[Consensus on the clinical diagnosis, treatment and prevention of cancer treatment-induced thrombocytopenia in China (2023 edition)].

Cancer treatment-induced thrombocytopenia (CTIT) is a common adverse event during anti-tumor treatment, of which incidence is related to tumor classification, regimens, course of chemotherapy, etc. CTIT may result in a series of events including bleeding, dose intensity reduction, chemotherapy delay, and in severe cases, even the need for platelet transfusion, ultimately affecting the implementation of treatment plan, increasing the cost of treatment, reducing treatment effect and quality of life, and leading to a poor prognosis. The treatment of CTIT should first identify the cause, assess the risk of bleeding, and then adopt treatment strategies according to the cause and severity of CTIT. The main treatments of CTIT include platelet transfusion, application of various types of platelet-producing drugs, and measures to reduce the additional loss of platelets. Among them, platelet-producing drugs mainly refer to platelet-stimulating factors, including recombinant human thrombopoietin (rhTPO), recombinant human interleukin 11(rhIL-11), and thrombopoietin receptor agonists (TPO-RAs). In addition, traditional Chinese medicine also has some assistance in raising platelets. Pharmacological prophylaxis in high-risk patients may help reduce the incidence and severity of CTIT. This consensus aims to support Chinese oncologists in the diagnosis and treatment of CTIT in China, reduce the risk of bleeding and improve the quality of life of patients.

Thrombopoietin receptor agonists for chemotherapy-induced thrombocytopenia: a new solution for an old problem.

Chemotherapy-induced thrombocytopenia (CIT) is common, resulting in increased bleeding risk and chemotherapy delays, dose reduction, and treatment discontinuation, which can negatively affect oncologic outcomes. The only agent approved by the US Food and Drug Administration to manage CIT (oprelvekin) was voluntarily withdrawn from the market by the manufacturer, leaving few options for patients. Therefore, patients experiencing CIT present a significant clinical challenge in daily practice. The availability of thrombopoietin receptor agonists has led to formal clinical trials describing efficacy in CIT as well as a rather extensive body of published observational data from off-label use in this setting but no formal regulatory indications for CIT to date. The accumulated data, however, have affected National Comprehensive Cancer Network guidelines, which now recommend consideration of TPO-RA clinical trials as well as off-label use of romiplostim. This review article details the evidence to date for the management of CIT with thrombopoietin receptor agonists (TPO-RAs), discussing the efficacy data, the specific circumstances when treatment is warranted (and when it is generally unnecessary), and safety considerations. Specific recommendations regarding patient selection, initiation, dosing, titration, and discontinuation for TPO-RA therapy in CIT are given, based on published data and expert opinion where evidence is lacking.

Alternatives to allogeneic platelet transfusion.

Allogeneic platelet transfusions are widely used for the prevention and treatment of bleeding in thrombocytopenia. Recent evidence suggests platelet transfusions have limited efficacy and are associated with uncertain immunomodulatory risks and concerns about viral or bacterial transmission. Alternatives to transfusion are a well-recognised tenet of Patient Blood Management, but there has been less focus on different strategies to reduce bleeding risk by comparison to platelet transfusion. Direct alternatives to platelet transfusion include agents to stimulate endogenous platelet production (thrombopoietin mimetics), optimising platelet adhesion to endothelium by treating anaemia or increasing von Willebrand factor levels (desmopressin), increasing formation of cross-linked fibrinogen (activated recombinant factor VII, fibrinogen concentrate or recombinant factor XIII), decreasing fibrinolysis (tranexamic acid or epsilon aminocaproic acid) or using artificial or modified platelets (cryopreserved platelets, lyophilised platelets, haemostatic particles, liposomes, engineered nanoparticles or infusible platelet membranes). The evidence base to support the use of these alternatives is variable, but an area of active research. Much of the current randomised controlled trial focus is on evaluation of the use of thrombopoietin mimetics and anti-fibrinolytics. It is also recognised that one alternative strategy to platelet transfusion is choosing not to transfuse at all.

[Economic evaluation of Thrombopoietin Receptor Agonists in the treatment of chronic primary immune thrombocytopenia]. / Evaluación económica del tratamiento de la trombocitopenia inmune primaria crónica refractaria con agonistas del receptor de la trombopoyetina.

