Successful Bone Marrow Plus Cord Blood Stem Cell Transplantation in a Girl who Developed Myelodysplastic Syndrome from Hepatitis-associated Aplastic Anemia Treated with Long-term Immunosuppressants and Growth Factors

A 9-year-old girl who had hepatitis-associated aplastic anemia was treated intermittently with methylprednisolone pulse therapy and growth factors (granulocyte-colony stimulating factor (G-CSF), recombinant human erythropoietin (rhEpo) and cyclosporin A (CyA) for over two years. At this time, there was hematological improvement, but chromosome analysis revealed monosomy 7.

After six months, there was progression to myelodysplastic syndrome (MDS) (stage in refractory anemia of excess blasts (RAEB)) with monosomy 7, monosomy 6, marker chromosome and with hematological deterioration.

She received bone marrow (1.57×10⁵ cells kg^?1 (patient body weight)) plus cord blood cell (0.3×10⁷ cells kg^?1 (patient body weight)) transplantation from her brother, 2 years and 7 months after the diagnosis of hepatitis-associated aplastic anemia. Engraftment was achieved after two weeks, and acute graft-versus-host disease occurred in a mild form after four weeks. Hematological remission has been continuous for 20 months after bone marrow transplantation. Transformation of hepatitis-associated aplastic anemia to MDS with the monosomy 7, monosomy 6 and marker chromosome in this patient was considered to have been related to the administration of high doses of immunosuppressive drugs plus growth factors.     

Measurements of immature platelets with haematology analysers are of limited value to separate immune thrombocytopenia from bone marrow failure

Detection of immature platelets in the circulation may help to dissect thrombocytopenia due to platelet destruction from bone marrow failure (BMF ). We prospectively tested the predictive value of immature platelets, measured as immature platelet fraction (IPF ) on the XE‐5000 (Sysmex, Kobe, Japan) or percentage of reticulated platelets (rPT ) on the CD Sapphire (Abbott Diagnostics, Santa Clara, CA, USA) to separate immune thrombocytopenia (ITP ) from BMF (leukaemia, myelodysplastic syndrome, aplastic anaemia). We analysed 58 samples of patients with BMF , 47 samples of patients with ITP and 97 controls. Median rPT (CD Sapphire) was increased to 9·0% in ITP and to 10·9% in BMF , compared to 1·9% in controls. Median IPF (XE‐5000) was 16·2% in ITP , 10·2% in BMF and 2·5% in controls. We found an inverse correlation between high fractions of immature platelets and low platelet counts in thrombocytopenic samples regardless of the diagnosis. In conclusion, we observed a broad overlap of immature platelets between ITP and BMF , which may be caused by an accelerated release of immature platelets in any thrombocytopenic state and decreased production in many patients with ITP . Despite this, IPF (XE‐5000) had some power to discriminate ITP from BMF , whereas rPT (CD Sapphire) was of no predictive value.     

Assessment of an immature platelet fraction (IPF) in peripheral thrombocytopenia

A new automated method to reliably quantify reticulated platelets, expressed as the immature platelet fraction (IPF), has been developed utilizing the XE-2100 blood cell counter with upgraded software (Sysmex, Kobe, Japan). The IPF is identified by flow cytometry techniques and the use of a nucleic acid specific dye in the reticulocyte/optical platelet channel. The clinical utility of this parameter was established in the laboratory diagnosis of thrombocytopenia due to increased peripheral platelet destruction, particularly autoimmune thrombocytopenic purpura (AITP) and thrombotic thrombocytopenic purpura (TTP). Reproducibility and stability results over 48 h were good. An IPF reference range in healthy individuals was established as 1.1-6.1%, with a mean of 3.4%. Patients in whom platelet destruction might be abnormal, were studied and two of these patients followed serially during the course of treatment. The IPF was raised in several disease states. The most significant increases in IPF values were found in patients with AITP (mean 22.3%, range 9.2-33.1%) and acute TTP (mean 17.2%, range 11.2-30.9%). Following patients during treatment demonstrated that as the platelet count recovered the IPF% fell. These results show that a rapid, inexpensive automated method for measuring the IPF% is feasible and should become a standard parameter in evaluating the thrombocytopenic patient.     

