Thrombocytopenia is a common medical problem for which the main treatment is platelet transfusion. Given the increasing use of platelets and the declining donor population, identification of a safe and effective platelet growth factor could improve the management of thrombocytopenia. Thrombopoietin (TPO), the c-Mpl ligand, is the primary physiologic regulator of megakaryocyte and platelet development. Since the purification of TPO in 1994, 2 recombinant forms of the c-Mpl ligand—recombinant human thrombopoietin (rhTPO) and pegylated recombinant human megakaryocyte growth and development factor (PEG-rHuMGDF)—have undergone extensive clinical investigation. Both have been shown to be potent stimulators of megakaryocyte growth and platelet production and are biologically active in reducing the thrombocytopenia of nonmyeloablative chemotherapy. However, neither TPO has demonstrated benefit in stem cell transplantation or leukemia chemotherapy. Other clinical studies have investigated the use of TPO in treating chronic nonchemotherapy-induced thrombocytopenia associated with myelodysplastic syndromes, idiopathic thrombocytopenic purpura, thrombocytopenia due to human immunodeficiency virus, and liver disease. Based solely on animal studies, TPO may be effective in reducing surgical thrombocytopenia and bleeding, ex vivo expansion of pluripotent stem cells, and as a radioprotectant. Ongoing and future studies will help define the clinical role of recombinant TPO and TPO mimetics in the treatment of chemotherapy- and nonchemotherapy-induced thrombocytopenia.

Introduction

Thrombocytopenia is a common problem in the management of patients with cancer and other conditions that affect hematopoietic cells. Thrombocytopenia may be occasionally encountered with conventional chemotherapy regimens used to treat solid tumors but can be a major clinical problem in the management of patients receiving dose-intensive chemotherapy, induction and consolidation therapy for leukemia, palliative chemotherapy following multiple previous regimens, and multiple cycles of certain chemotherapeutic regimens.1Multiagent regimens such as MAID (mesna, adriamycin, ifosfamide, and dacarbazine) and ICE (ifosfamide, carboplatin, and etoposide) used in the treatment of lymphoma, sarcoma, breast, ovarian, and germ cell tumors often produce thrombocytopenia that requires dose modifications, platelet transfusions, or both to prevent bleeding complications.1,2 Thrombocytopenia associated with the use of newer chemotherapy agents such as gemcitabine may limit their use in patients with lung, breast, or ovarian cancer. Additionally, patients with associated bone marrow failure have a higher risk of severe thrombocytopenia and bleeding complications with any chemotherapy regimen.

Thrombocytopenia is also a frequent problem in the management of nonchemotherapy patients with myelodysplastic syndrome (MDS), idiopathic thrombocytopenic purpura (ITP), chronic liver disease, and acquired immunodeficiency syndrome (AIDS).3-6The chronic thrombocytopenia observed in these conditions results from defective or diminished platelet production or enhanced immunologic and nonimmunologic platelet destruction and may be associated with abnormal platelet function.3-6 Furthermore, patients undergoing liver transplantation, cardiovascular surgery, requiring intra-aortic balloon counterpulsation, or receiving supportive intensive care often experience severe, acute thrombocytopenia that is associated with increased mortality.7,8 

Platelet transfusion therapy is currently the only acute treatment for severe thrombocytopenia. Although temporarily effective in controlling severe thrombocytopenia, platelet transfusion therapy is associated with several problems, including refractoriness and alloimmunization, transmission of infectious agents, and transfusion reactions.9-15 The limited supply of blood products can also be problematic. The use of dose-intensive chemotherapy regimens and hematopoietic progenitor cell transplantation, as well as intensive support for the medical and surgical patient, has resulted in an increasing demand for platelet products; this demand is likely to escalate in an attempt to improve clinical outcomes for oncology and nononcology patients. The limitations of platelet transfusions and the increased costs associated with the complications of such transfusions have prompted a search for growth factors that stimulate platelet production, thereby reducing or eliminating the need for platelet transfusions.1,2 

Thrombopoietic growth factors

Over the past 2 decades, a number of hematopoietic growth factors with thrombopoietic activity have been identified, including recombinant granulocyte-macrophage colony-stimulating factor (GM-CSF); stem cell factor (c-kit ligand or steel factor); interleukin 1 (IL-1), IL-3, IL-6, and IL-11; and thrombopoietin (TPO).16-26Early clinical studies of many of these cytokines, including IL-1, IL-3, IL-6, and IL-11, showed their ability to stimulate platelet production directly or indirectly in patients with chemotherapy-induced thrombocytopenia.27-32 In phase 1 studies, administration of IL-1α before or after carboplatin therapy increased platelet counts and was effective at attenuating thrombocytopenia associated with chemotherapy.27,32 Similarly, both IL-6 and IL-11 have been shown to produce an increase in platelet counts and accelerate platelet recovery after chemotherapy.29,30Despite its relatively modest effect on megakaryocyte and platelet production, IL-11 has been shown to reduce the need for platelet transfusions in patients with chemotherapy-induced thrombocytopenia.31,33 

Although ILs stimulate thrombopoiesis, their action on platelets is not their principal physiologic function. Recently, gene-targeting studies have shown that the primary physiologic function of IL-11 is to maintain female fertility; it is not essential for hematopoiesis either in normal physiology or in response to hematopoietic stress.34,35 Furthermore, the pleiotropic effect of ILs often results in unwanted or unacceptable toxic effects, including hyperbilirubinemia, rapid induction of anemia, fever, fatigue, chills, hypotension, and headache.27,32,36-38 Although administration of IL-11 reduces the need for platelet transfusions by approximately a third in patients with severe chemotherapy-induced thrombocytopenia, it is associated with mild peripheral edema, dyspnea, conjunctival redness, and a low incidence of atrial arrhythmias and syncope.30,33 Thus, despite the ability of ILs to ameliorate thrombocytopenia in a subset of patients treated with conventional chemotherapy, the moderate toxicity encountered with IL treatment may interfere with its therapeutic effect and potential use as a thrombopoietic agent.

In contrast to ILs, TPO, also known as c-Mpl ligand, is a relatively lineage-specific cytokine that stimulates megakaryocyte growth and maturation in vitro and is a potent in vivo thrombopoietic growth factor. Gene-targeting studies have established that TPO is the most important physiologic regulator of steady-state megakaryocyte and platelet production.39-41 Cloning of the c-Mpl ligand led to the clinical development of various preparations of TPO, including recombinant human thrombopoietin (rhTPO) and pegylated recombinant human megakaryocyte growth and development factor (PEG-rHuMGDF). This article presents an overview of the biology of the recombinant thrombopoietins, the results of all phase 1 and 2 clinical studies of different preparations of TPO, and the problems encountered in their development.

TPO and its biology

Isolation of c-Mpl ligand

The identification of TPO as the ligand for the c-Mpl receptor was heralded with much enthusiasm, as almost 40 years had elapsed since the proposal of the existence of a factor that regulates megakaryocytopoiesis and generation of platelets.42 As is often the case in research, several independent laboratories simultaneously reported the identification and molecular cloning of TPO.17-21 A sentinel discovery that preceded the purification and molecular cloning of TPO was the cloning of the retroviral oncogene, v-mpl, from the murine myeloproliferative leukemia virus.43 Subsequently, the cellular homologue, c-mpl, was cloned and shown to encode a membrane protein that possessed substantial homology with receptors for ILs and colony-stimulating factors, indicating that it might function as a hematopoietic receptor.44 Further support for the role of c-Mpl as the putative thrombopoietin receptor is provided by the presence of c-Mpl mRNA and protein primarily in platelets, megakaryocytes, and a subpopulation of CD34+ cells and the absolute requirement for the presence of a functional c-Mpl to stimulate progenitor cells of the megakaryocyte lineage in bone marrow cultures.45 Several groups were able to isolate, purify, and clone its ligand, which was initially referred to as the c-Mpl ligand, megakaryocyte growth and development factor, megapoietin, or thrombopoietin.17-21 

Structure and biologic properties of TPO

TPO is synthesized primarily in the liver as a single 353-amino acid precursor protein. On removal of the 21-amino acid signal peptide, the mature molecule consists of 2 domains: a receptor-binding domain that shows considerable homology to erythropoietin and a carbohydrate-rich carboxy-terminus of the protein that is highly glycosylated and important in maintaining protein stability (Figure1).18,19,46-48 

Fig. 1.

Domain structure shows features unique to endogenous TPO.

The amino terminus (first 153 amino acids) of TPO contains 4 conserved cysteine residues and is 17% identical to erythropoietin (EPO; ∼50% identical if neutral substitutions are taken into account). It contains the entire receptor-binding region. The shaded boxes show the predicted α-helical regions of TPO. The carboxyl terminus (amino acids 154 to 332) of the molecule appears to be unique to TPO and contains theN-linked glycosylation sites indicated by solid arrows. Adapted from Foster and Hunt47 with permission.

Fig. 1.

Domain structure shows features unique to endogenous TPO.

The amino terminus (first 153 amino acids) of TPO contains 4 conserved cysteine residues and is 17% identical to erythropoietin (EPO; ∼50% identical if neutral substitutions are taken into account). It contains the entire receptor-binding region. The shaded boxes show the predicted α-helical regions of TPO. The carboxyl terminus (amino acids 154 to 332) of the molecule appears to be unique to TPO and contains theN-linked glycosylation sites indicated by solid arrows. Adapted from Foster and Hunt47 with permission.

TPO levels usually increase in response to the decline in platelet mass and remain elevated during persistent thrombocytopenia. Although hepatic transcription and translation of the TPO gene appears to be constant,49,50 most studies indicate that the circulating platelet mass directly determines the circulating level of TPO.20,51-56 Transfusion of platelets into thrombocytopenic animals or humans has resulted in a decrease in plasma TPO levels20,52-54,57 and similar results have been observed when normal platelets are transfused into c-Mpl–deficient mice.51 These findings indicate that TPO is constitutively synthesized in the liver and removed from circulation by binding to the c-Mpl receptor on platelets and possibly bone marrow megakaryocytes.

