Stimulate Platelet Production Research Articles

Structural basis of MPL activation by thrombopoietin

AbstractMyeloproliferative leukemia protein (MPL), also known as thrombopoietin (TPO) receptor, is a class I cytokine receptor that is expressed on hematopoietic progenitors, promoting growth and differentiation toward the megakaryocyte lineage and is critical for normal platelet production. Mutations in MPL, TPO, or Janus kinase 2 (JAK2) have been implicated in multiple diseases from congenital thrombocytopenias to myeloproliferative neoplasms. The ligand for MPL, TPO, stimulates platelet production by inducing MPL dimerization and results in an active conformation that allows downstream JAK2/STAT5 signaling. Despite the biological importance of this pathway, the molecular signaling mechanism remained unclear. Here, we present a 3.39-Å cryo-electron microscopy structure of the ectodomain of mouse MPL bound to TPO. The structure revealed both low and high affinity sites between MPL and TPO that contain several pathologic mutations. To better understand TPO-driven MPL signaling, we expanded upon this structure by molecular dynamic (MD) simulations to model the full-length human MPL/TPO complex, and showed that MPL D4-D4 domain interactions are functionally relevant in activity assays. To build on our understanding of downstream activation, we added JAK2 to the MPL/TPO complex by MD simulations. This ternary complex illustrates JAK2 dimerization through the pseudokinase domain, illustrates residues important for MPL interactions, and reveals the constitutive activation mechanism of patient mutant V617F. The model also suggests the mechanism of JAK2 tyrosine kinase domain transphosphorylation. Overall, our studies illuminate TPO/MPL/JAK2 signaling mechanisms and provide additional insight into the nature of receptor signaling, which will further benefit human health.

Structural Basis for Activation of the Thrombopoietin Receptor (MPL) By Its Ligand (TPO)

Myeloproliferative leukemia protein (MPL), also known as thrombopoietin receptor, is a class I cytokine receptor expressed on hematopoietic progenitors that is critical for normal platelet production by promoting myeloid progenitor growth and differentiation toward the megakaryocyte lineage. The ligand for MPL, thrombopoietin (TPO), stimulates platelet production through binding MPL via two sequential sites, a high- and then a low-affinity interface that together induce MPL dimerization and result in an active conformation allowing downstream JAK2/STAT5 signaling. Despite intense interest in characterizing the molecular mechanism of the interaction between MPL and TPO over the last several decades, no experimental structural evidence has been reported. Here we present a 3.39 Å cryo-EM structure of the ectodomain of mouse MPL bound to TPO. TPO features both low and high affinity sites that each bind to one MPL monomer in agreement with previous functional studies. Our structure reveals that several patient mutations associated with congenital amegakaryocytic thrombocytopenia (CAMT) and aplastic anemia are found at MPL-TPO interaction sites. The structure further explains additional patient mutations, as well as published experimental mutations that have previously been the basis of structure-function studies. Building on this structure, we generated a molecular dynamics simulation model of the full length human MPL-TPO complex, revealing that TPO binding to full length MPL results in unexpected MPL-MPL intramolecular interactions that stabilize the dimer to allow for efficient signaling. Functional characterization of deleterious mutations predicted by our structure and modelling have revealed additional residues critical to MPL-TPO and MPL-MPL interactions, providing further validation of our proposed structure. Overall, our studies reveal new insight into a receptor-cytokine interaction critical for normal hematopoiesis and platelet production that are frequently mutated in disease, provide the structural basis underlying existing experimental findings, and provide additional insight into the nature of hematopoietic growth factor signaling which could benefit future therapeutic research.