PURPOSE: To develop a tool to assist the decision-making for selection of Thrombopoyetin Receptor Agonists of adult patients with chronic immune primary thrombocytopenia (PTI). METHODS: Stochastic cost-effectiveness analysis with a 6-Health- States Markov model: stable, bleeding type 2, 3 or 4, post-type 4 bleeding and death. Each simulation analyzes a randomly generated scenario that describes patients characteristics, results measured in quality adjusted life years (QALYs) and costs (in ?2011). Distributions were obtained from the Spanish data of the European health survey of 2009, the INE estimate of population for 2011 and the 6-months clinical studies for Eltrombopag and Romiplostim. Utility values were obtained from the literature and the costs from Spanish official rates lists. A set of 10.000 random scenarios were generated and the patients evolution of each scenario was simulated during a time horizon of five years (in 2-weeks cycles). National Health System Perspective was used and the annual discount rate was set at 3%. RESULTS: Eltrombopag showed more effectiveness in 9.983 scenarios and there was no difference in 17. In 7.048 scenarios the alternative Eltombopag was dominant. It was cost-effective in another 19 (threshold 30,000 ??/AVAC). CONCLUSIONS: Eltrombopag was the most cost-effective alternative in 70,67% of the simulated scenarios and its use could produce lower costs to the NHS.

Objetivo: Desarrollar una herramienta de apoyo a la decisión en la selección de agonistas del receptor de trombopoyetina en el tratamiento de pacientes adultos con trombocitopenia inmune primaria crónica (PTI) refractaria. Métodos: Análisis coste-efectividad estocástico con un modelo de Markov de seis estados: estable, sangrado tipo 2, 3 ó 4, post-sangrado 4 y muerte. Cada simulación analiza un escenario aleatoriamente generado que describe las características del paciente, los resultados medidos en años de vida ajustados a calidad (AVACs) y los costes (en ?2011). Se obtuvieron distribuciones a partir de los datos para España de la Encuesta Europea de Salud de 2009, de la estimación de población para 2011 del INE, de los estudios a 6 meses de Eltrombopag y Romiplostim, de las utilidades obtenidas de la bibliografía y de las tarifas oficiales en España para procesos y actividad. Se generaron 10.000 escenarios aleatorios y se simuló la evolución de los pacientes de cada escenario durante un horizonte temporal de cinco años (ciclos de dos semanas). Perspectiva del Sistema Nacional de Salud (SNS). Tasa de descuento anual del 3% para costes y efectos. Resultados: En 9.983 escenarios Eltrombopag mostró mayor efectividad y en 17 no hubo diferencias. Eltombopag fue la alternativa dominante en 7.048 escenarios y la más coste efectiva en otros 19 (umbral 30.000 ?/AVAC). Conclusiones: Eltrombopag es la alternativa más coste-efectiva en el 70,67% de los escenarios simulados, por lo que su uso podría producir menores costes al SNS.

Combination of recombinant factor VIIa and fibrinogen corrects clot formation in primary immune thrombocytopenia at very low platelet counts.

Haemostatic treatment modalities alternative to platelet transfusion are desirable to control serious acute bleeds in primary immune thrombocytopenia (ITP). This study challenged the hypothesis that recombinant activated factor VII (rFVIIa) combined with fibrinogen concentrate may correct whole blood (WB) clot formation in ITP. Blood from ITP patients (n = 12) was drawn into tubes containing 3·2% citrate and corn trypsin inhibitor 18·3 µg/ml. WB [mean platelet count 22 × 10(9) /l (range 0-42)] was spiked in vitro with buffer, donor platelets (+40 × 10(9) /l), rFVIIa (1 or 4 µg/ml), fibrinogen (1 or 3 mg/ml), or combinations of rFVIIa and fibrinogen. Coagulation profiles were recorded by tissue factor (0·03 pmol/l) activated thromboelastometry. Coagulation in ITP was characterized by a prolonged clotting time (CT, 1490 s (mean)) and a low maximum velocity (MaxVel, 3·4 mm × 100/s) and maximum clot firmness (MCF, 38·2 mm). Fibrinogen showed no haemostatic effect, whereas rFVIIa reduced the CT and increased the MaxVel. The combination of fibrinogen and rFVIIa revealed a significant synergistic effect, improving all parameters (CT 794 s, MaxVel 7·9 mm × 100/s, MCF 50·7 mm) even at very low platelet counts. These data suggest that rFVIIa combined with fibrinogen corrects the coagulopathy of ITP even at very low platelet counts, and may represent an alternative to platelet transfusion.