Determinants of platelet count in pediatric patients with congenital cyanotic heart disease: Role of immature platelet fraction

Objectives

Congenital heart defects are common noninfectious causes of mortality in children. Bleeding and thrombosis are both limiting factors in the management of such patients. We assessed the frequency of thrombocytopenia in pediatric patients with congenital cyanotic heart disease (CCHD) and evaluated determinants of platelet count including immature platelet fraction (IPF) and their role in the pathogenesis of thrombocytopenia.

Methods

Forty‐six children and adolescents with CCHD during pre‐catheter visits were studied; median age was 20.5 months. Complete blood count including IPF as a marker of platelet production and reticulated hemoglobin content (RET‐He) as a marker of red cell production and iron status were done on Sysmex XE 2100 (Sysmex, Japan). C‐reactive protein, prothrombin time (PT), Activated partial thromboplastin time (APTT) were also assessed.

Results

Thrombocytopenia was found in 6 patients (13%). PT was prolonged (P = .016) and IPF was significantly higher in patients with thrombocytopenia compared with patients with normal platelet count (14.15 ± 5.2% vs 6.68 ± 3.39%; P = .003). Platelet count was negatively correlated with IPF while significant positive correlations were found between IPF and hemoglobin, red blood cells (RBCs) count, hematocrit (Hct), PT, reticulocytes count, and immature reticulocyte fraction.

Conclusions

We suggest that elevated IPF in CCHD patients with thrombocytopenia may denote peripheral platelets destruction as an underlying mechanism. Hemoglobin level, RBCs count, Hct, and RET‐He were not significant determinants for platelet count in CCHD.     

Reliability of automated platelet counts: Comparison with manual method and utility for prediction of clinical bleeding

The 20 × 10⁹/L (20,000/μl) threshold for prophylactic platelet transfusion may be unnecessarily high. The widespread use of this threshold may reflect lack of confidence in the reliability of low platelet counts. We evaluated the performance of automated platelet counts and their relation to clinical bleeding. First, we prepared serial blood dilutions with “target” platelet counts from 2 to 40 × 10⁹/L For the 48 measurements on 2 × 10⁹/L “target” dilutions, values of 1 or 2 × 10⁹/L were obtained with the Sysmex NE-8000 analyzer (mean 1.44 × 10⁹/L; SD 0.31 × 10⁹/L). Similarly, for 5 × 10⁹/L “target” counts, automated counts were 3-6 × 10⁹/L (mean 4.42 × 10⁹/L; SD 0.18 × 10⁹/L). Similar results were observed with all other “target” levels, with coefficients of variation (CV) from 6.39% to 7.71% with 10-40 × 10⁹/L “target” values. Secondly, we compared triplicate automated and manual platelet counts on thrombocytopenic patients with platelet counts from 4–30 × 10⁹/L. The triplicate automated platelet counts differed by no more than 5 × 10⁹/L among themselves, whereas the manual counts varied by as much as 30 × 10⁹/L. Mean platelet counts: automated, 14.40 × 10⁹/L (CV 10.12%); manual, 16.48 × 10⁹/L (CV 30.39%) (P = 0.038 for counts; P < 0.001 for CV). Finally, we prospectively evaluated bleeding in thrombocytopenic patients (1,809 patient-days of observation). Univariate and multivariate logistic regression analysis revealed highly significant correlations between the automated platelet count and major and minor bleeding manifestations. Thus, automated platelet counts are highly reliable and accurately predict clinical bleeding. The use of automated analyzers should facilitate improved prophylactic platelet transfusion protocols.     

New quantitative parameters on a recently introduced automated blood cell counter – the XE 2100TM

The XE 2100 (Sysmex Corporation) is a cell counter that furthers the technology of fluorescent flow cytometry developed from the earlier range of Sysmex analysers. The new diagnostic features are a nucleated red cell count (NRBC), the ability to measure platelets by impedance as well as an ‘optical’ platelet count using a fluorescence dye and an immature granulocyte (IG) count. The NRBC count was highly correlated (r=0.97) with the manual reference count. For counts below 100 × 10⁹/l the ‘optical’ method and the immunocount gave good a correlation (r=0.97) optical and impedance counts were also well correlated (r=0.89). The use of the ‘optical’ platelet count significantly improves the reliability of low platelet counts. The IG count correlated with visual counts (r=0.81) and allows the detection of immature cells at an earlier stage in the laboratory process. The introduction of fluorescent flow cytometric analysis allows extended quantification of additional cell populations and so potentially improves screening and monitoring of various pathological conditions.     