Some investigators have alternatively suggested that local production of TPO by bone marrow stromal cells is increased during thrombocytopenia and stimulates megakaryocyte growth.58Direct evidence to support the relative contribution of this mechanism to platelet production is lacking; but in experiments in which livers from TPO−/− mice were transplanted into normal mice, at least 60% of the platelet production could be accounted for by hepatic TPO production.59 Furthermore, in patients with hepatic failure undergoing liver transplantation, the low platelet counts and undetectable TPO levels before transplantation became normal after transplantation, suggesting that the liver is responsible for virtually all TPO production.60,61 

As predicted, TPO increases the size, ploidy, and number of megakaryocytes and stimulates the expression of platelet-specific markers.20,62,63 In addition to acting as a potent megakaryocyte colony-stimulating factor, TPO has a synergistic effect on the growth of myeloid and erythroid precursors when combined with other hematopoietic growth factors such as erythropoietin or stem cell factor.64-66 The role of TPO as the principal physiologic regulator of platelet production has been confirmed in studies of mutant mice lacking the ability to produce either TPO (TPO−/−) or its receptor (c-Mpl−/−).67-71 Genetic elimination of TPO or c-Mpl results in an 85% to 95% reduction in the number of circulating platelets, megakaryocytes, and megakaryocyte progenitor cells.39,70,71 Although TPO-deficient mice are severely thrombocytopenic, they are healthy and show no signs of spontaneous hemorrhage, implying that TPO-independent mechanisms for platelet production exist.70 Recently, “double knock-out” mice that lack the genes for c-Mpl and one other growth factor or its receptor (GM-CSF, IL-3, IL-11, IL-6, or leukemia inhibitory factor) have been created to investigate this observation. The double knock-out mice had no additional defect in platelets or their precursors, indicating that GM-CSF, IL-3, IL-11, IL-6, or leukemia inhibitory factor alone are not responsible for the basal platelet production seen in the absence of TPO signaling.40,41,72 

TPO and its receptor also have a major effect on production of primitive pluripotent stem cells and progenitor cells from other lineages. Despite normal red and white blood cell numbers, mice deficient in TPO (TPO−/−) or c-Mpl (c-Mpl−/−) exhibit a 60% to 70% reduction in the number of erythroid and myeloid progenitor cells compared with control animals.39,70 In addition, the ability of hematopoietic cells from these animals to reconstitute the hematopoietic organs of irradiated normal mice is also substantially reduced.73Administration of TPO to TPO−/− mice expanded the bone marrow and spleen progenitor pools of all hematopoietic lineages and increased the number of circulating platelets.39 

Despite its activity on hematopoietic stem cells and early megakaryocytopoiesis, TPO has little effect on platelets and on the late stages of megakaryocyte development.74-77 This contrasts with granulocyte colony-stimulating factor (G-CSF) and GM-CSF, for which an action on late myeloid precursor cells and mature myeloid cells is well established.78,79 Perhaps the most significant consequence of TPO's minimal effect on late-stage megakaryocytes is the inability of TPO to hasten platelet shedding from megakaryocytes. In fact, if anything, TPO inhibits platelet shedding.74 

Although TPO does not directly cause platelet activation, at pharmacologically high doses it does have a modest effect on mature platelets by increasing their reactivity to some aggregation stimuli; TPO-treated platelets require half as much adenosine diphosphate for a response.80,81 Other hematopoietic growth factors also reduce the threshold for platelet activation but the clinical relevance is uncertain. This effect may be mediated by TPO-dependent activation of phosphatidylinositide 3-kinase, which in turn phosphorylates Thr306 and Ser473 of platelet protein kinase Bα, an important antiapoptotic protein.82 However, TPO does not prevent apoptosis of platelets during ex vivo storage.83,84 

Recombinant TPO

Since TPO was first cloned, several recombinant TPOs have been developed for clinical evaluation. There are 2 of these preparations, rhTPO and PEG-rHuMGDF (Figure 2), that have undergone considerable preclinical and clinical evaluation. The amino acid sequence of rhTPO (Pharmacia, Peapack, NJ; formerly developed by Genentech, South San Francisco, CA) is identical to that of endogenous TPO. rhTPO is produced in mammalian cells and is glycosylated. Nonetheless, its molecular weight is 90 kd, less than the 95 kd of the native molecule.85 PEG-rHuMGDF (Amgen, Thousand Oaks, CA), is produced in Escherichia coli and consists of the receptor-binding, 163 amino-terminal amino acids of native TPO. It is conjugated to a 20-kd polyethylene glycol moiety to increase its circulatory half-life and possesses all the biologic activity of native TPO.86 These 2 recombinant thrombopoietins have similar pharmacologic characteristics and show profound in vitro and in vivo effects on megakaryocyte development and platelet production (Table1).87,88 

Fig. 2.

Molecular structures of rhTPO and PEG-rHuMGDF exhibit specific differences.

rhTPO is a glycosylated full-length TPO molecule, whereas PEG-rHuMGDF is a truncated molecule consisting of the receptor-binding portion of native TPO conjugated to a 20-kd polyethylene glycol moiety. Adapted from Begley and Basser88 with permission.

Fig. 2.

Molecular structures of rhTPO and PEG-rHuMGDF exhibit specific differences.

rhTPO is a glycosylated full-length TPO molecule, whereas PEG-rHuMGDF is a truncated molecule consisting of the receptor-binding portion of native TPO conjugated to a 20-kd polyethylene glycol moiety. Adapted from Begley and Basser88 with permission.

Table 1.

Pharmacologic characteristics of rhTPO and PEG-rHuMGDF

CharacteristicrhTPOPEG-rHuMGDF
Terminalt1/2(h) 24-40 31 
Kd for c-Mp1 (pM) 150 150  
Platelet clearance (mL/h/1091.28 1.28  
Time to onset of effect in mice (d) 3  
Time to peak effect in mice (d) 4-6 4-6  
Truncation No Yes 
Glycosylation Yes No 
Pegylation No Yes  
Human route tested Intravenous Subcutaneous 
CharacteristicrhTPOPEG-rHuMGDF
Terminalt1/2(h) 24-40 31 
Kd for c-Mp1 (pM) 150 150  
Platelet clearance (mL/h/1091.28 1.28  
Time to onset of effect in mice (d) 3  
Time to peak effect in mice (d) 4-6 4-6  
Truncation No Yes 
Glycosylation Yes No 
Pegylation No Yes  
Human route tested Intravenous Subcutaneous 

PEG-rHuMGDF, pegylated recombinant human megakaryocyte growth and development factor; rhTPO, recombinant human thrombopoietin.

In healthy animals, TPO exerts its peripheral blood effects exclusively on platelets; with no increase in white or red blood cells. Administration of either form of recombinant TPO to healthy nonhuman primates results in a dose-dependent increase in megakaryocyte number, size, and ploidy and up to a 5-fold increase in circulating platelet counts.89 There is a requisite lag time of 4 to 5 days before the platelet count rises; this reflects the finding that TPO acts primarily on early, not late, precursor cells. In murine models of severe chemotherapy- or radiation-induced thrombocytopenia or both, daily administration of recombinant TPO increases megakaryocyte numbers in the bone marrow, ameliorates the depth and duration of thrombocytopenia, and reduces the severity of leukopenia and anemia.86,90 Similar results have been observed with recombinant TPO in nonhuman primate models of chemotherapy- and radiation-induced thrombocytopenia.91-93 

In addition to these recombinant forms of TPO, several other molecules that bind and activate c-Mpl are being tested. One of these molecules is a fusion protein of TPO and IL-3. Administration of this molecule has been shown to increase platelet count in animals, but it has been found to be immunogenic and is no longer under development.94 Recently, great interest has been focused on the development of TPO peptide95 and nonpeptide96 mimetics. These mimetics are designed to bind to the TPO receptor but have no sequence homology with endogenous TPO.

Finally, a TPO receptor “potentiating” peptide has been developed that binds to c-Mpl at a location distant from the TPO binding area and prevents receptor internalization after TPO binding, thereby increasing TPO action. An analogous peptide that binds the erythropoietin receptor and potentiates erythropoietin has also recently been described.97 

Clinical studies of recombinant TPO

Nonmyeloablative treatments

The stimulatory effects of PEG-rHuMGDF and rhTPO on megakaryocyte and platelet production have been demonstrated in several clinical trials (Table 2).98-107PEG-rHuMGDF, the most widely studied recombinant TPO, has produced dose-dependent increases in platelet counts in patients with advanced malignancies and attenuated chemotherapy-induced thrombocytopenia in randomized, placebo-controlled clinical trials.98,99,105-107 When administered before chemotherapy as a daily subcutaneous injection, PEG-rHuMGDF produced a dose-dependent increase in peripheral blood platelet counts and a modest increase in megakaryocyte, erythroid, and myeloid progenitor cell levels in patients with advanced cancer.99 No evidence of platelet activation or altered platelet function was observed with PEG-rHuMGDF administration.108 

Table 2.