Efficacy of radiation countermeasures depends on radiation quality.

The detonation of a nuclear weapon or a nuclear accident represent possible events with significant exposure to mixed neutron/γ-radiation fields. Although radiation countermeasures generally have been studied in subjects exposed to pure photons (γ or X rays), the mechanisms of injury of these low linear energy transfer (LET) radiations are different from those of high-LET radiation such as neutrons, and these differences may affect countermeasure efficacy. We compared 30-day survival in mice after varying doses of pure γ and mixed neutron/γ (mixed field) radiation (MF, Dn/Dt = 0.65), and also examined peripheral blood cells, bone marrow cell reconstitution, and cytokine expression. Mixed-field-irradiated mice displayed prolonged defects in T-cell populations compared to mice irradiated with pure γ photons. In mouse survival assays, the growth factor granulocyte colony-stimulating factor (G-CSF) was effective as a (post-irradiation) mitigator against both γ-photons and mixed-field radiation, while the thrombopoietin (TPO) mimetic ALXN4100TPO was effective only against γ irradiation. The results indicate that radiation countermeasures should be tested against radiation qualities appropriate for specific scenarios before inclusion in response plans.

Supportive care and chelation therapy in MDS: are we saving lives or just lowering iron?

The myelodysplastic syndromes (MDS) are characterized by cytopenias and risk of transformation to acute myeloid leukemia (AML). Although new treatments are available, a mainstay in MDS remains supportive care, which aims to minimize the impact of cytopenias and transfusion of blood products. Red blood cell (RBC) transfusions place patients at risk of iron overload (IOL). In beta-thalassemia major (BTM), IOL from chronic RBC transfusions inevitably leads to organ dysfunction and death. With iron chelation therapy (ICT), survival in BTM improved from the second decade to near normal and correlated with ICT compliance. Effects of ICT in BTM include reversal of cardiac arrhythmias, improvement in left ventricular ejection fraction, arrest of hepatic fibrosis, and reduction of glucose intolerance. It is not clear whether these specific outcomes are applicable to MDS. Although retrospective, recent studies in MDS suggest an adverse effect of transfusion dependence and IOL on survival and AML transformation, and that lowering iron minimizes this impact. These data raise important points that warrant further study. ICT is potentially toxic and cumbersome, is costly, and in MDS patients should be initiated only after weighing potential risks against benefits until further data are available to better justify its use. Since most MDS patients eventually require RBC transfusions, the public health implications both of transfusion dependence and ICT in MDS are considerable. This paper summarizes the impact of cytopenias in MDS and treatment approaches to minimize their impact, with a focus on RBC transfusions and their complications, particularly with respect to iron overload.

Trombocitosis en la consulta de oncohematología. Descripción, diagnóstico etiológico y evolución / Thrombocytosis in the oncology-haematology clinic: Description, aetiological diagnosis and outcome

Introducción: La trombocitosis es un motivo frecuente de consulta en oncohematología infantil, y precisa un importante número de visitas y determinaciones analíticas para su diagnóstico o resolución. Nuestro objetivo ha sido evaluar las características de los pacientes derivados a nuestro servicio en los últimos meses para estudio de trombocitosis y establecer cuáles deberían ser los pacientes que precisan un estudio más exhaustivo en el hospital. Pacientes y métodos: Se determina en 33 pacientes derivados por este motivo el sexo, el rango de edad, la procedencia, los antecedentes personales y familiares, el grado de trombocitosis por la que se consulta y la cifra máxima durante el seguimiento, el motivo por el que se realiza la primera analítica, los valores hematimétricos en la primera analítica (hemoglobina [Hb], volumen corpuscular medio [VCM], hemoglobina corpuscular media [HCM], leucocitos [linfocitos y neutrófilos]), las exploraciones complementarias realizadas, el diagnóstico y el número de visitas que precisaron antes del alta. Se clasifica la trombocitosis en leve (500-700 × 103/μl), moderada (700-900 × 103/μl), grave (900-1.000 × 103/μl) y extrema (> 1.000 × 103/μl). Resultados: No hubo predominancia de sexos. El 45 % de los pacientes eran menores de 2 años. Procedían en un 55 % de su centro de salud. La cifra media de plaquetas por la que consultaron fue de 669.000 (trombocitosis leve). En el seguimiento llegaron a cifras extremas el 24 %. En el 28 % la analítica se había realizado por un cuadro infeccioso. La exploración complementaria más solicitada fue el metabolismo del hierro (en el 82 %). Todos se corresponden con trombocitosis secundarias (el 48 % reactivas a infecciones, el 24 % secundarias a ferropenia y el 15 % por ambas causas). El número medio de visitas ha sido de 5,12. Conclusiones: El hallazgo de la trombocitosis es en la mayoría de los casos casual o en el contexto de un cuadro infeccioso y, además, son leves. Dada la baja incidencia de trombocitosis primaria en la infancia y que el diagnóstico es de exclusión, se debería iniciar el estudio de la misma en la consulta de atención primaria, descartando inicialmente una causa infecciosa, inflamatoria o secundaria a sangrado. Una vez descartadas estas causas la prueba complementaria más rentable es el metabolismo del hierro, dada la asociación de ferropenia con trombocitosis. Si también se excluye esta etiología y se comprueba la persistencia de la trombocitosis, estaría indicado derivar al paciente a un servicio de oncohematología infantil para completar estudio (AU)