Romiplostim monotherapy in thrombocytopenic patients with myelodysplastic syndromes: long‐term safety and efficacy

Romiplostim can improve platelet counts in about 50% of patients with low‐ or intermediate 1‐risk (lower risk) myelodysplastic syndromes (MDS) and thrombocytopenia, but its long‐term toxicity and efficacy are not known. This open‐label extension study evaluated the long‐term safety and efficacy of romiplostim in 60 patients with lower risk MDS and platelet counts ≤50 × 10⁹/l. The primary endpoint was adverse event (AE) incidence. Secondary endpoints were efficacy parameters, including bleeding events and platelet response. Median (range) treatment time in the extension study and the median observation times thereafter were 25 (2–181) and 57 (11–209) weeks, respectively. Treatment‐related AEs and serious AEs were reported in 14/60 (23%) and 4/60 (7%) patients, respectively. Progression to acute myeloid leukaemia (AML) occurred in two patients after 44 and 46 weeks. Patients (n = 34, 57%) with a platelet response were further evaluated for length of response. Median (range) response duration was 33 (7–174) weeks; 28/34 (82%) patients had a continuous response. Five of 34 patients (15%) had grade ≥3 bleeding events; three when the platelet count was >50 × 10⁹/l. There were no new safety concerns and the rate of progression to AML was low; response to romiplostim was maintained for most patients.     

Platelet production and platelet destruction: assessing mechanisms of treatment effect in immune thrombocytopenia

This study investigated the immature platelet fraction (IPF) in assessing treatment effects in immune thrombocytopenia (ITP). IPF was measured on the Sysmex XE2100 autoanalyzer. The mean absolute-IPF (A-IPF) was lower for ITP patients than for healthy controls (3.2 vs 7.8 × 10?/L, P < .01), whereas IPF percentage was greater (29.2% vs 3.2%, P < .01). All 5 patients with a platelet response to Eltrombopag, a thrombopoietic agent, but none responding to an anti-FcγRIII antibody, had corresponding A-IPF responses. Seven of 7 patients responding to RhoD immuneglobulin (anti-D) and 6 of 8 responding to intravenous immunoglobulin (IVIG) did not have corresponding increases in A-IPF, but 2 with IVIG and 1 with IVIG anti-D did. This supports inhibition of platelet destruction as the primary mechanism of intravenous anti-D and IVIG, although IVIG may also enhance thrombopoiesis. Plasma glycocalicin, released during platelet destruction, normalized as glycocalicin index, was higher in ITP patients than controls (31.36 vs 1.75, P = .001). There was an inverse correlation between glycocalicin index and A-IPF in ITP patients (r2 = -0.578, P = .015), demonstrating the relationship between platelet production and destruction. Nonresponders to thrombopoietic agents had increased megakaryocytes but not increased A-IPF, suggesting that antibodies blocked platelet release. In conclusion, A-IPF measures real-time thrombopoiesis, providing insight into mechanisms of treatment effect.     

Immature platelet fraction measurement in patients with chronic liver disease: a convenient marker for evaluating cirrhotic change

Platelet number is often used as an indicator of the severity of liver disease. Although inadequate thrombopoietin production and decreased platelet production have been proposed as major causes of cirrhotic thrombocytopenia, the underlying mechanism has not yet been fully clarified. We examined whether the measurement of the immature platelet fraction (IPF) in thrombocytopenic patients with liver dysfunction is useful as a rapid and noninvasive method for the differential diagnosis of chronic liver diseases. We examined 20 liver cirrhosis patients, 56 patients with chronic hepatitis, 9 patients with fatty liver, and 86 patients without liver disease. The percentage value of IPF (IPF%) was measured using an XE-2100 multiparameter automatic hematology analyzer. Using a receiver operating characteristic curve, we found diagnostic significance of the absolute platelet count and the absolute number of the IPF between cirrhotic patients and noncirrhotic patients, and developed a powerful multivariate discriminant analysis (MDA) function based on the platelet count and the IPF%. The diagnostic accuracy obtained by the MDA function was superior to that obtained by the absolute number of platelets and the IPF. We therefore propose our IPF% measurement for the diagnosis of liver cirrhosis.     