Thrombocytosis and Megakaryocyte Changes Associated with Pure Red Cell Aplasia

J.A., R.G.W, and G.R. contributed equally to this abstract. Introduction: Acquired pure red cell aplasia (PRCA) is a rare hematological disorder characterized by normocytic, normochromic anemia with severe reticulocytopenia and a notable absence of identifiable erythroid precursors in the bone marrow. PRCA is often without a clear etiology and can be associated with conditions such as Large Granular Lymphocytic (LGL) Leukemias, B-cell dyscrasias, thymomas, immunodeficiencies or infections. Discovery of an IgG inhibitor in primary autoimmune pure red cell aplasia (PRCA) has been an immensely significant breakthrough. This finding has not only shed light on the critical role of IgG inhibitors in the pathophysiology of PRCA but has also underscored the importance of aberrant T cell activity in this condition. Thrombocytosis refers to an elevation in platelet count of >450,000/μL, which is not a typical feature associated with PRCA. Based on observations in our clinical practice of patients with PRCA with concomitant thrombocytosis, we screened our cohort for the combination of these features to discover potential mechanisms underlying this phenomenon. Methods: In order to address this objective, we conducted a retrospective analysis of medical records from 90 patients diagnosed with acquired PRCA at the University of Texas Southwestern and Cleveland Clinic Foundation between 2000 and 2022. Our primary goal was to determine the frequency of thrombocytosis among these patients. For those in the subset displaying thrombocytosis, longitudinal hemoglobin and platelet counts before and after treatment of the condition were also collected for relevant comparisons utilizing paired t-tests after assessing for normality with Shapiro-Wilk test. Results: Among a cohort of 90 patients diagnosed with acquired PRCA, 12 individuals exhibited thrombocytosis at the time of PRCA diagnosis (Table 1). Demographically, this subset of individuals had a median age of 53.5, and a 3:1 ratio of women to men. 66.7% of those with thrombocytosis also exhibited megakaryocyte hyperplasia on bone marrow biopsy. Molecular analysis ruled out the presence of myeloproliferative neoplasms, and blood work was obtained ruling out iron deficiency. Patients were treated for PRCA with a variety of medications including prednisone, cyclosporine, tacrolimus, rituximab, and cyclophosphamide. After treatment, 8 of the 12 patients demonstrated complete resolution of their PRCA, defined as transfusion independence with hemoglobin levels above 10. Further examination of those 8 patients with resolution of PRCA revealed statistically significant differences in mean hemoglobin and platelet counts before and after treatment (Figure 1). Specifically, the mean hemoglobin count increased from 7.9 (SD 2.5) g/dL to11.6 (1.8) g/dL (p=0.0168), while the platelet count decreased from 557,300 (SD 104,700) /μL to 414,400 (SD 109,600) /μL (p=0.0049). Conclusion: Our report is the most comprehensive case series to date reporting on thrombocytosis in patients with acquired PRCA. We speculate that the arrest of erythropoiesis generating PRCA may lead to skewing of precursors towards the megakaryocytic lineage. Alternatively, EPO production secondary to PRCA could stimulate TPO receptors and thus stimulate platelet production. Clinicians should be aware of this particular PRCA presentation. Further investigations will aid in elucidating whether one of these mechanisms may also be playing a role in PRCA-related thrombocytosis. The significance of thrombocytosis or megakaryocyte alterations in PRCA remains enigmatic, necessitating further investigations to elucidate their role in disease pathophysiology, as well as their utility as biomarkers for treatment response prediction. The limitations of this study encompass a small sample size and the absence of post-treatment bone marrow biopsy results, which would have facilitated the visual assessment of any resolved megakaryocyte abnormalities. Future directions include examining megakaryocyte hyperplasia and other megakaryocyte abnormalities such as hyper- and hypo-lobation associated with PRCA.

Hetrombopag plus recombinant human thrombopoietin for chemotherapy-induced thrombocytopenia in patients with solid tumors

BackgroundChemotherapy-induced thrombocytopenia (CIT) is a common hematological complication in patients with cancer. Hetrombopag is a novel thrombopoietin receptor agonist that has shown an additive effect in stimulating platelet production when combined with recombinant human thrombopoietin (rhTPO). ObjectivesThis multicenter retrospective cohort study aimed to evaluate the efficacy and safety of hetrombopag plus rhTPO compared with rhTPO alone for CIT. MethodsA total of 294 patients with solid tumors and CIT (platelet count, <50 × 109/L) who received either rhTPO plus hetrombopag (146 patients) or rhTPO alone (148 patients) at 3 centers from January to December 2022 were included in the study. The primary outcome was a platelet count at least 50 × 109/L higher than the baseline value within 14 days. Chemotherapy dose reductions/delays, bleeding, and adverse events were reported. ResultsOne hundred twenty patients (82.2%) in the rhTPO-hetrombopag group vs 100 patients (67.6%) in the rhTPO group achieved the primary outcome (P = .005). This significant difference persisted in adjusted analysis (odds ratio, 2.01; 95% CI, 1.12-3.60). A total of 115 patients (78.8%) in the rhTPO-hetrombopag group and 101 patients (68.2%) in the rhTPO group avoided chemotherapy dose reductions/delays (P = .041). There was no significant difference in bleeding rates, and adverse events were mild and similar between the 2 groups. No deaths occurred. ConclusionCompared to rhTPO alone, our findings suggest that the combination of hetrombopag and rhTPO is safe and more effective in patients with CIT.

Immunomodulation with romiplostim as a second-line strategy in primary immune thrombocytopenia: The iROM study.