Introduction: Thrombocytosis is a common cause of patient referral to a paediatric haematologist specialist which requires a significant number of laboratory tests and visits to confirm the diagnosis. The aim of our study has been to analyse the characteristics of patients referred to our centre for specialised thrombocytosis assessment. Based on this assessment we established the criteria patients must fulfil to be recommend for further hospital study. Patients and methods: We categorised the 33 patients referred for thrombocytosis assessment according to sex, age, origin, personal and family history, platelet count at diagnosis and the reason why the red and white blood count at diagnosis (Haemoglobin, mean corpuscular volume, meen corpuscular haemoglobin, leukocyte, neutrophils, lymphocyte and monocyte count), maximum platelet count during follow-up and other complementary examinations were done. The final diagnosis itself and number of previous visits before were also considered. The classification used to grade thrombocytosis was: low (500-700 × 103/μl), mild (700-900 × 103/μl), severe (900-1.000 × 103/μl) and extreme (> 1.000 × 103/μl). Results: There was no predominance of males or females. 45 % of patients were under 2 years old and 55 % of them came from their primary care centre. The mean platelet count at the first medical visit was 669,000 (mild thrombocytosis). During follow-up, 24 % of the patients reached extreme platelets levels. In 28 % the initial blood count was performed because of an infection. The most frequently requested laboratory test was iron metabolism (82 % of the cases). All cases correspond to secondary thrombocytosis (48 % were reactive to infections, 24 % secondary to iron deficiency, and 15 % were associated to both causes). The mean number of visits before hospital discharge was 5.12. Conclusions: The finding of thrombocytosis in the majority of the cases studied was casual or in the context of an infectious process. Most of the thrombocytosis were mild. Due to the extremely low incidence of primary thrombocytosis in childhood and the fact that diagnosis is made by exclusion of other possibilities, the initial study of these patients should be done in primary care centres. The first conditions to be ruled out are infectious, inflammatory or bleeding processes. Once these causes are excluded, the most useful complementary test is to measure iron level given the relation between iron deficiency and thrombocytosis. Once these causes are ruled out and thrombocytosis persists, it would then be indicated to refer the patient to a paediatric oncology-haematology department for a more exhaustive follow-up (AU)

Cytokines in combination to treat radiation-induced myelosuppresssion: evaluation of SCF + glycosylated EPO + pegylated G-CSF as an emergency treatment in highly irradiated monkeys.

Multicytokine therapy may be useful to counteract radiation-induced myelosuppression. We assessed the stem cell factor + glycosylated erythropoietin + pegylated granulocyte colony-stimulating factor combination (SEG) as an emergency treatment. SEG in highly irradiated monkeys efficacy appeared to be restricted to granulopoiesis. Early administration of Erythropoietin did not prevent radiation-induced anemia.

Thrombopoietin protects against in vitro and in vivo cardiotoxicity induced by doxorubicin.

