hemophiliacs.

Adeno-associated virus (AAV) vectors are promising modalities of gene therapy to address unmet medical needs. However, anti-AAV neutralizing antibodies (NAbs) hamper the vector-mediated therapeutic effect. Therefore, NAb prevalence in the target population is vital in designing clinical trials with AAV vectors. Hence, updating the seroprevalence of anti-AAV NAbs, herein we analyzed sera from 100 healthy individuals and 216 hemophiliacs in Japan. In both groups, the overall seroprevalence against various AAV serotypes was 20%-30%, and the ratio of the NAb-positive population increased with age. The seroprevalence did not differ between healthy participants and hemophiliacs and was not biased by the concomitant blood-borne viral infections. The high neutralizing activity, which strongly inhibits the transduction with all serotypes in vitro, was mostly found in people in their 60s or of older age. The multivariate analysis suggested that "60s or older age" was the only independent factor related to the high titer of NAbs. Conversely, a large proportion of younger hemophiliacs was seronegative, rendering them eligible for AAV-mediated gene therapy in Japan. Compared with our previous study, the peak of seroprevalences has shifted to older populations, indicating that natural AAV exposure in the elderly occurred in their youth but not during the last decade.

Convenience Comparison Study of Reconstitution Devices for the Blood Coagulation Factor VIII Products Rurioctocog Alfa Pegol and Antihemophilic Factor (Recombinant).

INTRODUCTION: The Baxject® II Hi-Flow device has previously been used to reconstitute the factor VIII products antihemophilic factor (recombinant) (ADVATE®) and rurioctocog alfa pegol (ADYNOVATE®). METHODS: In this crossover study in healthy men, the convenience of an advanced device, Baxject III® with and without a nonslip sleeve, was compared with that of Baxject II Hi-Flow. The primary endpoint was the operational time for reconstitution; secondary endpoints included participants' assessment of the usability of the devices for reconstitution and their preference for using each of the devices. RESULTS: Twelve healthy adult men (mean ± standard deviation [SD] age: 36.7 ± 7.0 years) and 12 healthy elderly men (mean ± SD age: 70.3 ± 4.8 years) participated in the study. In the adult group, the mean operational time for reconstitution was shorter using Baxject III (mean ± SD: 19.7 ± 2.7 and 19.9 ± 5.2 seconds with and without a nonslip sleeve, respectively) than when using Baxject II Hi-Flow (49.6 ± 7.2 seconds, P < 0.0001 for both comparisons). Adult participants rated preference (P < 0.0001) and ease of reconstitution (P < 0.0001) as higher for Baxject III with a nonslip sleeve than for Baxject II Hi-Flow. Results were consistent regardless of age group or the use of the nonslip sleeve. CONCLUSION: Owing to the convenience of Baxject III, this device will improve the reconstitution process for patients with hemophilia treated with rurioctocog alfa pegol or antihemophilic factor (recombinant) at home.

Effect of transfusion on the venous blood lactate level in very low-birthweight infants.

BACKGROUND: In recent years the blood lactate level can be easily and quickly measured with a small amount of blood, and the availability of an arterial blood lactate level has been reported as an indicator of oxygen deficit in adults. To determine whether venous blood lactate level can serve as such a marker for determining the indications for transfusion, blood lactate and hemoglobin level were monitored before and after transfusion. METHODS: The study subjects consisted of 12 very low-birthweight infants admitted to the neonatal intensive care unit and who had transfusion between June 2005 and June 2007. The data on the blood lactate and hemoglobin were collected retrospectively by referring to the clinical records. RESULTS: A total of 18 transfusions was performed. There was no significant relationship between venous blood lactate and hemoglobin concentration before transfusion. The subjects were classified into two groups according to the lactate level before transfusion: > or =3.3 mmol/L and <3.3 mmol/L. In the high-lactate group the lactate decreased significantly after transfusion (P < 0.01) and it continued to decrease thereafter. In the low-lactate group, however, the lactate remained unchanged. CONCLUSIONS: Venous blood lactate measurements may offer some additional information regarding the optimal time for performing a transfusion. To the authors' knowledge this is the first report to study the changes in lactate levels using venous blood sampling in red blood cell transfusion in very low-birthweight infants.