Correlation between immature platelet fraction and reticulated platelets. Usefulness in the etiology diagnosis of thrombocytopenia

Background: Reticulated platelets (RP) are a surrogate marker for megakaryocytic activity, but the limitation of this determination is the lack of standardization of methodology. The determination of the immature platelet fraction (IPF) is performed in a simple, automated, and reproducible way between laboratories. We analyzed the correlation between IPF and RP, and usefulness of IPF in patients with thrombocytopenia. Methods: RP were determined by flow cytometry using double staining with thiazole orange and CD61 PerCP^®. IPF was performed with Sysmex XE2100 analyzer. We used a control group with normal platelets, and thrombocytopenic patients were classified into three groups: Group 1. Central thrombocytopenia, Group 2. Thrombocytopenia as a result of enhanced peripheral platelet destruction, and Group 3. Peripheral non‐immune thrombocytopenia by abnormal distribution. Results: Fourteen controls and 66 patients were analyzed. Group 1: 25 patients, they had mean and confidence interval 95% (95% CI) for IPF 8.67% (6.49–10.46%) and RP 4.08% (2.86–5.30%). Group 2: 20 patients, they had mean and 95%CI for IPF 16.80% (12.20–21.39%) and RP 16.14% (9.89–22.40%). Group 3: 21 patients, they had mean and 95% CI for IPF 9.04% (6.95–11.14%) and RP 5.23% (3.41–7.05%). The overall Pearson linear correlation between IPF and RP was r: 0.65. There were statistically significant differences in values of IPF and RP between Group 2 and the other two groups (P < 0.01). Conclusion: There is a good correlation between IPF and RP mainly in thrombocytopenia by peripheral destruction. Determination of IPF is an easy technique in their implementation, standardized and reproducible, so it could be a useful screening technique in patients with thrombocytopenia.     

Myelodysplastic syndrome in transformation to acute myeloid leukemia presenting with diabetes insipidus: due to pituitary infiltration association with abnormalities of chromosomes 3 and 7

A 31-yr-old woman with myelodysplastic syndrome (MDS) in transformation to acute myeloid leukemia (AML) presented with initial symptoms of polyuria and polydipsia. Cytogenetics revealed monosomy 7 and translocation (3;3)(q21;q26). The initial symptoms, in conjunction with a low serum level of anti-diuretic hormone (ADH) and magnetic resonance imaging (MRI) findings demonstrating loss of the "bright spot" of the neurohypophysis, indicated diabetes insipidus (DI), e.g. caused by leukemic infiltration of the neurohypophysis. After induction chemotherapy the patient's bone marrow revealed blast persistence, and following a second course of chemotherapy and normalisation of MRI, an allogeneic peripheral blood stem cell transplantation (PBSCT) from the patient's HLA-identical brother was performed, resulting in ongoing complete remission. Recently, Lavabre-Bertrand et al. reported an association of AML with DI, elevated platelet counts, and monosomy 7 and chromosome 3 abnormalities in three patients (Eur. J. Haematol. 2001: 66: 66-69). Our report of an MDS with trilineage dysplasia and these karyotypic changes associated with DI indicates that this new entity may also include preleukemic cases.     

Immature platelet fraction related parameters in the differential diagnosis of thrombocytopenia

Abstract

The pathogenesis of thrombocytopenia can be divided into increased destruction (ID) of platelets in the peripheral blood and decreased production (DP) of platelets in the bone marrow. This study aimed to analyze the efficacy of immature platelet fraction (IPF) related parameters, including the IPF count (IPF#), IPF percentage (IPF%) and highly fluorescence IPF percentage (H-IPF%), measured by XN-9000, in the differential diagnosis of thrombocytopenia. One hundred and twenty healthy volunteers were enrolled in the healthy control (HC) group, and 180 thrombocytopenia patients were grouped into either the increased destruction (ID) group or the decreased production (DP) group according to their final diagnosis. IPF# was significantly lower in the DP group than in the ID and HC groups (P < .01). Among the three groups, the ID group had the highest IPF% and H-IPF%, and the HC group had the lowest IPF% and H-IPF%. The differences between the three groups were all statistically significant (P < .01). In differentiating the ID patients from the DP patients, the areas under the operating characteristics curve of IPF#, IPF% and H-IPF% were 0.859, 0.944 and 0.930, respectively. False positive rates were below 0.04 when IPF#, IPF% and H-IPF% were above 2.65, 7.55 and 2.35, respectively. IPF related parameters showed high efficacy in the differential diagnosis of thrombocytopenia. However, due to the small numerical values of the IPF related parameters in some thrombocytopenia patients, the fluctuations of IPF% and H-IPF% should also be taken into consideration. Though H-IPF% is a new parameter, its effectiveness in the differential diagnosis of thrombocytopenia is not better than IPF%’s.     