BACKGROUND: Doxorubicin (DOX) is an important antineoplastic agent. However, the associated cardiotoxicity, possibly mediated by the production of reactive oxygen species, has remained a significant and dose-limiting clinical problem. Our hypothesis is that the hematopoietic/megakaryocytopoietic growth factor thrombopoietin (TPO) protects against DOX-induced cardiotoxicity and might involve antiapoptotic mechanism exerted on cardiomyocytes. METHODS AND RESULTS: In vitro investigations on H9C2 cell line and spontaneously beating cells of primary, neonatal rat ventricle, as well as an in vivo study in a mouse model of DOX-induced acute cardiomyopathy, were performed. Our results showed that pretreatment with TPO significantly increased viability of DOX-injured H9C2 cells and beating rates of neonatal myocytes, with effects similar to those of dexrazoxane, a clinically approved cardiac protective agent. TPO ameliorated DOX-induced apoptosis of H9C2 cells as demonstrated by assays of annexin V, active caspase-3, and mitochondrial membrane potential. In the mouse model, administration of TPO (12.5 microg/kg IP for 3 alternate days) significantly reduced DOX-induced (20 mg/kg) cardiotoxicity, including low blood cell count, cardiomyocyte lesions (apoptosis, vacuolization, and myofibrillar loss), and animal mortality. Using Doppler echocardiography, we observed increased heart rate, fractional shortening, and cardiac output in animals pretreated with TPO compared with those receiving DOX alone. CONCLUSIONS: These data have provided the first evidence that TPO is a protective agent against DOX-induced cardiac injury. We propose to further explore an integrated program, incorporating TPO with other protocols, for treatment of DOX-induced cardiotoxicity and other forms of cardiomyopathy.

Safety and efficacy of transfusions of autologous cryopreserved platelets derived from recombinant human thrombopoietin to support chemotherapy-associated severe thrombocytopenia: a randomised cross-over study.

BACKGROUND: The increasing demand for platelet products, and concern over the transfusion-associated risks of alloimmunisation and infections, have motivated a search for improved methods aimed at keeping exposure to donor antigens to a minimum. Transfusion of thrombopoietin-derived autologous platelets might provide an alternative strategy. We aimed to compare the safety and efficacy of this strategy with that of transfusion with fresh allogeneic platelets in patients with severe chemotherapy-induced thrombocytopenia. METHODS: 20 patients with gynaecological malignancies were treated with two doses of 1.2 microg/kg recombinant human thrombopoietin. From day 12, we aimed to collect 50 units of platelets from these patients by plateletpheresis. Harvested platelets were cryopreserved in ThromboSol and 2% dimethyl sulfoxide (DMSO) for use in subsequent autologous transfusions. Patients then received carboplatin for up to six cycles. Patients were randomly assigned to group A (n=10), which received allogeneic fresh platelets at the first instance of severe thrombocytopenia (platelet count <15,000/microL) and then autologous cryopreserved platelets at the next, or to group B (n=10), which received first autologous and then allogeneic platelets. In subsequent cycles, all patients received autologous platelets while available. The primary endpoint was platelet count increment corrected for the number of platelets transfused and the patients' body-surface area. Analysis was by intention to treat. FINDINGS: Treatment with recombinant human thrombopoietin significantly increased platelet count (median 2.3-fold [range 1.5-3.3], p<0.0001) in all but one patient in group A. The median number of platelets collected per patient was 53 units (14-66) in two collections (one to three). There was no significant difference in the corrected platelet count increments (CCIs) between the 19 paired transfusions of cryopreserved autologous platelets and fresh allogeneic platelets (median 1-h CCI 15.7 vs 19.8, p=0.398; median 24-h CCI 13.0 vs 18.1, p=0.398). 14 of the 19 patients had a good response (1-h CCI >7.5) to their first transfusion of allogeneic platelets. By contrast, all patients had a good response to their first transfusion of autologous platelets (p=0.063). Moreover, no significant decrease in the CCIs (p=0.405) was seen over six cycles after autologous platelet transfusions (n=63). No transfusion reactions or any serious adverse event was recorded during autologous platelet transfusions. INTERPRETATION: Recombinant human thrombopoietin facilitated collection of multiple units of platelets, which could be cryopreserved and reinfused to counteract severe thrombocytopenia during multicycle chemotherapy. Transfusion of autologous cryopreserved platelets derived from recombinant human thrombopoietin can provide a viable strategy to minimise the risks of allogeneic platelet transfusions and provide a long-lasting supply of platelet support.