Correction to: Efficacy and safety of full-length pegylated recombinant factor VIII with extended half-life in previously treated patients with hemophilia A: comparison of data between the general and Japanese study populations.

The authors would like to correct the error in Table 2 in the original publication of the article. The "Blood type" is not described in any part of "Results" and "Discussion" and had no impact on the conclusion hence the bottom of the table is removed.

Efficacy and safety of full-length pegylated recombinant factor VIII with extended half-life in previously treated patients with hemophilia A: comparison of data between the general and Japanese study populations.

Rurioctocog alfa pegol (BAX 855) is a novel third-generation recombinant factor VIII whose active ingredient is chemically modified with polyethylene glycol. A global multicenter phase 2/3 study of the product in 137 patients (including 11 patients from Japan) with severe hemophilia A aged 12-65 years, reported an extended half-life and a good tolerability profile, as well as a significantly lower annualized bleeding rate in the prophylactic treatment arm than in the on-demand treatment arm. Using descriptive statistics, a post hoc analysis was performed to compare the pharmacokinetics, safety, and efficacy profiles of the product in the Japanese subpopulation and the overall population. Extended half-life was demonstrated in the Japanese subpopulation. The mean [standard deviation (SD)] annualized bleeding rates in the prophylactic treatment arm were 3.7 (4.7) for the overall population (n = 120) and 4.0 (3.4) for the Japanese subpopulation (n = 11). The proportion of bleeds reported as excellent or good was 94.9% (149/157) in the overall population, whereas that in the Japanese subpopulation was 92.3% (12/13). No FVIII inhibition or anaphylactic reaction was reported in the Japanese subpopulation. The post hoc comparisons demonstrated similar pharmacokinetic, safety, and efficacy profiles between the overall population and the Japanese subpopulation.

HLA-B polymorphism in Japanese HIV-1-infected long-term surviving hemophiliacs.

Approximately 30% of patients with hemophilia in Japan were infected with human immunodeficiency virus (HIV) in early 1980s through contaminated blood products. In 1995, a cohort of HIV-infected, asymptomatic patients with hemophilia was set up for follow-up study. Although the patients met the criteria for long-term non-progressor (LTNP) at the entry to the cohort, some of them later developed lymphopenia during five more years of observation. We collected blood samples from 80 long-term survivors; 42 of them did not require antiviral therapy, but the rest were under treatment. Analysis of HLA-B genotype revealed that carriers of known HIV-resistant alleles such as HLA-B*5701, B*5801, and alleles of B27 antigenic group were not increased in frequency, but that HLA-B*1507 was increased in the cohort (6.25% vs. 1.03%, OR = 6.40, p = 0.039). We also observed the decrease in carriers of HLA-B*5401 (3.75% vs. 14.95%, OR = 0.22, p = 0.016). HLAB* 5401 is a relatively common allele in East Asian populations and belongs to the same B22 antigenic group as B55 and B56 which were reported to associate with rapid progression. Our data indicated that HLA class I is one of the host factors involved in the retardation of HIV disease progression as also reported in the previous studies; however, the alleles associated with this resistance were not the same because of divergent host genetic background.

The International Immune Tolerance Induction Study and its follow-up study on Japanese hemophilia A patients with inhibitors.

The International Immune Tolerance Induction (I-ITI) Study in hemophilia A patients with inhibitors included 16 Japanese patients among a total of 115 test subjects. The results within this group of Japanese patients were 11 cases of I-ITI off-study, three cases of I-ITI on-study, and two cases of tolerance on prophylaxis. There was no significant difference in success rate between the low-dose and high-dose groups (Study I). Successively, independent follow-up survey in Japan was conducted in 14 cases, with consent (Study II). Ten cases were off-study at the end of the I-ITI Study. Of these 10 cases, seven of seven successful cases remained clinical successes at the end of the follow-up study, one partial success became a full success while a second relapsed, and one failure was subsequently evaluated as a partial success. Four cases that were on-study at the end of I-ITI Study were classified as three successes and one failure at the end of the follow-up study. As a result, the status at the end of follow-up study was: 11 ITI successes (78.6 %); one partial success; one failure; and one relapse. Thus, the ITI follow-up study was helpful in providing a long-term prognostic determination of inhibitors.