Thrombopoietin receptor agonists (TPO-RAs) stimulate platelet production, which might restore immunological tolerance in primary immune thrombocytopenia (ITP). The iROM study investigated romiplostim's immunomodulatory effects. Thirteen patients (median age, 31 years) who previously received first-line treatment received romiplostim for 22 weeks, followed by monitoring until week 52. In addition to immunological data, secondary end-points included the sustained remission off-treatment (SROT) rate at 1 year, romiplostim dose, platelet count and bleedings. Scheduled discontinuation of romiplostim and SROT were achieved in six patients with newly diagnosed ITP, whereas the remaining seven patients relapsed. Romiplostim dose titration was lower and platelet count response was stronger in patients with SROT than in relapsed patients. In all patients, regulatory T lymphocyte (Treg) counts increased until study completion and the counts were higher in patients with SROT. Interleukin (IL)-4, IL-9 and IL-17F levels decreased significantly in all patients. FOXP3 (Treg), GATA3 (Th2) mRNA expression and transforming growth factor-β levels increased in patients with SROT. Treatment with romiplostim modulates the immune system and possibly influences ITP prognosis. A rapid increase in platelet counts is likely important for inducing immune tolerance. Better outcomes might be achieved at an early stage of autoimmunity, but clinical studies are needed for confirmation.

PEGylated thrombopoietin mimetic, JNJ‑26366821 a novel prophylactic radiation countermeasure for acute radiation injury

Thrombopoietin (TPO) is the primary regulator of platelet generation and a stimulator of multilineage hematopoietic recovery following exposure to total body irradiation (TBI). JNJ‑26366821, a novel PEGylated TPO mimetic peptide, stimulates platelet production without developing neutralizing antibodies or causing any adverse effects. Administration of a single dose of JNJ‑26366821 demonstrated its efficacy as a prophylactic countermeasure in various mouse strains (males CD2F1, C3H/HeN, and male and female C57BL/6J) exposed to Co-60 gamma TBI. A dose dependent survival efficacy of JNJ‑26366821 (− 24 h) was identified in male CD2F1 mice exposed to a supralethal dose of radiation. A single dose of JNJ‑26366821 administered 24, 12, or 2 h pre-radiation resulted in 100% survival from a lethal dose of TBI with a dose reduction factor of 1.36. There was significantly accelerated recovery from radiation-induced peripheral blood neutropenia and thrombocytopenia in animals pre-treated with JNJ‑26366821. The drug also increased bone marrow cellularity and megakaryocytes, accelerated multi-lineage hematopoietic recovery, and alleviated radiation-induced soluble markers of bone marrow aplasia and endothelial damage. These results indicate that JNJ‑26366821 is a promising prophylactic radiation countermeasure for hematopoietic acute radiation syndrome with a broad window for medical management in a radiological or nuclear event.

Roles and mechanism of IL-11 in vascular diseases.

Vascular diseases are the leading cause of morbidity and mortality worldwide. Therefore, effective treatment strategies that can reduce the risk of vascular diseases are urgently needed. The relationship between Interleukin-11 (IL-11) and development of vascular diseases has gained increasing attention. IL-11, a target for therapeutic research, was initially thought to participate in stimulating platelet production. Additional research concluded that IL-11 is effective in treating several vascular diseases. However, the function and mechanism of IL-11 in these diseases remain unknown. This review summarizes IL-11 expression, function, and signal transduction mechanism. This study also focuses on the role of IL-11 in coronary artery disease, hypertension, pulmonary hypertension, cerebrovascular disease, aortic disease, and other vascular diseases and its potential as a therapeutic target. Consequently, this study provides new insight into the clinical diagnosis and treatment of vascular diseases.

Rilzabrutinib versus placebo in adults and adolescents with persistent or chronic immune thrombocytopenia: LUNA 3 phase III study.