Single administration of thrombopoietin to lethally irradiated mice prevents infectious and thrombotic events leading to mortality.

A sufficiently high dose of thrombopoietin to overcome initial c-mpl-mediated clearance stimulates hematopoietic reconstitution following myelosuppressive treatment. We studied the efficacy of thrombopoietin on survival after supralethal total body irradiation (9 Gy) of C57BL6/J mice and the occurrence of infectious and thrombotic complications in comparison with a bone marrow graft or prophylactic antibiotic treatment. Administration of 0.3 microg thrombopoietin, 2 hours after irradiation, protected 62% of the mice as opposed to no survival in placebo controls. A graft with a supraoptimal number of syngeneic bone marrow cells (10(6) cells) fully prevented mortality, whereas antibiotic treatment was ineffective. Blood cell recovery was observed in the thrombopoietin-treated mice but not in the placebo or antibiotic-treated group. Bone marrow and spleen cellularity as well as colony-forming unit granulocyte-macrophage and burst-forming unit erythroid were considerably increased in thrombopoietin-treated mice relative to controls. Histologic examination at day 11 revealed numerous petechiae and vascular obstructions within the brain microvasculature of placebo-treated mice, which was correlated with hypercoagulation and hypofibrinolysis. Thrombopoietin treatment prevented coagulation/fibrinolysis disorder and vascular thrombosis. High fibrinogen levels were related to bacterial infections in 67% of placebo-treated mice and predicted mortality, whereas the majority of the thrombopoietin-treated mice did not show high fibrinogen levels and endotoxin was not detectable in plasma. We conclude that thrombopoietin administration prevents mortality in mice subjected to 9-Gy total body irradiation both by interfering in the cascade leading to thrombotic complications and by amelioration of neutrophil and platelet recovery and thus protects against infections and hemorrhages.

Use of thrombopoietic growth factors in acute leukemia.

Several hematopoietic growth factors have been shown to affect megakaryocyte development, and two, interleukin (IL)-11 and thrombopoietin (TPO) are presently being evaluated for use in patients with thrombocytopenia. In two studies patients who required one or more platelet transfusions during their first course of chemotherapy were found to require fewer platelet transfusions if their second cycle was augmented with IL-11. The drug was generally safe, with cardiovascular compromise the only significant complication occurring in a minority of patients. Although these reports included patients with various malignancies, studies of IL-11 in patients with myeloproliferative disorders have not been presented. In several clinical trials in cancer patients treatment with TPO was safe, and when administered early following a moderately aggressive cytotoxic insult was effective in accelerating platelet recovery. In addition, in both pre-clinical and clinical trials, TPO given to stem cell donors during mobilization lead to accelerated hematopoietic recovery. Finally, TPO appears safe when administered to patients with acute myelogenous leukemia (AML), both with respect to acute toxicity and long-term outcome of the leukemia. However, when used following a 7-day course of standard chemotherapy, the agent does not appear to accelerate platelet recovery. As such, additional clinical trials to test different growth factor regimens are ongoing. A number of studies have suggested that megakaryocytic growth factors may play a role in the biology of myeloproliferative disorders. Given the potential for adversely affecting patients with these disorders, the affects of IL-11 or TPO in patients with AML must continue to be carefully studied.

Memorial lecture. Megakaryocytic growth factors: is there a new approach for management of thrombocytopenia in patients with malignancies?

C-mpl ligand acts primarily as a lineage-specific hematopoietic growth factor by promoting proliferation of megakaryocyte precursors and their differentiation into megakaryocytes and platelets. In addition to the ability of c-mpl ligand to support megakaryocytic development from CD34+ precursor cells, several lines of evidence also point to a stimulatory effect on hematopoietic stem cells. When recombinant thrombopoietin or pegylated megakaryocyte growth and development factor is administered to normal animals or humans, there is a dose-dependent increase in the platelet count. When administered following chemotherapy in animal models or humans, c-mpl ligands reduce the duration and sometimes the degree of thrombocytopenia. The issue of whether clinically relevant thrombocytopenia can be ameliorated has so far been more difficult to resolve. Because severe thrombocytopenia is not commonly seen with standard chemotherapy regimens, clinical studies examining c-mpl ligands for their ability to ameliorate chemotherapy-induced thrombocytopenia will focus on treatment of acute leukemias and bone marrow transplantation. The potential utility of c-mpl ligands for treatment of myelodysplastic syndromes, aplastic anemias, or in HIV infection, will have to be evaluated in the future. Possibly the greatest potential of thrombopoietic growth factors in the near future may be in transfusion medicine, to collect and to store platelets from healthy donors or in autologous settings.