Beyond the platelet count: immature platelet fraction and thromboelastometry correlate with bleeding in patients with immune thrombocytopenia

Platelet counts (PC) estimate bleeding risk in Immune Thrombocytopenia (ITP). We investigated whether measures of thromboelastometry and absolute immature platelet fraction (A‐IPF) would correlate better with acute bleeding score (ABS) than PC or mean platelet volume (MPV). Simultaneous determination of ABS, complete blood count and thromboelastometry was performed in 141 ITP patients; 112 underwent A‐IPF testing. Subgroup analyses were performed for paediatric subjects, PC <60 × 10⁹/l and <30 × 10⁹/l. PC significantly inversely correlated with ABS in all subjects, PC <30 × 10⁹/l and total paediatric cohort. MPV did not correlate with ABS in any subgroup. Thromboelastometry measures of clot firmness, but not PC, significantly correlated with ABS in all subjects with PC <60 × 10⁹/l, and children with PC <60 × 10⁹/l and <30 × 10⁹/l. A‐IPF demonstrated stronger correlation with ABS than did PC among all subjects, those with PC <60 × 10⁹/l, all children and children with PC <30 × 10⁹/l (r = ?0·37; r = ?0·34; r = ?0·44; r = ?0·60) versus ABS with PC (r = ?0·36; ns; r = ?0·32; ns). Stronger correlations of both thromboelastometry measures of clot firmness and A‐IPF than PC with ABS suggest factors beyond PC, i.e. related to platelet function, contribute to ITP bleeding pathophysiology. Thromboelastometry, A‐IPF and ABS can be incorporated into routine or acute visits.     

Stromal cell-derived factor-1 rs2297630 polymorphism associated with platelet production and treatment response in Chinese patients with chronic immune thrombocytopenia

Stromal cell-derived factor-1 (SDF-1), signaling through CXCR4, is implicated in megakaryopoiesis and platelet production. SDF-1 rs2297630 is a functional polymorphism in linkage disequilibrium with other functional variants in SDF-1. This study aimed to investigate the role of SDF-1 rs2297630 in chronic ITP. The genotypes were determined by polymerase chain reaction-restriction fragment length polymorphism and confirmed by direct sequencing. Immature platelet fraction (IPF) was performed using Sysmex XE-2100. Anti-platelet autoantibodies were assayed by enzyme-linked immunosorbent assay. The main characteristics at diagnosis and the outcome of chronic ITP in 201 Chinese patients were retrospectively reviewed. There was no significant difference in either genotype or allelic distribution between ITP patients and the controls (p = 0.114; p = 0.787). However, both heterozygote (GA) and homozygote minor allele (AA) patients had significantly increased megakaryocyte quantity compared to homozygote genotype (GG) patients at diagnosis (p = 0.011). The mean IPF values of GA and AA genotype patients were higher than those observed in the GG genotype patients when platelet counts ≤50 × 10⁹/L at diagnosis (p = 0.007). Patients with GA and AA genotype showed a higher response rate to standard treatments than patients with GG genotype (p < 0.001). In particular, GA and AA genotype patients had a significantly increased chance of responding to steroids, intravenous immunoglobulin (IVIG), and thrombopoietin analogs (p = 0.007; p = 0.029; p = 0.034, respectively). No significant difference was found between anti-platelet antibodies and genotypes (p = 0.296). In summary, the SDF-1 rs2297630 was associated with platelet production and treatment response in Chinese patients with chronic ITP.     