Clinical studies of 2 forms of recombinant thrombopoietin in chemotherapy-induced thrombocytopenia

FormInvestigatorDiseaseNo. of patientsChemotherapyDose (μg/kg)RoutePrechemotherapy phasePostchemotherapy phase
DosingOutcomeDosingOutcome
PEG-rHuMGDF Fanucchi et al98 Non-small-cell lung cancer PEG-rHuMGDF = 41 Carboplatin, paclitaxel 0.03-5.0 SC Daily up to 10 d Increase in platelets in 2 of 3 patients treated* Daily up to 16 d Increased recovery of platelets to baseline level  
   Placebo = 12       Nadir platelet counts increased 
 Basser et al99,105  Advanced cancer PEG-rHuMGDF = 31 Carboplatin, cyclophosphamide 0.03-5.0 SC Daily up to 10 d Dose-related increase in platelets
0.03-0.1 μg/kg per day 
Daily for 7 to 20 d Increased recovery of platelets to baseline level  
   Placebo = 10     → no increase in platelets
0.3-1.0 μg/kg per day 
 Platelet nadir earlier but nadir platelet count unaffected 
        → median 3-fold increase in platelets   
 Crawford et al100 Non-small-cell lung cancer PEG-rHuMGDF = 30 Carboplatin, paclitaxel 2.5-5.0 SC NA NA  Daily up to 7 d Dose-related increase in platelet nadir 
   Placebo = 10       Reduction in number of platelet transfusions required in first 2 cycles 
          Dose-limiting thrombocytopenia in later cycles 
          Anti-TPO antibodies in 2 patients  
 Basser et al106 Advanced cancer PEG-rHuMGDF = 68 Carboplatin, cisplatin 1.0-10.0 SC NA NA  Multiple daily doses up to 7 d Increased platelet nadir (47.5 × 109/L versus 35.5×109/L; P =.003) and decreased duration of grade 3 or 4 thrombocytopenia (0 versus 3 days; P =.004) with PEG-rHuMGDF  
rhTPO Vadhan-Raj et al101,102  Sarcoma rhTPO = 12 Doxorubicin, ifosfamide 0.3-2.4 IV Single Dose-related 1.3-3.6-fold increase in platelets 1, 2, 3, or 7 daily doses Decreased (schedule-dependent) thrombocytopenia Increased platelet recovery 
 Vadhan-Raj et al103,104  Gynecologic malignancy rhTPO = 29 Carboplatin 0.6-3.6 SC Single Dose-related 1.1-3.5-fold increase in platelets 4 daily doses Decrease in both degree and duration of severe thrombocytopenia and the need for platelet transfusions 
FormInvestigatorDiseaseNo. of patientsChemotherapyDose (μg/kg)RoutePrechemotherapy phasePostchemotherapy phase
DosingOutcomeDosingOutcome
PEG-rHuMGDF Fanucchi et al98 Non-small-cell lung cancer PEG-rHuMGDF = 41 Carboplatin, paclitaxel 0.03-5.0 SC Daily up to 10 d Increase in platelets in 2 of 3 patients treated* Daily up to 16 d Increased recovery of platelets to baseline level  
   Placebo = 12       Nadir platelet counts increased 
 Basser et al99,105  Advanced cancer PEG-rHuMGDF = 31 Carboplatin, cyclophosphamide 0.03-5.0 SC Daily up to 10 d Dose-related increase in platelets
0.03-0.1 μg/kg per day 
Daily for 7 to 20 d Increased recovery of platelets to baseline level  
   Placebo = 10     → no increase in platelets
0.3-1.0 μg/kg per day 
 Platelet nadir earlier but nadir platelet count unaffected 
        → median 3-fold increase in platelets   
 Crawford et al100 Non-small-cell lung cancer PEG-rHuMGDF = 30 Carboplatin, paclitaxel 2.5-5.0 SC NA NA  Daily up to 7 d Dose-related increase in platelet nadir 
   Placebo = 10       Reduction in number of platelet transfusions required in first 2 cycles 
          Dose-limiting thrombocytopenia in later cycles 
          Anti-TPO antibodies in 2 patients  
 Basser et al106 Advanced cancer PEG-rHuMGDF = 68 Carboplatin, cisplatin 1.0-10.0 SC NA NA  Multiple daily doses up to 7 d Increased platelet nadir (47.5 × 109/L versus 35.5×109/L; P =.003) and decreased duration of grade 3 or 4 thrombocytopenia (0 versus 3 days; P =.004) with PEG-rHuMGDF  
rhTPO Vadhan-Raj et al101,102  Sarcoma rhTPO = 12 Doxorubicin, ifosfamide 0.3-2.4 IV Single Dose-related 1.3-3.6-fold increase in platelets 1, 2, 3, or 7 daily doses Decreased (schedule-dependent) thrombocytopenia Increased platelet recovery 
 Vadhan-Raj et al103,104  Gynecologic malignancy rhTPO = 29 Carboplatin 0.6-3.6 SC Single Dose-related 1.1-3.5-fold increase in platelets 4 daily doses Decrease in both degree and duration of severe thrombocytopenia and the need for platelet transfusions 

NA indicates not applicable; PEG-rHuMGDF, pegylated recombinant human megakaryocyte growth and development factor; rhTPO, recombinant human thrombopoietin; SC subcutaneous; and IV, intravenous.

*

Six patients (3 placebo, 3 PEG-rHuMGDF) were evaluated in the prechemotherapy phase.

Seventeen patients (4placebo, 13 PEG-rHuMGDF) were evaluated in the prechemotherapy phase.

Subsequent trials evaluated the effects of PEG-rHuMGDF on hematologic recovery after chemotherapy. A randomized, placebo-controlled, dose-escalation study evaluated the effects of PEG-rHuMGDF after carboplatin-paclitaxel chemotherapy in 53 patients with lung cancer.98 Patients treated with PEG-rHuMGDF after chemotherapy had a higher median nadir platelet count (188 × 109/L) than did placebo-treated patients (111 × 109/L) and also showed more rapid recovery of platelet counts (14 days versus > 21 days). The need for platelet transfusions was unaffected because the chemotherapy regimen used did not frequently generate severe thrombocytopenia. In another randomized study of 41 patients with advanced cancer undergoing chemotherapy with carboplatin and cyclophosphamide, treatment with PEG-rHuMGDF enhanced platelet recovery in a dose-related manner.99 Although the platelet nadir occurred earlier in the PEG-rHuMGDF–treated group, its depth was unchanged. Similar results were observed in a dose-scheduling trial of PEG-rHuMGDF with G-CSF carried out in patients with non–small-cell lung cancer treated with carboplatin-paclitaxel.100 PEG-rHuMGDF–treated patients had a higher platelet nadir than did placebo-treated patients (89 × 109/L versus 27 × 109/L in cycle 1). Moreover, the need for transfusion was lower in the PEG-rHuMGDF group than in the placebo group (17% versus 64% in the first 2 cycles). However, in the later cycles, thrombocytopenia became dose limiting in all treatment groups.

A recent study examined the efficacy of different doses and schedules of PEG-rHuMGDF in 68 patients with advanced cancer.106Patients received 1 cycle of carboplatin and cyclophosphamide and were then randomly assigned to receive PEG-rHuMGDF or placebo after the second and subsequent cycles of carboplatin and cyclophosphamide chemotherapy. The platelet nadir was higher and the duration of grade 3 or 4 thrombocytopenia shorter when PEG-rHuMGDF was administered to patients who received the same dose of chemotherapy for at least 2 cycles. No evidence of an effect on platelet nadir was observed when PEG-rHuMGDF was given before chemotherapy. Unlike in animal chemotherapy models (in which multilineage responses are often seen), no effect of recombinant TPOs on red or white blood cell recovery has been seen in humans.

rhTPO has also produced a dose-dependent increase in platelet counts in patients with sarcomas and gynecologic malignancies.101-104,109 A phase 1 and 2 study examined the effect of rhTPO on megakaryocyte and platelet production before and after chemotherapy with doxorubicin and ifosfamide in patients with sarcomas who were at high risk of developing chemotherapy-induced thrombocytopenia. When given intravenously before chemotherapy, a single dose of rhTPO was associated with a dose-dependent increase in peripheral platelets that began on day 4 and peaked on day 12 in most patients.102This increase in platelet number was accompanied by a 4-fold increase in bone marrow megakaryocytes and a marked expansion and mobilization of erythroid, myeloid, and megakaryocyte progenitor cells. A single dose of rhTPO given intravenously after chemotherapy with doxorubicin and ifosfamide decreased the incidence of thrombocytopenia in some patients.101 A second trial investigated the clinical safety and activity of rhTPO administered subcutaneously to previously treated patients with gynecologic malignancies before and after chemotherapy with carboplatin.104 As observed in the previous study, administration of a single subcutaneous dose of rhTPO before chemotherapy produced a modest dose-dependent rise in circulating platelet counts. Administration of multiple doses of rhTPO after carboplatin chemotherapy produced an earlier platelet count nadir but effectively reduced the depth of the platelet nadir and the duration of severe thrombocytopenia (Figure3). The need for platelet transfusions decreased by 75% (Figure4).104 

Fig. 3.

rhTPO increases nadir platelet count.

In patients undergoing intensive chemotherapy for gynecologic malignancy, rhTPO (given on days 2, 4, 6, and 8 after chemotherapy) increased the nadir platelet count. The platelet nadir also occurred earlier in patients treated with rhTPO than in untreated patients. Figure provided by Pharmacia (Peapack, NJ) from data in study by Vadhan-Raj et al.104 

Fig. 3.

rhTPO increases nadir platelet count.

In patients undergoing intensive chemotherapy for gynecologic malignancy, rhTPO (given on days 2, 4, 6, and 8 after chemotherapy) increased the nadir platelet count. The platelet nadir also occurred earlier in patients treated with rhTPO than in untreated patients. Figure provided by Pharmacia (Peapack, NJ) from data in study by Vadhan-Raj et al.104 

Fig. 4.

rhTPO decreases the need for platelet transfusions.

In patients undergoing intensive chemotherapy for gynecologic malignancy, rhTPO decreased thrombocytopenia and the need for platelet transfusions. Figure provided by Pharmacia (Peapack, NJ) from data in study by Vadhan-Raj et al.104 

Fig. 4.

rhTPO decreases the need for platelet transfusions.

In patients undergoing intensive chemotherapy for gynecologic malignancy, rhTPO decreased thrombocytopenia and the need for platelet transfusions. Figure provided by Pharmacia (Peapack, NJ) from data in study by Vadhan-Raj et al.104 

Myeloablative treatments

Prolonged and severe chemotherapy-induced thrombocytopenia is a major cause of morbidity in patients receiving intensive chemotherapy for acute leukemia and those undergoing blood stem cell transplantation.110,111 In recent years, several studies have evaluated the safety and efficacy of PEG-rHuMGDF and rhTPO in the management of thrombocytopenia associated with chemotherapy for acute leukemia and stem cell transplantation (Tables3 and4).112-125 

Table 3.