Update on thrombopoietin in preclinical and clinical trials.

Thrombopoietin, also termed the c-Mpl ligand, is a lineage-dominant hematopoietic factor that primarily regulates megakaryopoiesis and thrombopoiesis. Treatment of normal animals with recombinant human megakaryocyte growth and development factor, a truncated molecule of the c-Mpl ligand, which is modified with polyethylene glycol (PEG-rHuMGDF), and glycosylated recombinant thrombopoietin stimulates the expansion of bone marrow megakaryocytes and their progenitors, and greatly enhances the production of morphologically and functionally normal platelets. In contrast, this cytokine has only minimal effects on peripheral leukocyte and erythrocyte counts. In myelosuppressed animals, PEG-rHuMGDF or glycosylated thrombopoietin accelerates multilineage hematopoietic recovery effectively improving thrombocytopenia and, in most models, leukopenia (or neutropenia) and anemia. In addition to daily multiple injections, even a single injection of PEG-rHuMGDF after myelosuppressive treatment is fully effective for hematopoietic recovery. In clinical trials, PEG-rHuMGDF or glycosylated recombinant human thrombopoietin potently stimulates thrombopoiesis in cancer patients before chemotherapy. The administration of PEG-rHuMGDF alone or in combination with recombinant human granulocyte colony-stimulating factor (rhG-CSF) reduces the duration of severe thrombocytopenia and in some cases platelet nadirs in patients with advanced cancers after dose-intensive chemotherapy. The recombinant hormone is well-tolerated with little drug-related toxicity.

Multilineage hematopoietic recovery by a single injection of pegylated recombinant human megakaryocyte growth and development factor in myelosuppressed mice.

Previous studies have shown that daily multiple administration of pegylated recombinant human megakaryocyte growth and development factor (PEG-rHuMGDF) markedly stimulates thrombopoiesis and effectively ameliorates thrombocytopenia, and in most cases anemia and neutropenia, in myelosuppressed animals. In this study, we evaluated the effects of a single intravenous injection of PEG-rHuMGDF on hematopoietic recovery after sublethal total-body irradiation in mice. A single injection of PEG-rHuMGDF (1 to 640 microg/kg) 1 hour after irradiation accelerated platelet, red blood cell (RBC), and white blood cell (WBC) recovery in a dose-dependent fashion. In the bone marrow of vehicle-treated mice, megakaryocytic, erythroid, and myeloid progenitors, as well as day 12 colony-forming unit-spleen (CFU-S), were dramatically decreased much earlier than the nadirs of peripheral blood cells, whereas megakaryocytes were modestly decreased. Treatment with PEG-rHuMGDF (80 microg/kg, an optimal dose) 1 hour after irradiation resulted in more rapid recovery of these four hematopoietic progenitors and also significantly facilitated megakaryocyte recovery. In addition, the same PEG-rHuMGDF administration schedule expanded bone marrow cells capable of rescuing lethally irradiated recipient mice. As the interval between irradiation and PEG-rHuMGDF treatment was longer, its effects on hematopoietic recovery were attenuated. In contrast to the effects of PEG-rHuMGDF, a single injection of recombinant human granulocyte colony-stimulating factor (rhG-CSF) 1 hour after irradiation exclusively accelerated WBC recovery, but only to a similar extent as PEG-rHuMGDF (80 microg/kg) treatment even when rhG-CSF doses were escalated to 1,000 microg/kg. This appeared related to different pharmacokinetics of these two factors after a single injection in irradiated mice. The concentrations of PEG-rHuMGDF after injection persisted in the plasma for a longer time compared with rhG-CSF. These results indicate that a single injection of PEG-rHuMGDF at an early time after irradiation is able to effectively improve thrombocytopenia, anemia, and leukopenia with concomitant accelerated recovery of both primitive and committed hematopoietic progenitors in irradiated mice. Our data also show that compared with the rhG-CSF shown to exert multilineage effects on hematopoiesis, PEG-rHuMGDF has more wide-ranging effects on peripheral blood cell recovery.