Transient atypical monocytosis mimic acute myelomonocytic leukemia in post‐chemotherapy patients receiving G‐CSF: report of two cases

Granulocyte colony‐stimulating factor (G‐CSF) is now widely used in patients with malignant disorders receiving intensive chemotherapy to increase leukocyte count and to upregulate phagocyte function during neutropenia. Monocytosis associated with G‐CSF has been reported in anecdotal literature. We report two cases of pseudoleukemia secondary to G‐CSF administration. Both patients initially presented with myelodysplastic syndrome with chromosome 7 abnormalities that evolved into acute myeloid leukemia. Case one had deletion 7q while case two initially had monosomy 7 and subsequently developed a balanced translocation between the short (p) arm of chromosome 1 and long (q) arm of chromosome 15. Following the induction chemotherapy and G‐CSF administration, both of these patients developed pseudoleukemia. Patient 1 had white blood cell (WBC) count of 26 × 10⁹/l with 72% monocytes, while patient two had WBC of 14.1 × 10⁹/l with 30% monocytes. In both patients the monocytosis resolved after the discontinuation of G‐CSF therapy. In summary, patients treated with G‐CSF should be followed closely. In those cases with pseudoleukemia discontinuation of the drug with no supplemental chemotherapy is probably enough to control the atypical monocytosis.     

Sole abnormalities of chromosome 7 in myeloid malignancies: spectrum, histopathologic correlates, and prognostic implications

Among 6,565 consecutive abnormal cytogenetic reports at our institution, 3,192 (49%) constituted sole abnormalities, of which 230 (7%) involved chromosome 7: monosomy 7 (n = 98), 7q- (n = 51), der(1;7)(q10;p10) (n = 44), balanced translocations (n = 15), ring 7 (n = 13), and 7p- (n = 9). The most frequent histopathologic correlates were myelodysplastic syndromes (MDS; 28%), acute myeloid leukemia (AML; 17%), secondary or therapy-related MDS/AML (13%), primary myelofibrosis (PMF; 7%), and chronic myelomonocytic leukemia (6%). Monosomy 7 was the most frequent in each one of these disease categories except PMF where 7q- was more frequent. In primary MDS, patients with der(1;7)(q10;p10) (n = 13), compared to those with monosomy 7 (n = 30) or 7q- (n = 15), were less likely (P = 0.04) to display excess blasts or multilineage dysplasia but overall and leukemia-free survival adjusted for these variables revealed no significant difference between the three groups (P = 0.57 and 0.81, respectively). The current study does not prognostically distinguish monosomy 7 from 7q- or der(1;7), in MDS.     

Significance of immature platelet fraction and CD41-positive cells at birth in early onset neonatal thrombocytopenia

Early thrombocytopenia is a common hematological abnormality in sick neonates. Here, we examined the relationship between early thrombocytopenia in neonates and parameters associated with thrombopoiesis to identify predictive factors at birth. Two hundred and forty-four neonates admitted to the neonatal intensive care unit were divided into thrombocytopenic (n = 55, 23%) and non-thrombocytopenic (n = 189, 77%) groups based on platelet counts, which were monitored within 72 h of birth. Immature platelet fraction (IPF) and platelet count at birth were determined simultaneously soon after phlebotomy with an automated hematology analyzer. Megakaryocytes and their precursors positive for CD41 in peripheral blood were examined at birth by flow cytometry. The thrombocytopenic group showed significantly higher IPF percentage and lower percentage of CD41⁺ mononuclear cells (MNCs) than did the non-thrombocytopenic group (P < 0.01). Moreover, the percentage of CD41⁺ MNCs significantly differentiated neonates with platelet counts >150 × 10³/μL at birth and nadir platelet count <150 × 10³/μL over the clinical course from neonates without thrombocytopenia. These observations suggest that the percentage of CD41⁺ MNCs at birth and IPF percentage are useful predictors of early thrombocytopenia in the majority of sick neonates.     

Multicentre,randomised phase III study of the efficacy and safety of eltrombopag in Chinese patients with chronic immune thrombocytopenia

Eltrombopag, a thrombopoietin receptor agonist, raises platelet counts and reduces bleeding in patients with immune thrombocytopenia (ITP ). In Chinese patients, eltrombopag was evaluated at an initial dose of 25 mg, vs. 50 mg for non‐Asians, because the plasma exposure of eltrombopag is higher in East Asians. A multicentre, double‐blind, randomised, placebo‐controlled, 8‐week, phase III study enrolled 155 patients with chronic, previously treated ITP . Dosage could be adjusted (25–75 mg/day) to maintain platelet counts 50–250 × 10⁹/l. The primary efficacy endpoint was the proportion of patients with a platelet count ≥50 × 10⁹/l after Day 42. Pharmacokinetics and pharmacodynamics of eltrombopag were analysed in an open‐label extension. After Day 42, 57·7% of eltrombopag‐treated and 6·0% of placebo‐treated patients achieved platelet counts ≥50 × 10⁹/l. Odds of achieving a platelet count ≥50 × 10⁹/l were 26·08 times greater with eltrombopag than placebo (P  <  0·001). Compared with placebo, time to response and duration of response were better with eltrombopag (P  <  0·001) and the odds of any bleeding were reduced by 72% (P  =  0·001). Tolerability, pharmacokinetics, and pharmacokinetics/pharmacodynamics were similar to previous findings in East Asian patients. In conclusion, in Chinese patients with chronic ITP , eltrombopag 25 mg once daily, elevated platelet counts to a safe range and reduced bleeding.     