Immune thrombocytopenia (ITP) is characterized by primarily autoantibody-mediated platelet destruction and impaired platelet production resulting in thrombocytopenia and an increased risk of bleeding. Other manifestations include increased risk of thrombosis and diminished quality of life. Current treatment approaches are directed toward lowering the rate of platelet destruction or stimulating platelet production to prevent bleeding. Rilzabrutinib is an oral, reversible, potent Bruton tyrosine kinase inhibitor that was specifically designed to treat immune-mediated diseases and mediates its therapeutic effect through a dual mechanism of action: (1) inhibiting B-cell activation and (2) interrupting antibody-coated cell phagocytosis by Fc gamma receptor in spleen and liver. A 24-week dose-finding phase I/II study of rilzabrutinib in patients with ITP showed a 40% platelet response (⩾2 consecutive platelet counts of ⩾50 × 109/L and increase from baseline ⩾20 × 109/L without rescue medication use) and a well-tolerated safety profile with only grade 1/2 transient adverse events across dose levels. Assess the efficacy and safety of oral rilzabrutinib in adult and adolescent patients with persistent or chronic ITP. Rilzabrutinib 400 mg BID is being evaluated in the ongoing LUNA 3 multicenter, double-blind, placebo-controlled phase III study. The primary endpoint is durable platelet response, defined as achieving platelet counts of ⩾50 × 109/L for at least two-thirds of ⩾8 available weekly scheduled platelet measurements during the last 12 weeks (including ⩾2 available measurements within the last 6 weeks) of the 24-week blinded treatment period in the absence of rescue therapy. Ethical guidelines and informed consent are followed. The LUNA 3 trial will further investigate rilzabrutinib's safety and efficacy in adult and adolescent patients, with the primary goal of addressing a major objective in treating patients with ITP: durability of platelet response. ClinicalTrials.gov NCT04562766: https://clinicaltrials.gov/ct2/show/NCT04562766; EU Clinical Trials Register EudraCT 2020-002063-60: https://www.clinicaltrialsregister.eu/ctr-search/search?query=2020-002063-60.

Novel chemical-structure TPOR agonist, TMEA, promotes megakaryocytes differentiation and thrombopoiesis via mTOR and ERK signalings

BackgroundNon-peptide thrombopoietin receptor (TPOR) agonists are promising therapies for the mitigation and treatment of thrombocytopenia. However, only few agents are available as safe and effective for stimulating platelet production for thrombocytopenic patients in the clinic. PurposeThis study aimed to develop a novel small molecule TPOR agonist and investigate its underlying regulation of function in megakaryocytes (MKs) differentiation and thrombopoiesis. MethodsA potential active compound that promotes MKs differentiation and thrombopoiesis was obtained by machine learning (ML). Meanwhile, the effect was verified in zebrafish model, HEL and Meg-01 cells. Next, the key regulatory target was identified by Drug Affinity Responsive Target Stabilization Assay (DARTS), Cellular Thermal Shift Assay (CETSA), and molecular simulation experiments. After that, RNA-sequencing (RNA-seq) was used to further confirm the associated pathways and evaluate the gene expression induced during MK differentiation. In vivo, irradiation (IR) mice, C57BL/6N-TPORem1cyagen (Tpor-/-) mice were constructed by CRISPR/Cas9 technology to examine the therapeutic effect of TMEA on thrombocytopenia. ResultsA natural chemical-structure small molecule TMEA was predicted to be a potential active compound based on ML. Obvious phenotypes of MKs differentiation were observed by TMEA induction in zebrafish model and TMEA could increase co-expression of CD41/CD42b, DNA content, and promote polyploidization and maturation of MKs in HEL and Meg-01 cells. Mechanically, TMEA could bind with TPOR protein and further regulate the PI3K/AKT/mTOR/P70S6K and MEK/ERK signal pathways. In vivo, TMEA evidently promoted platelet regeneration in mice with radiation-induced thrombocytopenia but had no effect on Tpor-/- and C57BL/6 (WT) mice. ConclusionTMEA could serve as a novel TPOR agonist to promote MKs differentiation and thrombopoiesis via mTOR and ERK signaling and could potentially be created as a promising new drug to treat thrombocytopenia.