Molecular cloning and functional expression of feline thrombopoietin.

Feline thrombopoietin (TPO) was molecularly cloned to establish a basis for cytokine therapy of thrombocytopenia in cats. cDNA clones covering the whole coding sequence of feline TPO were isolated from feline liver. The feline TPO cDNA obtained in this study contained an open reading frame encoding 349 amino acid residues. The predicted amino acid sequence of feline TPO shared 78.7, 69.9, 72.9 and 83.0% similarity with sequences of human, murine, rat and canine TPO, respectively. Four cysteine residues and two of four N-glycosylation sites that are conserved among species were also found at the corresponding positions in feline TPO. The feline TPO cDNA fragment encoding the whole amino acid coding region was recloned into an expression vector, and the resulting vector was transfected into 293T cells using the calcium phosphate method. The supernatant of the transfected 293T cells stimulated the proliferation of a human megakaryoblastic leukemia cell line (UT-7/TPO) cells in a dose dependent manner, indicating that the feline TPO cDNA obtained in this study encodes biologically active feline TPO.

Improvement of thrombocytopenia following bone marrow transplantation by pegylated recombinant human megakaryocyte growth and development factor in mice.

We examined whether pegylated recombinant human megakaryocyte growth and development factor (PEG-rHuMGDF) is capable of improving thrombocytopenia and promoting thrombopoietic reconstitution following lethal irradiation and bone marrow transplantation (BMT) in mice. Immediately after receiving 10 Gy whole body irradiation (day 0), male C3H/HeN mice were inoculated with 10(6) bone marrow cells obtained from syngeneic mice. Circulating platelet counts decreased to below 4% of the normal counts with a nadir on day 10, and then returned to the normal level on day 28 in the control mice undergoing BMT. Subcutaneous consecutive treatment with PEG-rHuMGDF at doses from 10 to 300 micrograms/kg/day from day 1 for 13 days significantly improved the platelet nadir and promoted platelet recovery. The white blood cell counts and hemoglobin concentration following BMT were not influenced by the PEG-rHuMGDF. PEG-rHuMGDF-injection starting from day 5 did not improve the platelet nadir following BMT. Furthermore, administration with PEG-rHuMGDF on alternate days at 55.7 micrograms/kg/day for 7 days or at an interval of 3 days at 78 micrograms/kg/day for 4 days (twice a week for 2 weeks) had a significant efficacy, but these administration regimens had less efficacy than consecutive administration at 30 micrograms/kg/day for 13 days. The numbers of megakaryocytes and megakaryocyte progenitor cells decreased to 5 and 0.2% of normal level, respectively, in the control mice. Consecutive administration of PEG-rHuMGDF enhanced the recovery of the mean number of these cells compared to those in vehicle-treated mice, although such effects were not statistically significant except for the number of megakaryocyte progenitors on day 12. These results suggest that consecutive treatment with PEG-rHuMGDF beginning from the day after BMT may be effective in improving thrombocytopenia following BMT.

The thrombocytopenia of cancer. Prospects for effective cytokine therapy.

The previous 10 years have witnessed the development of increasing needs for platelet transfusion in support of aggressive therapies of malignancy. Despite gains in our understanding of platelet preparation, storage, and transfusion, alternative therapies are clearly desirable. During the late 1980s at least six distinct cytokines that display effects on megakaryocyte growth and differentiation-IL-3, KL, GM-CSF, IL-6, IL-11, and LIF- and a synthetic growth factor, PIXY 321, were cloned and characterized. Although none of these cytokines fulfill all of the physiologic roles of thrombopoietin, in its absence several have undergone extensive preclinical and preliminary clinical testing. Of these, IL-11 and PIXY 321 hold promise for clinical amelioration of thrombocytopenia in cancer patients. With the recent cloning of thrombopoietin and its promise in preclinical trials, the role of each of these recombinant proteins in clinical medicine is undergoing careful evaluation. As with erythropoietin and G-CSF before it, given its normal role in the regulation of platelet production, Tpo would appear to provide the greatest physiologic stimulus to platelet production in states of natural and iatrogenic marrow failure. Careful clinical trials of the agent are needed to determine whether the hormone will fulfill this promise. The following decade will most certainly see the resolution of many of the complications of thrombocytopenia and its transfusional support.