[Acquired von Willebrand syndrome with autoimmune hemolytic anemia].

A 17-year-old female underwent examination due to the chief complaints of epistaxis and bleeding from the gums. In addition to hemolytic anemia (Coombs positivity), she was found to have a marked reduction in the von Willebrand factor antigen (vWF:Ag) and ristocetin cofactor activity (vWF:RCo). Anti vWF antibodies mainly comprised of IgG1 fractions were also detected, so acquired von Willebrand syndrome (AvWS) with autoimmune hemolytic anemia (AIHA) was diagnosed. The recovery of vWF:Ag and vWF:RCo following administration of a factor VIII concentrate containing vWF was good, but the half life was short at about two hours and the collagen binding activity and vWF:RCo in mixing tests with control plasma were only mildly inhibited. Therefore, the anti vWF antibody in the present case was thought not to inhibit vWF function but to be involved in clearance promotion. Hemostatic control during hemorrhage was performed using a factor VIII concentrate containing vWF. With the administration of prednisolone thereafter, anti vWF antibodies disappeared at the same time as there was improvement in the AIHA. Throughout the course the patient contracted acute hepatitis A, became pregnant and gave birth, but there were no clear effects on the clinical phenotype of AvWS.

[Antiphospholipid antibodies in children with idiopathic thrombocytopenic purpura].

Three antiphospholipid antibodies (aPLs), namely, antiphosphatidylinositol antibody (antiinositol antibody), antiphosphatidylserine antibody (antiserine antibody), and anticardiolipin. beta 2-glycoprotein I complex antibody (antiCL. beta 2-GPI antibody), were determined in 49 children with idiopathic thrombocytopenic purpura (ITP) consisting of 14 newly-diagnosed cases and 35 chronic cases. Determination of aPL was performed twice in the newly-diagnosed patients, once each during the acute and convalescent phases, and once in the chronic patients. The positive rates in the acute and convalescent phases of the newly-diagnosed group and in the chronic group were, respectively, 14.3%, 28.6%, and 18.8% for the antiinositol antibody, 14.3%, 14.3%, and 15.6% for the antiserine antibody, and 21.4%, 28.6%, and 25.0% for either of these 2 antibodies. Thus, antiinositol and antiserine aPLs were present at high incidences; however, all patients were negative for the antiCL. beta 2-GPI antibody. No correlation was noted between either the antiinositol or the antiserine antibody and peripheral platelet count, anti-GP IIb/IIIa antibody or PAIgG. Thus, although some aPLs are present in both acute and chronic pediatric ITP, the aPLs seems to be of an infectious disease type. No results that suggest possible involvement of aPLs in ITP pathology were obtained.

Long-term follow-up of children with refractory immune thrombocytopenia treated with rituximab.

Data on long-term outcomes of children with refractory immune thrombocytopenia (ITP) treated with rituximab are limited. We retrospectively analyzed the long-term effect of rituximab on 22 pediatric ITP patients (11 boys and 11 girls). Compete response (CR) (platelet count ≥100 × 10(9)/L) and partial response (PR) (platelet count 30-99 × 10(9)/L) were achieved in nine (41 %) and two (9 %) patients, respectively. Of the 11 responders, eight subsequently relapsed 2-26 months after initial rituximab treatment. The 5-year relapse-free rate was 14 % (3/22, 95 % confidence interval: 0-27 %) with a median follow-up period of 6.4 years. Five initial responders with subsequent relapse and one non-responder received multiple rituximab treatments of nine courses; all patients responded to the second rituximab therapy without any significant toxicity. All eight patients who relapsed after an initial response and six of 11 non-responders achieved CR or PR with subsequent treatment, including repeated courses of rituximab, splenectomy, steroids, and other immunomodulating agents. Our findings indicated that the sustained effect of rituximab on children with refractory ITP is low, but that the long-term outcome of ITP itself is not poor. Furthermore, repeated rituximab administration may be a promising therapy for those who relapse after an initial response.