Use of Thrombopoietin Agonists in Immune Thrombocytopenia Purpura

Introduction: Immune/idiopathic thrombocytopenia purpura (ITP) is a condition characterized by low platelet counts. Current chronic treatment options for ITP include rituximab, a thrombopoietin (TPO) agonist, or a splenectomy. Rituximab, a CD20 monoclonal antibody that depletes B-cells to disrupt the production of harmful antibodies (Zaja et al. 2003), has historically been the primary treatment for ITP. However, early studies have shown that while effective in the short term, rituximab has a drop in efficacy after 180 days (Zaja et al. 2003). TPO agonists are a newer mechanism that mimic endogenous TPO by stimulating the maturation and proliferation of megakaryocytes to stimulate platelet production (Nugent et al. 2009). Current TPO agonists include avatrombopag, eltrombopag, fostamatinib, and romiplostim. We explored the use of these TPO agonists practically in the hematology/oncology practice at Pennsylvania Hospital (PAH) to evaluate efficacy and responses to each therapy. Methods: A retrospective chart review was conducted through the University of Pennsylvania Epic system investigating chronic treatment of ITP at PAH hematology/oncology department since 2012 with a diagnosis of ITP. Results: 231 charts were analyzed. 121 patients did not require chronic treatment (1 patient was admitted for workup while the chart review occurred, and treatment is still being determined). 10 had chronic treatment via steroids, mainly prednisone, and 1 only had a family history of ITP with no ITP diagnosis. 8 charts were unable to be reviewed due to privacy concerns. This left 91 charts which were reviewed for chronic management of ITP. Of these 91 charts, 7 only presented for a second opinion on management and did not receive treatment. 15 underwent a splenectomy, but only 3 did not first receive a secondary therapy. 48 received rituximab, but only 15 (31.25%) had a sustained response. 66 patients eventually received a TPO agonist. Of these 66 patients, only 22 (33.33%) received a single therapy. The other 44 received multiple TPO agonists or rituximab and then a TPO agonist. All four TPO agonists were used, and the responses to these drugs are summarized in table 1. Conclusions: Of the 231 patients presenting to the PAH hematology/oncology clinic, 91 required long term, non-steroidal therapy. Our findings support previous studies that show a decline in long term efficacy of rituximab with only 31.25% (15) of patients achieving a sustained response. The other 68.65% (33) required an additional therapy. Interestingly, all 3 secondary therapies (rituximab, TPO agonist, and splenectomy) were utilized by the clinicians and 72.5% (66) of patients required a TPO agonist. About one-third (22) of these 66 patients had a sustained response on a single TPO agonist on their first try. However, two-thirds (44) of the patients had to switch to another TPO agonist before finding an acceptable response. Eltrombopag and romiplostim are relatively older than fostamatinib and avatrombopag, however the newer drugs are beginning to make their way into therapy with 5 patients on ongoing therapy with fostamatinib (3) and avatrombopag (2). The downfall of the TPO agonists is that most require sustained long-term therapy with only 5 patients total achieving a sustained response without therapy. This is especially difficult for romiplostim which requires patients to come in weekly for an injection. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

Administration of recombinant human thrombopoietin is associated with alleviated thrombocytopenia in adult intensive care unit patients with pneumonia: A single-center retrospective study.

Background: Recombinant human thrombopoietin (rhTPO) is reported to stimulate platelet production and increase peripheral platelet counts; it is primarily used to manage chemotherapy-induced thrombocytopenia and idiopathic thrombocytopenic purpura. However, the effect of rhTPO in patients with pneumonia and thrombocytopenia remains uncertain. Objective: To assess the association of rhTPO and platelet counts in ICU patients with pneumonia and thrombocytopenia. Materials and Methods: A retrospective cohort study was performed in the ICU department, Nanfang Hospital, Southern Medical University, Guangzhou, China. From January 2016 to April 2021, patients with pneumonia and thrombocytopenia were allocated to two groups—the rhTPO and no-rhTPO groups—according to whether they received rhTPO treatment or not during their ICU stay. Demographical and clinical data were collected and analyzed using statistical software; p < 0.05 was considered statistically significant. Results: Out of 327 patients, 149 were in the rhTPO group and 178 were in the no-rhTPO group. Within the first 7 days, platelet counts increased more for patients in the rhTPO group compared with those in the no-rhTPO group (99.21 ± 102.613 vs. 2.08 ± 43.877, p = 0.000). The clinical recovery rate of platelets increased within 7 days (65.8 vs. 18.5%, p = 0.000) and, after 7 days of enrollment, hemorrhagic scores decreased more apparently in the rhTPO group (2.81 ± 2.856 vs. 1.16 ± 2.123, p = 0.000). Further, bleeding events ceased in 66.7% of the patients in the rhTPO group compared with 37.3% of the patients in the no-rhTPO group (p = 0.000). Less red-blood-cells transfusions were needed in the rhTPO group (3.639 ± 4.630 vs. 5.818 ± 6.858, p = 0.009). Furthermore, through logistic regression, rhTPO administration was found to be an independent indicator that affected the platelet recovery rate within 7 days. Conclusion: This study finds that rhTPO administration is associated with increased platelet counts, alleviated bleeding, and reduced blood transfusion. For patients with pneumonia and thrombocytopenia, rhTPO may be an effective therapeutic drug; however, more RCT trails are needed to confirm our observation.