Clinical studies of 2 forms of recombinant thrombopoietin as adjunct to induction/consolidation chemotherapy in patients with newly diagnosed acute myelogenous leukemia

FormInvestigatorNo. of patientsChemotherapyDose (μg/kg)RouteInductionConsolidation
DosingOutcomeDosingOutcome
PEG-rHuMGDF Archimbaud et al112 PEG-rHuMGDF = 38 Daunorubicin, cytarabine, etoposide 2.5 or 5.0 SC Multiple or single Dose-related increase in median platelet count Multiple or single Dose-related increase in remission median platelet count  
  Placebo = 12     No difference between groups in time to platelet recovery or platelet transfusion requirements  No difference between groups in time to platelet recovery or platelet transfusion requirements  
 Schiffer et al114 PEG-rHuMGDF = 24 Daunorubicin, cytarabine +/− high-dose cytarabine 2.5 or 5.0 SC Multiple No difference between groups in time to platelet count ≥ 20 × 109/L or platelet transfusion requirements Multiple No difference between groups in time to platelet recovery to ≥ 20 × 109/L and ≥ 50 × 109/L or platelet transfusion requirements 
  Placebo = 11        
rhTPO Cripe et al115 rhTPO = 28 Idarubicin, cytarabine 2.5-5.0 IV Multiple No difference between rhTPO and historical controls in time to platelet count ≥ 20 × 109/L or ≥ 50 × 109/L NA NA 
FormInvestigatorNo. of patientsChemotherapyDose (μg/kg)RouteInductionConsolidation
DosingOutcomeDosingOutcome
PEG-rHuMGDF Archimbaud et al112 PEG-rHuMGDF = 38 Daunorubicin, cytarabine, etoposide 2.5 or 5.0 SC Multiple or single Dose-related increase in median platelet count Multiple or single Dose-related increase in remission median platelet count  
  Placebo = 12     No difference between groups in time to platelet recovery or platelet transfusion requirements  No difference between groups in time to platelet recovery or platelet transfusion requirements  
 Schiffer et al114 PEG-rHuMGDF = 24 Daunorubicin, cytarabine +/− high-dose cytarabine 2.5 or 5.0 SC Multiple No difference between groups in time to platelet count ≥ 20 × 109/L or platelet transfusion requirements Multiple No difference between groups in time to platelet recovery to ≥ 20 × 109/L and ≥ 50 × 109/L or platelet transfusion requirements 
  Placebo = 11        
rhTPO Cripe et al115 rhTPO = 28 Idarubicin, cytarabine 2.5-5.0 IV Multiple No difference between rhTPO and historical controls in time to platelet count ≥ 20 × 109/L or ≥ 50 × 109/L NA NA 

NA indicates not applicable; PEG-rHuMGDF, pegylated recombinant human megakaryocyte growth and development factor; rhTPO, recombinant human thrombopoietin; SC, subcutaneous; and IV, intravenous.

Table 4.

Clinical studies of 2 forms of recombinant thrombopoietin in patients receiving myeloablative chemotherapy followed by stem cell transplantation

FormInvestigatorNo. of patientsChemotherapyDose (μg/kg)RouteStem cell mobilizationStem cell engraftment
DosingOutcomeDosingOutcome
PEG-rHuMGDF Glaspy et al116 PEG-rHuMGDF = 39 STAMP (high-dose) 1, 3, 10, or 30 SC NA NA Multiple Dose-related increase in median platelet count with PEG-rHuMGDF  
  Placebo = 14       No difference between groups in time to platelet recovery to ≥ 20 × 109/L or platelet transfusion requirements  
 Bolwell et al117 PEG-rHuMGDF = 36 STAMP I 1.0, 2.5, 5.0, 7.5, or 10.0 SC NA NA Multiple (until platelet recovery ≥ 50 × 109/L) Dose-related increase in platelet counts with PEG-rHuMGDF as compared with placebo 
  Placebo = 11       No difference between groups in time to platelet recovery to ≥ 20 × 109/L or platelet transfusion requirements 
 Beveridge et al118 PEG-rHuMGDF = 40

Placebo = 10 
STAMP V 5 or 10.0 SC NA NA Multiple (until platelet recovery ≥ 25 × 109/L) No difference between groups in time to platelet recovery, severe thrombocytopenia, or platelet transfusion requirements  
rhTPO Nash et al120 rhTPO = 38 with delayed platelet recovery after stem cell transplantation NA 0.6-2.4 IV NA NA Single and multiple until platelet recovery ≥ 20 × 109/L No effect on time to platelet recovery to ≥ 20 × 109/L or platelet transfusion requirements  
 Somlo et al121 rhTPO plus G-CSF = 26
G-CSF alone = 20 
Cisplatin, etoposide, cyclophosphamide 0.6-2.4 IV Single and multiple Increased CD34+ cell yields with rhTPO plus G-CSF as compared with G-CSF alone NA Reduced time to platelet recovery and platelet transfusion requirements with rhTPO plus G-CSF-mobilized cells  
 Linker et al123 rhTPO plus G-CSF = 101

G-CSF alone = 28 
 1.5 IV Single or multiple Increased CD34+ cell number per apheresis and number of patients reaching minimum graft (CD34+ > 2 × 106/kg) with rhTPO plus G-CSF as compared with G-CSF alone Single or multiple No effect on time to platelet recovery to ≥ 20 × 109/L or platelet transfusion requirements 
FormInvestigatorNo. of patientsChemotherapyDose (μg/kg)RouteStem cell mobilizationStem cell engraftment
DosingOutcomeDosingOutcome
PEG-rHuMGDF Glaspy et al116 PEG-rHuMGDF = 39 STAMP (high-dose) 1, 3, 10, or 30 SC NA NA Multiple Dose-related increase in median platelet count with PEG-rHuMGDF  
  Placebo = 14       No difference between groups in time to platelet recovery to ≥ 20 × 109/L or platelet transfusion requirements  
 Bolwell et al117 PEG-rHuMGDF = 36 STAMP I 1.0, 2.5, 5.0, 7.5, or 10.0 SC NA NA Multiple (until platelet recovery ≥ 50 × 109/L) Dose-related increase in platelet counts with PEG-rHuMGDF as compared with placebo 
  Placebo = 11       No difference between groups in time to platelet recovery to ≥ 20 × 109/L or platelet transfusion requirements 
 Beveridge et al118 PEG-rHuMGDF = 40

Placebo = 10 
STAMP V 5 or 10.0 SC NA NA Multiple (until platelet recovery ≥ 25 × 109/L) No difference between groups in time to platelet recovery, severe thrombocytopenia, or platelet transfusion requirements  
rhTPO Nash et al120 rhTPO = 38 with delayed platelet recovery after stem cell transplantation NA 0.6-2.4 IV NA NA Single and multiple until platelet recovery ≥ 20 × 109/L No effect on time to platelet recovery to ≥ 20 × 109/L or platelet transfusion requirements  
 Somlo et al121 rhTPO plus G-CSF = 26
G-CSF alone = 20 
Cisplatin, etoposide, cyclophosphamide 0.6-2.4 IV Single and multiple Increased CD34+ cell yields with rhTPO plus G-CSF as compared with G-CSF alone NA Reduced time to platelet recovery and platelet transfusion requirements with rhTPO plus G-CSF-mobilized cells  
 Linker et al123 rhTPO plus G-CSF = 101

G-CSF alone = 28 
 1.5 IV Single or multiple Increased CD34+ cell number per apheresis and number of patients reaching minimum graft (CD34+ > 2 × 106/kg) with rhTPO plus G-CSF as compared with G-CSF alone Single or multiple No effect on time to platelet recovery to ≥ 20 × 109/L or platelet transfusion requirements 

G-CSF indicates granulocyte colony-stimulating factor; NA, not applicable; PEG-rHuMGDF, pegylated recombinant human megakaryocyte growth and development factor; rhTPO, recombinant human thrombopoietin; STAMP, solid tumor autologous marrow program.

In contrast to their effect in the nonmyeloablative setting, PEG-rHuMGDF and rhTPO have not had a clinically significant effect on platelet production when administered to patients receiving dose-intensive therapy for acute leukemia and those undergoing stem cell transplantation after chemotherapy. Moderate increase in peak platelet counts and reduction in time to full platelet recovery were often achieved in patients treated with PEG-rHuMGDF and rhTPO. However, no improvement in time to recovery to a platelet count more than or equal to 20 × 109/L and no reduction in the need for platelet transfusions were observed in these studies.112-114,119,120 

In preclinical studies, treatment with TPO before bone marrow harvesting accelerated platelet reconstitution in recipient mice after transplantation, suggesting that this approach may be effective in shortening the time to platelet independence after stem cell transplantation.126 With this approach, one study showed that administration of rhTPO to patients during mobilization of peripheral blood progenitor cells increased CD34+ yield before stem cell transplantation. Small, statistically significant improvements in neutrophil recovery and platelet and erythrocyte transfusion requirements were noted after transplantation.121 

MDS

Hematopoietic growth factors have had some success in ameliorating the neutropenia and anemia associated with myelodysplastic syndrome (MDS). The recombinant TPOs may have a similar benefit: some in vitro studies have shown that bone marrow cells of patients with MDS can differentiate into the megakaryocytic lineage when exposed to recombinant TPO.127,128 Because of the underlying heterogeneity of MDS, some individuals might have responsive marrow whereas others might not. Endogenous TPO levels are normal to slightly elevated in MDS,129 so whether they can help predict responsiveness to exogenous TPO requires further investigation. In a preliminary report, various intravenous doses of PEG-rHuMGDF were given daily for 14 days to 21 Japanese patients with MDS (refractory anemia and refractory anemia with ringed sideroblasts) with platelet counts less than 30 × 109/L. The peak effect of PEG-rHuMGDF occurred 5 to 6 weeks later with an average doubling of platelet count; responses were seen in a third of the patients, and a multilineage effect was observed in a few patients.130 

Human immunodeficiency virus

Several studies have examined thrombocytopenia in patients or primates infected with human immunodeficiency virus (HIV) with respect to peripheral platelet mass turnover, marrow megakaryocytopoiesis, and endogenous TPO levels.3,131,132 A 10-fold disparity between the reduced platelet production and expanded megakaryocyte mass was observed in the bone marrow of HIV patients with thrombocytopenia.132 This suggests that, despite the expanded megakaryocyte mass, HIV-infected megakaryocytes have a high rate of apoptosis and resultant ineffective thrombopoiesis and thrombocytopenia. Harker and colleagues131 showed that, in thrombocytopenic chimpanzees infected with HIV, administration of PEG-rHuMGDF rapidly eliminated thrombocytopenia. With normal or slightly elevated endogenous TPO levels in 6 HIV-infected patients, platelet counts in all patients increased 10-fold within 14 days of the start of PEG-rHuMGDF treatment.132 This increase was not associated with change in the megakaryocyte mass, platelet life span, or viral load. What appeared to occur was an increase in the rate of effective platelet production from the bone marrow megakaryocytes of these individuals. These data suggest that, in HIV-related immune thrombocytopenic purpura, TPO can be expected to produce clinically beneficial increases in platelet counts.