Thrombocytopenia model with minimal manipulation of blood cells allowing whole blood assessment of platelet function

In vitro models of thrombocytopenia are useful research tools. Previously published models have shortcomings altering properties of platelets and other blood components. The aim of the present study was to develop a whole blood method to induce thrombocytopenia with minimal manipulation, and to describe platelet function in induced thrombocytopenia in individuals with healthy platelets. Hirudin anticoagulated blood was obtained from 20 healthy volunteers. One part of the blood was gently centrifuged at 130g for 15 minutes. The platelet-rich plasma was replaced with phosphate-buffered saline to establish thrombocytopenia. Various levels of thrombocytopenia were achieved by combining different volumes of baseline whole blood and thrombocytopenic blood. Platelet counts were measured by flow cytometry (Navios, Beckman Coulter) and routine haematological analyser (Sysmex XE-5000). Platelet function was analysed by impedance aggregometry (Multiplate® Analyzer, Roche) and by flow cytometry (Navios, Beckman Coulter) using collagen, adenosine diphosphate, thrombin receptor activating peptide-6 and ristocetin as agonists. Median baseline platelet count was 227×10⁹/l. The in vitro model yielded median platelet counts at 51×10⁹/l (range 26–93×10⁹/l). We observed minor, yet significant, changes in platelet size and maturity from baseline to modelled thrombocytopenia. In the thrombocytopenic samples, significant and positive linear associations were found between platelet count and platelet aggregation across all agonists (all p-values<0.001). Platelet function assessed by flow cytometry showed minimal alterations in the thrombocytopenic samples. A new whole blood-based model of thrombocytopenia was established and validated. This new model serves as a useful future tool, particularly to explore platelet function in patients with thrombocytopenia.     

Donor Leukocyte Infusion for Japanese Patients with Relapsed Leukemia After Allogeneic Bone Marrow Transplantation: Indications and Dose Escalation

Abstract: To clarify the role of dose escalation of donor leukocyte infusion (DLI) in the treatment of relapsed leukemia after allogeneic bone marrow transplant (BMT), data from 100 patients were collected from 46 facilities in Japan and analyzed with respect to indications and infused cell dose. Complete remission (CR) was achieved in 11 of 12 (91%) patients with relapsed chronic myelogenous leukemia (CML) in the chronic phase, 3 of 11 (27%) with CML in the acute phase, 8 of 21 (38%) with acute myelogenous leukemia (AML), 6 of 23 (25%) with acute lymphoblastic leukemia (ALL), and 5 of 11 (45%) with myelodysplastic syndrome (MDS). The probability of remaining in CR at 3 years was 82% in CML patients in the chronic phase, but 0% in those with CML in the acute phase, 7% in those with AML, 0% with ALL, and 33% with MDS. Acute graft‐versus‐host disease (GVHD) (≥2) developed in 31 of 89 (34%) patients with human leukocyte antigen identical related donors and was fatal for 7 (7%). A leukocyte dose of 1 × 10⁷/kg of recipient body weight with CML in the chronic phase, 3 × 10⁷/kg of recipient body weight with MDS, and 1 × 10⁸/kg of recipient body weight with acute leukemia appeared to be optimal as an initial dose of DLI. However, the minimal dose of leukocyte developing fatal GVHD was 7 × 10⁷/kg of recipient body weight. These suggest that a relatively small dose of DLI ranging from 1 × 10⁷/kg to 5 × 10⁷/kg of recipient body weight should be administered initially then the infused escalating dose 2 or 3 months later in patients with CML in the chronic phase and MDS. However, a large number of leukocytes around 1 × 10⁸/kg are needed to induce graft versus leukemia effects in patients with acute leukemia despite a 7% fatality in GVHD.