Safety, tolerance, pharmacokinetic and pharmacodynamic properties of thrombopoietin mimetic peptide for injection in Chinese healthy volunteers: a randomized, placebo-controlled, double-blind study

The thrombopoietin mimetic peptide for injection is a second-generation thrombopoietin receptor agonist (TPO-RA) used in the treatment of patients with immune thrombocytopenia. The aim of the present study was to assess the safety, tolerance, pharmacokinetic and pharmacodynamic properties of thrombopoietin mimetic peptide for injection in Chinese healthy volunteers. A randomized, placebo-controlled, double-blind, dose-escalation study was conducted in healthy Chinese subjects aged 18–50 years. Thirty subjects received single subcutaneous injection of 0.3 μg/kg, 1.0 μg/kg, 2.0 μg/kg thrombopoietin mimetic peptide or placebo. Thrombopoietin mimetic peptide was safe and well tolerated at doses of 0.3–2.0 μg/kg. There was no significant change in mean platelet count (PLT) from baseline at the 0.3 μg/kg or placebo groups. The mean PLT of subjects in the 1.0 μg/kg and 2.0 μg/kg groups peaked at day 12 (± 1), began to decline around day 17, and returned to the baseline level at day 28 (± 1). Platelet aggregation rates of the three dose groups showed no significant change before and after administration. Serum concentrations of thrombopoietin mimetic peptide in all subjects were below the quantization limit. This was the first study to demonstrate that subcutaneous injection of thrombopoietin mimetic peptide at doses of 0.3–2.0 μg/kg was safe and well tolerated in Chinese healthy subjects. As a second-generation TPO-RA, thrombopoietin mimetic peptide is effective at improving PLT after single subcutaneous injection at dose of ≥1 μg/kg. P lain l anguage s ummary What is the context? ● Immune thrombocytopenia (ITP) is a rare, serious autoimmune disorder characterized by low platelet count (PLT) without an alternate cause. The treatment goal of ITP is to increase the platelet count to a safe level that can stop active bleeding and reduce the risks of future bleeding. ● Thrombopoietin receptor agonists (TPO-RAs, e.g. eltrombopag, avatrombopag, hetrombopag, and romiplostim) have shown high response rates in stimulating platelet production and reducing the risk of bleeding. TPO-RAs provide ITP patients with well-tolerated, long-term treatment choices. What is new? ● The thrombopoietin mimetic peptide for injection is a new TPO-RAs developed by Shandong Quangang Pharmaceutical Co., Ltd. (China). ● This study showed that thrombopoietin mimetic peptide is effective at improving PLT after a single subcutaneous injection. ● The thrombopoietin mimetic peptide is safe and well-tolerated in Chinese healthy subjects. What is the impact? ● This study provides evidence for the further development potential of the thrombopoietin mimetic peptide.

Thrombocytopenia and Therapeutic Strategies after Allogeneic Hematopoietic Stem Cell Transplantation.

Thrombocytopenia following allogeneic hematopoietic stem cell transplantation is a usual complication and can lead to high morbidity and mortality. New strategies, such as the use of another graft versus host-disease prophylaxis, alternative donors, and management of infections, have improved the survival of these patients. The mechanisms are unknown; therefore, the identification of new strategies to manage this potentially serious problem is needed. Thrombopoietin receptor agonists are currently available to stimulate platelet production. Some small retrospective studies have reported their potential efficacy in an allogeneic stem cell transplant setting, confirming good tolerability. Recent studies with higher numbers of patients also support their safety and efficacy in this setting, hence establishing the use of these drugs as a promising strategy for this post-transplant complication. However, prospective trials are needed to confirm these results.

Megakaryocyte Diversity in Ontogeny, Functions and Cell-Cell Interactions.

Hematopoietic stem cells (HSCs) rely on local interactions in the bone marrow (BM) microenvironment with stromal cells and other hematopoietic cells that facilitate their survival and proliferation, and also regulate their functions. HSCs and multipotent progenitor cells differentiate into lineage-specific progenitors that generate all blood and immune cells. Megakaryocytes (Mks) are hematopoietic cells responsible for producing blood platelets, which are essential for normal hemostasis and blood coagulation. Although the most prominent function of Mks is platelet production (thrombopoiesis), other increasingly recognized functions include HSC maintenance and host immune response. However, whether and how these diverse programs are executed by different Mk subpopulations remains poorly understood. This Perspective summarizes our current understanding of diversity in ontogeny, functions and cell-cell interactions. Cumulative evidence suggests that BM microenvironment dysfunction, partly caused by mutated Mks, can induce or alter the progression of a variety of hematologic malignancies, including myeloproliferative neoplasms (MPNs) and other disorders associated with tissue scarring (fibrosis). Therefore, as an example of the heterogeneous functions of Mks in malignant hematopoiesis, we will discuss the role of Mks in the onset and progression of BM fibrosis. In this regard, abnormal interactions between of Mks and other immune cells might directly contribute to fibrotic diseases. Overall, further understanding of megakaryopoiesis and how Mks interact with HSCs and immune cells has potential clinical implications for stem cell transplantation and other therapies for hematologic malignancies, as well as for treatments to stimulate platelet production and prevent thrombocytopenia.