A similar response has been recently seen in 3 of 4 Japanese patients with non–HIV-related ITP treated with intravenous PEG-rHuMGDF.133 One patient with ITP has been successfully treated twice weekly with subcutaneous PEG-rHuMGDF for over 3 years.134 

Liver disease

Recent understanding of TPO biology suggests that reduced hepatic production of TPO may play a major role in thrombocytopenia associated with liver disease. TPO is produced primarily in the liver, and thrombocytopenia in animals seems to be proportional to the extent of liver resection.135 In addition, after transplantation of healthy livers into TPO−/− mice, platelet counts returned toward normal, suggesting that the majority of TPO is produced in the liver.59 An association between low platelet counts (median, 84 × 109/L; range, 26 × 109/L-112 × 109/L) and low levels of TPO (median, < 20 pg/mL; range, < 20-182 pg/mL) has been reported in patients before orthotopic liver transplantation.60,61Within 4 days of orthotopic liver transplantation, TPO levels rose above normal and were accompanied by increased amounts of reticulated platelets. At 14 days after transplantation, platelet counts were normal in 14 of 18 patients (median, 254 × 109/L; range, 70 × 109/L-398 × 109/L) and TPO levels returned to normal in 14 of 18 patients (median, 59 pg/mL; range, < 20-639 pg/mL). No appreciable change in spleen size was observed. In multivariate analysis, the increase in TPO was the only variable that correlated with the increase in platelet count. Thus, TPO could potentially be used to reduce hemorrhage in patients with thrombocytopenia due to liver disease or to prepare such patients for liver transplantation.

Surgery

Approximately 40% of all platelet transfusions are used in surgical settings.136 Preoperative and postoperative thrombocytopenia complicates surgical procedures and mandates platelet transfusions. No clinical studies have targeted this important area. However, in dogs, the administration of PEG-rHuMGDF 4 days before surgery decreased thrombocytopenia after cardiopulmonary bypass.137 Despite its 5-day lag time before platelet rise, judicial administration of TPO before surgery may ameliorate preoperative and postoperative thrombocytopenia and reduce the need for platelet transfusions.

Transfusion medicine

The striking effect of TPO on the mobilization of CD34+ cells and expansion of multilineage stem cell progenitor pools in vivo led to an evaluation of its activity in 3 areas: mobilization of peripheral blood stem cells before stem cell transplantation, ex vivo expansion of pluripotent stem cells from umbilical cord blood or bone marrow, and increase in the yield of platelet apheresis from healthy donors.

Stem cell mobilization.

Recently, several pilot studies evaluated the activity of various doses and schedules of rhTPO or PEG-rHuMGDF in combination with G-CSF and chemotherapy as part of a mobilization regimen for stem cell transplantation (Table 4).99,121-123,138,139 In contrast to peak progenitor cell numbers on days 5 to 7 usually obtained with G-CSF alone, a peak on days 12 to 15 was produced by the combination of PEG-rHuMGDF and G-CSF. However, since a full pharmacodynamic response profile to PEG-rHuMGDF was not performed in this study, the exact day of peak stem cell mobilization was not determined. The addition of rhTPO to G-CSF for chemotherapy mobilization regimens substantially increased CD34+ yields (Figure5). The promising results observed in these early studies were recently confirmed in a large randomized phase 2 study of rhTPO in patients undergoing high-dose chemotherapy and transplantation of peripheral blood stem cells (Table4).121 Treatment with rhTPO in various doses and schedules reduced the number of aphereses needed to reach a target graft (ie, CD34+ > 5 × 106/kg) and, compared with placebo treatment, increased the percentage of patients reaching a target graft (from 46% in the placebo group to 79% in the rhTPO group), as well as the percentage of patients reaching the minimum target graft (ie, CD34+ > 2 × 106/kg) (from 75% in the placebo group to 94% in the rhTPO group). These studies demonstrate the ability of rhTPO to mobilize CD34+cells safely and effectively and increase the harvest of CD34+ cells used for stem cell transplantation.

Fig. 5.

PEG-rHuMGDF increases peripheral blood progenitor cells (PBPCs).

Patients undergoing PBPC transplantation underwent stem cell mobilization with chemotherapy and G-CSF with or without PEG-rHuMGDF. Use of PEG-rHuMGDF increased granulocyte-macrophage colony-forming cells (GM-CFCs), megakaryocyte colony-forming cells (Meg-CFCs), erythroid burst-forming units (BFU-Es), and CD34+ cells. Adapted from Basser et al99 with permission.

Fig. 5.

PEG-rHuMGDF increases peripheral blood progenitor cells (PBPCs).

Patients undergoing PBPC transplantation underwent stem cell mobilization with chemotherapy and G-CSF with or without PEG-rHuMGDF. Use of PEG-rHuMGDF increased granulocyte-macrophage colony-forming cells (GM-CFCs), megakaryocyte colony-forming cells (Meg-CFCs), erythroid burst-forming units (BFU-Es), and CD34+ cells. Adapted from Basser et al99 with permission.

Ex vivo expansion of primitive stem cells.

The role of TPO in the expansion and prolonged survival of primitive stem cells derived from bone marrow or umbilical cord blood has been the focus of several recent investigations. Yagi and colleagues140 demonstrated that administration of TPO alone can sustain ex vivo expansion of hematopoietic stem cells in long-term bone marrow cultures (LTBMCs) from mice (Figure6). The continuous presence of TPO resulted in the generation of long- and short-term colony-forming cells and maintained the relative amount of high-proliferative-potential colony-forming cells (Figure 7). Most important, competitive repopulation studies found that the TPO-treated LTBMC cells were as effective as fresh marrow. Subsequent data from this research group suggest that the expanded population of stem cells, when transplanted into recipient mice, is adequate for the long-term repopulation.

Fig. 6.

TPO promotes ex vivo stem cell expansion.

During a 5-month, long-term murine bone marrow culture, the total number of nonadherent (NA) cells increased in the presence of TPO. Reprinted from Yagi et al140 with permission.

Fig. 6.

TPO promotes ex vivo stem cell expansion.

During a 5-month, long-term murine bone marrow culture, the total number of nonadherent (NA) cells increased in the presence of TPO. Reprinted from Yagi et al140 with permission.

Fig. 7.

Colony-forming cells (CFCs) and high-proliferative-potential (HPP) CFCs are maintained during ex vivo stem cell expansion.

The total number of CFCs (A) and HPP CFCs (B) were measured in long-term bone marrow cultures. Totals were compared between cultures grown in the presence (TPO) or absence (control) of TPO and uncultured (fresh) bone marrow. Reprinted from Yagi et al140 with permission.

Fig. 7.

Colony-forming cells (CFCs) and high-proliferative-potential (HPP) CFCs are maintained during ex vivo stem cell expansion.

The total number of CFCs (A) and HPP CFCs (B) were measured in long-term bone marrow cultures. Totals were compared between cultures grown in the presence (TPO) or absence (control) of TPO and uncultured (fresh) bone marrow. Reprinted from Yagi et al140 with permission.

Piacibello and colleagues141 showed that the use of growth factors could expand human cord blood CD34+ cells ex vivo by many million-fold in total number; the CD34+ component and the lineage-specific progenitors increased proportionately. Although TPO alone and Flt 3 ligand alone were insufficient in stimulating sustained growth, a combination of the 2 growth factors accounted for this rapid increase in cell number during 24 weeks in culture. However, whether the expanded cell population can be used clinically for transplantation has not been demonstrated.

Platelet apheresis.

Extensive studies have shown that healthy apheresis donors maximally increase their platelet count 10 to 14 days after a single injection of PEG-rHuMGDF.136,142,143 The rise in platelet count is dose dependent and leads directly to an increase in the apheresis platelet yield (Figure 8). The platelets collected have normal aggregation responses and normal function on transfusion into thrombocytopenic recipients. Transfusions with the higher platelet doses extended the duration of transfusion independence and possibly reduced bleeding episodes when compared with standard doses. The corrected count increment was also improved when patients were transfused with platelets mobilized by PEG-rHuMGDF rather than with those harvested from control donors.

Fig. 8.

PEG-rHuMGDF increases the yield of platelet apheresis.

Administration of a single dose of PEG-rHuMGDF produced a dose-dependent increase in platelet count and apheresis yield 15 days later. Reprinted from Kuter et al142 with permission.

Fig. 8.

PEG-rHuMGDF increases the yield of platelet apheresis.

Administration of a single dose of PEG-rHuMGDF produced a dose-dependent increase in platelet count and apheresis yield 15 days later. Reprinted from Kuter et al142 with permission.