Eltrombopag in paediatric immune thrombocytopenia: Iron metabolism modulation in mesenchymal stromal cells.

SummaryImmune thrombocytopenia (ITP) is an autoimmune disease caused by platelet destruction mediated by auto‐antibody production. It is characterized by a compromised immune system and alteration of the inflammatory response. Mesenchymal stromal cells (MSCs) play an important role in modulating immune and inflammatory processes, exerting immune‐suppressing and anti‐inflammatory properties. In ITP‐MSCs the activity and survival are strongly impaired. Eltrombopag (ELT) is a thrombopoietin receptor agonist approved in chronic ITP for stimulating platelet production. It has immunomodulating properties by stimulating T and B regulatory cell activity and by promoting a macrophage switch from the pro‐inflammatory to the anti‐inflammatory phenotype. ELT also exhibits iron‐chelating properties. Iron is a crucial element involved in several physiologic processes, but its intracellular accumulation determines cell damages. Therefore, for the first time we analysed the effect of ELT on ITP‐MSCs demonstrating its ability to restore survival and activity of MSCs directly and to promote their survival and proliferation indirectly, by iron metabolism modulation.

Avatrombopag Use in Patient With Thromboembolic Risks Listed for Combined Coronary Artery Bypass Grafting and Liver Transplant: A Case Report

Thrombocytopenia commonly occurs in patients with advanced liver disease and can be a contraindication in patients needing combined coronary artery bypass grafting (CABG) and liver transplant (LT). Thrombopoietin receptor agonists, including avatrombopag, are part of a novel drug class and stimulate platelet production. Avatrombopag is indicated in the perioperative setting to avoid platelet transfusions, which carry several disadvantages. Avatrombopag was shown to be safe and effective in patients with chronic liver disease. This study describes the successful use of avatrombopag in a patient with thromboembolic risks in preparation for a combined CABG and LT. Larger clinical trials are necessary to validate our results.

Treatment of severe thrombocytopenia with platelet biotherapic and Phosphorus

Background Thrombocytopenia is a decrease in the number of platelets in the blood resulting from disorders of production, distribution or destruction, leading to bleeding. The respective treatment of thrombocytopenia is usually based on corticotherapy and platelet transfusion. Aims Report the evolution of homeopathic treatment in a 12 years old Maltese canine cardiopathic patient with thrombocytopenia associated with the diagnosis of Ehrlichia canis. Methods Replacement of corticoid and transfusion by Platelet Biotherapic, to stimulate platelet production and high diluted Phosphorus to contain the bleeding. Results On 01/28/19, the patient presented thrombocytopenia of 5.000/mm³ with large platelets, petechials and suffusion hemorrhages in dorsal and ventral regions, extensive acute cervical hematoma due to blood collection; anisocytosis, polychromasia and Howell Jolly corpuscle in red series; and also reactive lymphocytes, monocytes active and toxic neutrophils in white series. On 01/30/19, initiated the administration of 3 globules of Platelet Biotherapic 12CH every 12 hours; 3 globules of Phosphorus 6CH every 24 hours, both oral; and topical phytotherapeutic use of Arnica montana once daily. Three days later, it was observed the complete absence of petechials and suffusion hemorrhages, with elevation of platelets count to 74.000/mm³ and mild leukocytosis with improvement in overall patient well-being. On 02/06/19, there was regression of hematoma and leukocytosis, in addition to the platelet count in 95.000/mm³. On 02/14/19, due to slight decrease in platelets to 92.000/mm³, the frequency of Phosphorus 6CH was changed every 12 hours. On 02/20/19, the new platelet count reached 126.000/mm³ with standardized morphology and no Howell Jolly record. Conclusion The homeopathies (High dilution medicine) have proven to be successful in the treatment of thrombocytopenia without causing side effects.

Emerging Therapies in Immune Thrombocytopenia.

Immune thrombocytopenia (ITP) is a rare autoimmune disorder caused by peripheral platelet destruction and inappropriate bone marrow production. The management of ITP is based on the utilization of steroids, intravenous immunoglobulins, rituximab, thrombopoietin receptor agonists (TPO-RAs), immunosuppressants and splenectomy. Recent advances in the understanding of its pathogenesis have opened new fields of therapeutic interventions. The phagocytosis of platelets by splenic macrophages could be inhibited by spleen tyrosine kinase (Syk) or Bruton tyrosine kinase (BTK) inhibitors. The clearance of antiplatelet antibodies could be accelerated by blocking the neonatal Fc receptor (FcRn), while new strategies targeting B cells and/or plasma cells could improve the reduction of pathogenic autoantibodies. The inhibition of the classical complement pathway that participates in platelet destruction also represents a new target. Platelet desialylation has emerged as a new mechanism of platelet destruction in ITP, and the inhibition of neuraminidase could dampen this phenomenon. T cells that support the autoimmune B cell response also represent an interesting target. Beyond the inhibition of the autoimmune response, new TPO-RAs that stimulate platelet production have been developed. The upcoming challenges will be the determination of predictive factors of response to treatments at a patient scale to optimize their management.