Radioprotection

Although there are no studies in humans, the potential for TPO to function as a radioprotectant is another area of clinical interest. In mice, administration of TPO 2 hours after exposure to sublethal total body irradiation (TBI) dramatically ameliorates the thrombocytopenia that is seen at day 10 in these mice.144 Mice treated with rhTPO before irradiation have a higher platelet count nadir (739 × 109/L) than do those that are untreated before irradiation (144 × 109/L); unirradiated control mice had a platelet count of 1123 × 109/L. This protective effect was enhanced when rhTPO was administered close to TBI. Stem cells appeared to be highly sensitive to the effects of rhTPO, possibly because it prevented apoptosis when given from 2 hours before until 2 hours after TBI. This very narrow window of protection underscores the importance of the timing of the administration of rhTPO vis-à-vis the irradiation. Furthermore, red and white blood cell counts also appeared to be protected somewhat by the administration of rhTPO close to the time of TBI (Figure9).144 These results suggest a major radioprotective effect of rhTPO on progenitor cells in the bone marrow. This finding is in line with previous data suggesting that pluripotential stem cells are sensitive to the presence of TPO and that TPO can support their survival.145,146 Similar results have been noted in subsequent studies that investigated the effect of rhTPO in mice exposed to lethal doses of TBI.147 Almost all the mice that received rhTPO close to the time of irradiation survived, whereas all the mice that received placebo died within 30 days of receiving TBI. Furthermore, recovery of blood counts in all lineages improved in those mice that received rhTPO within several hours of TBI.

Fig. 9.

TPO improves regeneration of bone marrow progenitors.

Mice receiving TBI with 6 Gy were treated with TPO 2 hours and 24 hours after irradiation. After 7 days, GM-CFUs (GM), BFU-Es (E), and Meg-CFCs (Meg) were measured per femur. Regeneration of bone marrow progenitors improved with the 2-hour but not the 24-hour post-TBI administration of TPO. Reprinted from Neelis et al144 with permission.

Fig. 9.

TPO improves regeneration of bone marrow progenitors.

Mice receiving TBI with 6 Gy were treated with TPO 2 hours and 24 hours after irradiation. After 7 days, GM-CFUs (GM), BFU-Es (E), and Meg-CFCs (Meg) were measured per femur. Regeneration of bone marrow progenitors improved with the 2-hour but not the 24-hour post-TBI administration of TPO. Reprinted from Neelis et al144 with permission.

Whether these results can be expanded to the chemotherapy setting has not been fully explored. Conceivably the antiapoptotic effects of TPO might lessen chemotherapy-induced apoptosis of pluripotential stem cells and thereby ameliorate the pancytopenia of chemotherapy.

Safety of recombinant TPO

In most of the studies with TPO in myeloablative and nonmyeloablative chemotherapy, patients also received myeloid growth factors. Although interactions of TPO with myeloid growth factors had been seen in one animal model,148 none have been noted an any clinical study.

Administration of multiple doses of PEG-rHuMGDF to some cancer patients and healthy volunteers was associated with an abrogation of its pharmacologic effect as a result of the development of neutralizing antibodies.85,100,149,150 These antibodies neutralized both the recombinant and endogenous TPO, resulting in thrombocytopenia. Thrombocytopenia occurred in 4 of 665 cancer/stem cell transplantation/leukemia patients given multiple doses and in 2 (1.2%) of 210 healthy volunteers who received 2 doses and in 11 (8.9%) of 124 healthy volunteers given 3 doses of PEG-rHuMGDF.85,149 No subject developed neutralizing antibodies or thrombocytopenia after a single injection. Evaluation of these thrombocytopenic subjects showed that the thrombocytopenia was due to the formation of an IgG antibody to PEG-rHuMGDF that cross-reacted with endogenous TPO and neutralized its biologic activity.85,149,150 In 2 patients, thrombocytopenia was associated with anemia and neutropenia, suggesting an effect on a stem cell population.85,150 Because endogenous TPO is produced in a constitutive fashion by the liver, thrombocytopenia ensues. PEG-rHuMGDF was withdrawn from clinical trials in the United States by Amgen in September 1998 because of this side effect.151 

A possible explanation for the immunogenicity of PEG-rHuMGDF administration may be that this molecule is truncated, nonglycosylated, and pegylated, in contrast to the full-length, glycosylated rhTPO molecule (Table 1). In addition, PEG-rHuMGDF has usually been administered subcutaneously, whereas full-length native rhTPO has been injected intravenously. Because TPO is a potent mobilizer of dendritic cells, injection of any form of TPO subcutaneously might enhance its immunogenicity. Support for this latter hypothesis comes from recent experiments in which PEG-ratMGDF was injected into rats once monthly for 3 months by either a subcutaneous or intravenous route. Most animals treated subcutaneously developed neutralizing antibodies and thrombocytopenia whereas those treated intravenously did not (personal communication, Dr A. Shimosaka, April 2002).

To date, the development of neutralizing antibodies in patients treated with intravenous rhTPO has not been reported, although one nonneutralizing antibody was found after subcutaneous injection of rhTPO.102,104 

Summary

The development of recombinant TPO has led to a wide number of discoveries describing the underlying biology of platelet production in normal and pathologic settings. Recombinant TPO has demonstrated a unique pharmacology, unlike other hematopoietic growth factors. Both rhTPO and PEG-rHuMGDF have a prolonged half-life of about 40 hours, and thus continuous dosing with TPO does not seem to be required; one or more appropriately timed doses may even be superior to multiple doses. TPO does not increase the platelet count for 5 days and has its peak effect 10 to 12 days later. It has little effect on mature megakaryocytes and may actually inhibit their shedding of platelets.

TPO is the most specific and effective growth factor identified to date for the prevention and treatment of thrombocytopenia. Preliminary clinical evidence indicates that TPO administration may be a helpful adjunct to the conventional approach of platelet transfusion therapy for some cancer patients with chemotherapy-induced thrombocytopenia. However, the studies with TPO, as with IL-11, mostly involved nonconventional chemotherapy regimens that caused considerable thrombocytopenia. For most routine chemotherapy regimens, clinically significant thrombocytopenia is relatively uncommon. The overall impact of TPO on the need for platelet transfusions will probably not be great, especially with the recent reduction in the threshold “trigger” for platelet transfusions to 10 × 109/L.152-154 

The failure to find any biologic effect in myeloablative regimens is still surprising given the success of the myeloid growth factors in these same settings. This is probably not simply due to inadequate dosing schemes; many have been tried. Rather it may reflect aspects of the clinical biology of TPO that are not yet recognized. The elevated endogenous TPO concentration in all of these settings may have already saturated the TPO receptor or, alternatively, may even prevent platelet shedding.155 

The ultimate clinical indications for recombinant TPO or TPO mimetics will certainly depend on the results of continuing and future studies. Further studies to elucidate their complex and unique biology will help to determine their optimal application in the treatment of thrombocytopenia. While the potential of TPO to reduce the extent of chemotherapy-induced thrombocytopenia and reduce the need for platelet transfusions in the nonmyeloablative chemotherapy setting may be enhanced with innovative dosing schemes, TPO will probably have its greatest impact in nononcology settings such as the stimulation of directed platelet apheresis donors. The thrombocytopenia associated with HIV infection, ITP, MDS, liver disease, radiation, surgery, and intensive care patients accounts for over half of all platelet transfusions136 and remains in need of an effective thrombopoietic growth factor.

Supported in part by grants from the National Institutes of Health HL54838 and HL61272 (D.J.K.), the National Institutes of Health, Medical Research Council, Canberra, Australia; Amgen Inc, Thousand Oaks, CA (C.G.B.); and Pharmacia Inc, Peapack, NJ (D.J.K., C.G.B.).

D.J.K. has been a consultant for and received research support from Amgen Inc, Kirin Inc, and Pharmacia Inc. C.G.B. was a consultant to Amgen Inc during the course of the clinical studies with PEG-rHuMGDF.

References

References
1
Kaushansky
K
The thrombocytopenia of cancer: prospects for effective cytokine therapy.
Hematol Oncol Clin North Am.
10
1996
431
455
2
Prow
D
Vadhan-Raj
S
Thrombopoietin: biology and potential clinical applications.
Oncology (Huntingt).
12
1998
1597
1604
1607-1598; discussion 1611-1594.
3
Ballem
PJ
Belzberg
A
Devine
DV
et al
Kinetic studies of the mechanism of thrombocytopenia in patients with human immunodeficiency virus infection.
N Engl J Med.
327
1992
1779
1784
4
Mittelman
M
Zeidman
A
Platelet function in the myelodysplastic syndromes.
Int J Hematol.
71
2000
95
98
5
Lazarus
AH
Ellis
J
Semple
JW
Mody
M
Crow
AR
Freedman
J
Comparison of platelet immunity in patients with SLE and with ITP.
Transfus Sci.
22
2000
19
27
6
Lawrence
SP
Lezotte
DC
Durham
JD
Kumpe
DA
Everson
GT
Bilir
BM
Course of thrombocytopenia of chronic liver disease after transjugular intrahepatic portosystemic shunts (TIPS): a retrospective analysis.
Dig Dis Sci.
40
1995
1575
1580
7
Alvarez
JM
Gates
R
Rowe
D
Brady
PW
Complications from intra-aortic balloon counterpulsation: a review of 303 cardiac surgical patients.
Eur J Cardiothorac Surg.
6
1992
530
535
8
Vonderheide
RH
Thadhani
R
Kuter
DJ
Association of thrombocytopenia with the use of intra-aortic balloon pumps.
Am J Med.
105
1998
27
32
9
Chiu
EK
Yuen
KY
Lie
AK
et al
A prospective study of symptomatic bacteremia following platelet transfusion and of its management.
Transfusion.
34
1994
950
954
10
Davda
RK
Collins
KA
Kitchens
CS
Case report: fatal Staphylococcus aureus sepsis from single-donor platelet transfusion.
Am J Med Sci.
307
1994
340
341
11
Chambers
LA
Kruskall
MS
Pacini
DG
Donovan
LM
Febrile reactions after platelet transfusion: the effect of single versus multiple donors.
Transfusion.
30
1990
219
221
12
Contreras
M
Diagnosis and treatment of patients refractory to platelet transfusions.
Blood Rev.
12
1998
215
221
13
Engelfriet
CP
Reesink
HW
Aster
RH
et al
Management of alloimmunized, refractory patients in need of platelet transfusions.
Vox Sang.
73
1997
191
198
14
Friedberg
RC
Mintz
PD
Causes of refractoriness to platelet transfusion.
Curr Opin Hematol.
2
1995
493
498
15
Novotny
VM
Prevention and management of platelet transfusion refractoriness.
Vox Sang.
76
1999
1
13
16
Kimura
H
Ishibashi
T
Shikama
Y
et al
Interleukin-1 beta (IL-1 beta) induces thrombocytosis in mice: possible implication of IL-6.
Blood.
76
1990
2493
2500
17
Bartley
TD
Bogenberger
J
Hunt
P
et al
Identification and cloning of a megakaryocyte growth and development factor that is a ligand for the cytokine receptor Mpl.
Cell.
77
1994
1117
1124
18
Lok
S
Kaushansky
K
Holly
RD
et al
Cloning and expression of murine thrombopoietin cDNA and stimulation of platelet production in vivo.
Nature.
369
1994
565
568
19
de Sauvage
FJ
Hass
PE
Spencer
SD
et al
Stimulation of megakaryocytopoiesis and thrombopoiesis by the c-Mpl ligand.
Nature.
369
1994
533
538
20
Kuter
DJ
Beeler
DL
Rosenberg
RD
The purification of megapoietin: a physiological regulator of megakaryocyte growth and platelet production.
Proc Natl Acad Sci U S A.
91
1994
11104
11108
21
Kato
T
Ogami
K
Shimada
Y
et al
Purification and characterization of thrombopoietin.
J Biochem.
118
1995
229
236
22
Metcalf
D
Burgess
AW
Johnson
GR
et al
In vitro actions on hemopoietic cells of recombinant murine GM-CSF purified after production in Escherichia coli: comparison with purified native GM-CSF.
J Cell Physiol.
128
1986
421
431
23
Debili
N
Masse
JM
Katz
A
Guichard
J
Breton-Gorius
J
Vainchenker
W
Effects of the recombinant hematopoietic growth factors interleukin-3, interleukin-6, stem cell factor, and leukemia inhibitory factor on the megakaryocytic differentiation of CD34+ cells.
Blood.
82
1993
84
95
24
Leonard
JP
Quinto
CM
Kozitza
MK
Neben
TY
Goldman
SJ
Recombinant human interleukin-11 stimulates multilineage hematopoietic recovery in mice after a myelosuppressive regimen of sublethal irradiation and carboplatin.
Blood.
83
1994
1499
1506
25
Metcalf
D
Begley
CG
Williamson
DJ
et al
Hemopoietic responses in mice injected with purified recombinant murine GM-CSF.
Exp Hematol.
15
1987
1
9
26
Metcalf
D
Begley
CG
Johnson
GR
Nicola
NA
Lopez
AF
Williamson
DJ
Effects of purified bacterially synthesized murine multi-CSF (IL-3) on hematopoiesis in normal adult mice.
Blood.
68
1986
46
57
27
Vadhan-Raj
S
Kudelka
AP
Garrison
L
et al
Effects of interleukin-1 alpha on carboplatin-induced thrombocytopenia in patients with recurrent ovarian cancer.
J Clin Oncol.
12
1994
707
714
28
Leonardi
V
Danova
M
Fincato
G
Palmeri
S
Interleukin 3 in the treatment of chemotherapy induced thrombocytopenia.
Oncol Rep.
5
1998
1459
1464
29
D'Hondt
V
Humblet
Y
Guillaume
T
et al
Thrombopoietic effects and toxicity of interleukin-6 in patients with ovarian cancer before and after chemotherapy: a multicentric placebo-controlled, randomized phase Ib study.
Blood.
85
1995
2347
2353
30
Gordon
MS
McCaskill-Stevens
WJ
Battiato
LA
et al
A phase I trial of recombinant human interleukin-11 (neumega rhIL-11 growth factor) in women with breast cancer receiving chemotherapy.
Blood.
87
1996
3615
3624
31
Tepler
I
Elias
L
Smith
JW
et al
A randomized placebo-controlled trial of recombinant human interleukin-11 in cancer patients with severe thrombocytopenia due to chemotherapy.
Blood.
87
1996
3607
3614
32
Smith
JWD
Longo
DL
Alvord
WG
et al
The effects of treatment with interleukin-1 alpha on platelet recovery after high-dose carboplatin.
N Engl J Med.
328
1993
756
761
33
Vredenburgh
JJ
Hussein
A
Fisher
D
et al
A randomized trial of recombinant human interleukin-11 following autologous bone marrow transplantation with peripheral blood progenitor cell support in patients with breast cancer.
Biol Blood Marrow Transplant.
4
1998
134
141
34
Nandurkar
HH
Robb
L
Tarlinton
D
Barnett
L
Kontgen
F
Begley
GC
Adult mice with targeted mutation of the interleukin-11 receptor (IL11Ra) display normal hematopoiesis.
Blood.
90
1997
2148
2159
35
Robb
L
Li
R
Hartley
L
Nandurkar
HH
Koentgen
F
Begley
CG
Infertility in female mice lacking the receptor for interleukin 11 is due to a defective uterine response to implantation.
Nat Med.
4
1998
303
308
36
Gordon
MS
Nemunaitis
J
Hoffman
R
et al
A phase I trial of recombinant human interleukin-6 in patients with myelodysplastic syndromes and thrombocytopenia.
Blood.
85
1995
3066
3076
37
Lazarus
HM
Winton
EF
Williams
SF
et al
Phase I multicenter trial of interleukin 6 therapy after autologous bone marrow transplantation in advanced breast cancer.
Bone Marrow Transplant.
15
1995
935
942
38
Nieken
J
Mulder
NH
Buter
J
et al
Recombinant human interleukin-6 induces a rapid and reversible anemia in cancer patients.
Blood.
86
1995
900
905
39
Carver-Moore
K
Broxmeyer
HE
Luoh
SM
et al
Low levels of erythroid and myeloid progenitors in thrombopoietin- and c-mpl-deficient mice.
Blood.
88
1996
803
808
40
Gainsford
T
Roberts
AW
Kimura
S
et al
Cytokine production and function in c-mpl-deficient mice: no physiologic role for interleukin-3 in residual megakaryocyte and platelet production.
Blood.
91
1998
2745
2752
41
Gainsford
T
Nandurkar
H
Metcalf
D
Robb
L
Begley
CG
Alexander
WS
The residual megakaryocyte and platelet production in c-mpl-deficient mice is not dependent on the actions of interleukin-6, interleukin-11, or leukemia inhibitory factor.
Blood.
95
2000
528
534
42
Kelemen
E
Cserhati
I
Tanos
B
Demonstration and some properties of human thrombopoietin in thrombocythaemic sera.
Acta Haematol.
20
1958
350
355
43
Wendling
F
Tambourin
P
The oncogene V-MPL, a putative truncated cytokine receptor which immortalized hemtopoietic progenitors.
Nouv Rev Fr Hematol.
33
1991
145
146
44
Vigon
I
Mornon
JP
Cocault
L
et al
Molecular cloning and characterization of MPL, the human homolog of the v-mpl oncogene: identification of a member of the hematopoietic growth factor receptor superfamily.
Proc Natl Acad Sci U S A.
89
1992
5640
5644
45
Methia
N
Louache
F
Vainchenker
W
Wendling
F
Oligodeoxynucleotides antisense to the proto-oncogene c-mpl specifically inhibit in vitro megakaryocytopoiesis.
Blood.
82
1993
1395
1401
46
Sohma
Y
Akahori
H
Seki
N
et al
Molecular cloning and chromosomal localization of the human thrombopoietin gene.
FEBS Letters.
353
1994
57
61
47
Foster
D
Hunt
P
The biological significance of truncated and full-length forms of Mpl ligand.
Thrombopoiesis and Thrombopoietins: Molecular, Cellular, Preclinical, and Clinical Biology.
Kuter
DJ
Hunt
P
Sheridan
W
Zucker-Franklin
D
1997
203
214
Humana Press
Totowa, NJ
48
Foster
DC
Sprecher
CA
Grant
FJ
et al
Human thrombopoietin: gene structure, cDNA sequence, expression, and chromosomal localization.
Proc Natl Acad Sci U S A.
91
1994
13023
13027
49
Yang
C
Li
YC
Kuter
DJ
The physiological response of thrombopoietin (c-Mpl ligand) to thrombocytopenia in the rat.
Br J Haematol.
105
1999
478
485
50
Stoffel
R
Wiestner
A
Skoda
RC
Thrombopoietin in thrombocytopenic mice: evidence against regulation at the mRNA level and for a direct regulatory role of platelets.
Blood.
87
1996
567
573
51
Fielder
PJ
Gurney
AL
Stefanich
E
et al
Regulation of thrombopoietin levels by c-mpl-mediated binding to platelets.