Effects of Eltrombopag on In Vitro Macrophage Polarization in Pediatric Immune Thrombocytopenia.

Immune Thrombocytopenia (ITP) is an autoimmune disease characterized by autoantibodies-mediated platelet destruction, a prevalence of M1 pro-inflammatory macrophage phenotype and an elevated T helper 1 and T helper 2 lymphocytes (Th1/Th2) ratio, resulting in impairment of inflammatory profile and immune response. Macrophages are immune cells, present as pro-inflammatory classically activated macrophages (M1) or as anti-inflammatory alternatively activated macrophages (M2). They have a key role in ITP, acting both as effector cells, phagocytizing platelets, and, as antigen presenting cells, stimulating auto-antibodies against platelets production. Eltrombopag (ELT) is a thrombopoietin receptor agonist licensed for chronic ITP to stimulate platelet production. Moreover, it improves T and B regulatory cells functions, suppresses T-cells activity, and inhibits monocytes activation. We analyzed the effect of ELT on macrophage phenotype polarization, proposing a new possible mechanism of action. We suggest it as a mediator of macrophage phenotype switch from the M1 pro-inflammatory type to the M2 anti-inflammatory one in paediatric patients with ITP, in order to reduce inflammatory state and restore the immune system function. Our results provide new insights into the therapy and the management of ITP, suggesting ELT also as immune-modulating drug.

The Impact of Ethnicity on the Response to Eltrombopag in Patients with Immune Thrombocytopenia (ITP) in Qatar: A Single Institution Experience

Background: Thrombopoietin receptor agonists stimulate platelet production . Eltrombopag olamine (ELT) is alow molecular weight, synthetic nonpeptide agent thathas been studied in multiple phase 3 trials and proved efficacy at a standard dose of 50 mg. ELT exposure has been reported to be different in different ethnic descents. In East Asian ITP patients, the area under the curve (AUC) was reported to be more than 85% those of non-East Asian descent Objectives: The objective of this study is to evaluate the efficacy of ELT in Arab and Asian ITP patients of the subcontinent of India by using a lower starting dose (12.5 mg) and maximum (50 mg) doses of ELT than the standard starting dosing of (50 mg) and maximum of (75 mg) approved in the USA and Europe. Methods A retrospective study was conducted to evaluate the role of ethnicity (Arab and East Asians) in response to ELT among ITP patients by reviewing patients' electronic medical records between Jan 2015 - Jan 2019. A total of 58 patients were identified. We examined retrospectively Arab (n = 41) and non-Arab Asian (n = 17) patients who are 18 years and older in Qatar, with previously treated chronic ITP who had a platelet count of &amp;lt; 30 000 /L and who presented with bleeding manifestations. Patients' responses were evaluated after receiving ELT for 3 months or more, as well as their response to different doses of ELT (100, 75, 50, 25, 12.5 or any alternating doses e.g., 50/25). Results: The response rate (platelet count of ‡ 50 000 /L) after 3 months of ELT treatment was comparable in the Arab (87.5%) and non-Arab (88.2%) patients. 26% of the Arab patients required 12.5 or 25 mg and 41.5 % required 50 mg of ELT to achieve an acceptable response. In the Non-Arab Asian group 17.6% required 25 mg and 52.9% required 50 mg of ELT to achieve acceptable response. 22% of the Arab patients and 35.3% of the Non-Arab Asians patients required 75 mg or more of ELT to achieve acceptable control. Further sub-analysis of this data showed that 70% of the Arab patients who achieved complete response were females (14/20) whereas 33.3% (3/9) were Asian females. Two-third the Arab patients who achieved clinical remission were females whereas more than Two-third were non-Arab Asian males. Conclusion: ELT is generally well tolerated and effectively achieves target platelet counts in adult ITP patients. Low doses (12.5 - 50 mg) of ELT were effective in achieving and maintaining safe platelet counts in most Arab patients. This helps in achieving the maximum benefit for the patient at the lower possible dose to prevent toxicity. Tailoring treatment guidelines to match ethnic variations will help in providing a more cost-effective approach for both the patients and the health care system. Disclosures No relevant conflicts of interest to declare.