Blood.
87
1996
2154
2161
52
Kuter
DJ
Rosenberg
RD
Appearance of a megakaryocyte growth-promoting activity, megapoietin, during acute thrombocytopenia in the rabbit.
Blood.
84
1994
1464
1472
53
Kuter
DJ
Rosenberg
RD
The reciprocal relationship of thrombopoietin (c-Mpl ligand) to changes in the platelet mass during busulfan-induced thrombocytopenia in the rabbit.
Blood.
85
1995
2720
2730
54
Kuter
DJ
The physiology of platelet production.
Stem Cells.
14
1996
88
101
55
Li
J
Xia
Y
Kuter
DJ
Interaction of thrombopoietin with the platelet c-mpl receptor in plasma: binding, internalization, stability and pharmacokinetics.
Br J Haematol.
106
1999
345
356
56
Broudy
VC
Lin
NL
Sabath
DF
Papayannopoulou
T
Kaushansky
K
Human platelets display high-affinity receptors for thrombopoietin.
Blood.
89
1997
1896
1904
57
Scheding
S
Bergmann
M
Shimosaka
A
et al
Human plasma thrombopoietin levels are regulated by binding to platelet thrombopoietin receptors in vivo.
Transfusion.
42
2002
321
327
58
Sungaran
R
Markovic
B
Chong
BH
Localization and regulation of thrombopoietin mRNa expression in human kidney, liver, bone marrow, and spleen using in situ hybridization.
Blood.
89
1997
101
107
59
Quin
S
Fu
F
Li
W
Chen
Q
de Sauvage
FJ
Primary role of the liver in thrombopoietin production shown by tissue-specific knockout.
Blood.
92
1998
2189
2191
60
Peck-Radosavljevic
M
Zacherl
J
Meng
YG
et al
Is inadequate thrombopoietin production a major cause of thrombocytopenia in cirrhosis of the liver?
J Hepatol.
27
1997
127
131
61
Peck-Radosavljevic
M
Wichlas
M
Zacherl
J
et al
Thrombopoietin induces rapid resolution of thrombocytopenia after orthotopic liver transplantation through increased platelet production.
Blood.
95
2000
795
801
62
Kaushansky
K
Lok
S
Holly
RD
et al
Promotion of megakaryocyte progenitor expansion and differentiation by the c-Mpl ligand thrombopoietin.
Nature.
369
1994
568
571
63
Broudy
VC
Lin
NL
Kaushansky
K
Thrombopoietin (c-mpl ligand) acts synergistically with erythropoietin, stem cell factor, and interleukin-11 to enhance murine megakaryocyte colony growth and increases megakaryocyte ploidy in vitro.
Blood.
85
1995
1719
1726
64
Rasko
JE
O'Flaherty
E
Begley
CG
Mpl ligand (MGDF) alone and in combination with stem cell factor (SCF) promotes proliferation and survival of human megakaryocyte, erythroid and granulocyte/macrophage progenitors.
Stem Cells.
15
1997
33
42
65
Ku
H
Yonemura
Y
Kaushansky
K
Ogawa
M
Thrombopoietin, the ligand for the Mpl receptor, synergizes with steel factor and other early acting cytokines in supporting proliferation of primitive hematopoietic progenitors of mice.
Blood.
87
1996
4544
4551
66
Sitnicka
E
Lin
N
Priestley
GV
et al
The effect of thrombopoietin on the proliferation and differentiation of murine hematopoietic stem cells.
Blood.
87
1996
4998
5005
67
de Sauvage
FJ
Carver-Moore
K
Luoh
SM
et al
Physiological regulation of early and late stages of megakaryocytopoiesis by thrombopoietin.
J Exp Med.
183
1996
651
656
68
de Sauvage
FJ
Villeval
JL
Shivdasani
RA
Regulation of megakaryocytopoiesis and platelet production: lessons from animal models.
J Lab Clin Med.
131
1998
496
501
69
Alexander
WS
Roberts
AW
Maurer
AB
Nicola
NA
Dunn
AR
Metcalf
D
Studies of the c-Mpl thrombopoietin receptor through gene disruption and activation.
Stem Cells.
14
1996
124
132
70
Alexander
WS
Roberts
AW
Nicola
NA
Li
R
Metcalf
D
Deficiencies in progenitor cells of multiple hematopoietic lineages and defective megakaryocytopoiesis in mice lacking the thrombopoietic receptor c-Mpl.
Blood.
87
1996
2162
2170
71
Gurney
AL
Carver-Moore
K
de Sauvage
FJ
Moore
MW
Thrombocytopenia in c-mpl-deficient mice.
Science.
265
1994
1445
1447
72
Scott
CL
Robb
L
Mansfield
R
Alexander
WS
Begley
CG
Granulocyte-macrophage colony-stimulating factor is not responsible for residual thrombopoiesis in mpl null mice.
Exp Hematol.
28
2000
1001
1007
73
Kimura
S
Roberts
AW
Metcalf
D
Alexander
WS
Hematopoietic stem cell deficiencies in mice lacking c-Mpl, the receptor for thrombopoietin.
Proc Natl Acad Sci U S A.
95
1998
1195
1200
74
Choi
ES
Hokom
MM
Chen
JL
et al
The role of megakaryocyte growth and development factor in terminal stages of thrombopoiesis.
Br J Haematol.
95
1996
227
233
75
Kojima
H
Hamazaki
Y
Nagata
Y
Todokoro
K
Nagasawa
T
Abe
T
Modulation of platelet activation in vitro by thrombopoietin.
Thromb Haemost.
74
1995
1541
1545
76
Ezumi
Y
Takayama
H
Okuma
M
Thrombopoietin, c-Mpl ligand, induces tyrosine phosphorylation of Tyk2, JAK2, and STAT3, and enhances agonists-induced aggregation in platelets in vitro.
FEBS Lett.
374
1995
48
52
77
Li
J
Kuter
DJ
The end is just the beginning: megakaryocyte apoptosis and platelet release.
Int J Hematol.
74
2001
365
374
78
Begley
CG
Lopez
AF
Nicola
NA
et al
Purified colony-stimulating factors enhance the survival of human neutrophils and eosinophils in vitro: a rapid and sensitive microassay for colony-stimulating factors.
Blood.
68
1986
162
166
79
Lopez
AF
Williamson
DJ
Gamble
JR
et al
Recombinant human granulocyte-macrophage colony-stimulating factor stimulates in vitro mature human neutrophil and eosinophil function, surface receptor expression, and survival.
J Clin Invest.
78
1986
1220
1228
80
Peng
J
Friese
P
Wolf
RF
et al
Relative reactivity of platelets from thrombopoietin- and interleukin-6-treated dogs.
Blood.
87
1996
4158
4163
81
Harker
LA
Hunt
P
Marzec
UM
et al
Regulation of platelet production and function by megakaryocyte growth and development factor in nonhuman primates.
Blood.
87
1996
1833
1844
82
Kroner
C
Eybrechts
K
Akkerman
JW
Dual regulation of platelet protein kinase B.
J Biol Chem.
275
2000
27790
27798
83
Snyder
E
Perrotta
P
Rinder
H
Baril
L
Nichol
J
Gilligan
D
Effect of recombinant human megakaryocyte growth and development factor coupled with polyethylene glycol on the platelet storage lesion.
Transfusion.
39
1999
258
264
84
Xia
Y
Li
J
Bertino
A
Kuter
DJ
Thrombopoietin and the TPO receptor during platelet storage.
Transfusion.
40
2000
976
987
85
Li
J
Yang
C
Xia
Y
et al
Thrombocytopenia caused by the development of antibodies to thrombopoietin.
Blood.
98
2001
3241
3248
86
Hokom
MM
Lacey
D
Kinstler
OB
et al
Pegylated megakaryocyte growth and development factor abrogates the lethal thrombocytopenia associated with carboplatin and irradiation in mice.
Blood.
86
1995
4486
4492
87
Sheridan
WP
Kuter
DJ
Mechanism of action and clinical trials of Mpl ligand.
Curr Opin Hematol.
4
1997
312
316
88
Begley
CG
Basser
RL
Biologic and structural differences of thrombopoietic growth factors.
Semin Hematol.
37
2000
19
27
89
Harker
LA
Marzec
UM
Hunt
P
et al
Dose-response effects of pegylated human megakaryocyte growth and development factor on platelet production and function in nonhuman primates.
Blood.
88
1996
511
521
90
Ulich
TR
del Castillo
J
Yin
S
et al
Megakaryocyte growth and development factor ameliorates carboplatin-induced thrombocytopenia in mice.
Blood.
86
1995
971
976
91
Harker
LA
Marzec
UM
Kelly
AB
et al
Prevention of thrombocytopenia and neutropenia in a nonhuman primate model of marrow suppressive chemotherapy by combining pegylated recombinant human megakaryocyte growth and development factor and recombinant human granulocyte colony-stimulating factor.
Blood.
89
1997
155
165
92
Akahori
H
Shibuya
K
Obuchi
M
et al
Effect of recombinant human thrombopoietin in nonhuman primates with chemotherapy-induced thrombocytopenia.
Br J Haematol.
94
1996
722
728
93
Neelis
KJ
Hartong
SC
Egeland
T
Thomas
GR
Eaton
DL
Wagemaker
G
The efficacy of single-dose administration of thrombopoietin with coadministration of either granulocyte/macrophage or granulocyte colony- stimulating factor in myelosuppressed rhesus monkeys.
Blood.
90
1997
2565
2573
94
Giri
JG
Smith
WG
Kahn
LE
et al
Promegapoietin, a chimeric growth factor for megakaryocyte and platelet restoration [abstract].
Blood.
90
1997
580a
95
Cwirla
SE
Balasubramanian
P
Duffin
DJ
et al
Peptide agonist of the thrombopoietin receptor as potent as the natural cytokine.
Science.
276
1997
1696
1699
96
Erickson-Miller
CL
Delorme
E
Tian
SS
et al
Discovery and characterization of a selective, non-peptidyl thrombopoietin receptor agonist [abstract].
Blood.
96
2000
675a
97
Naranda
T
Wong
K
Kaufman
RI
Goldstein
A
Olsson
L
Activation of erythropoietin receptor in the absence of hormone by a peptide that binds to a domain different from the hormone binding site.
Proc Natl Acad Sci U S A.
96
1999
7569
7574
98
Fanucchi
M
Glaspy
J
Crawford
J
et al
Effects of polyethylene glycol-conjugated recombinant human megakaryocyte growth and development factor on platelet counts after chemotherapy for lung cancer.
N Engl J Med.
336
1997
404
409
99
Basser
RL
Rasko
JE
Clarke
K
et al
Randomized, blinded, placebo-controlled phase I trial of pegylated recombinant human megakaryocyte growth and development factor with filgrastim after dose-intensive chemotherapy in patients with advanced cancer.
Blood.
89
1997
3118
3128
100
Crawford
J
Glaspy
J
Belani
C
et al
A randomized, placebo-controlled, blinded, dose scheduling trial of pegylated recombinant human megakaryocyte growth and development factor (PEG-HUMGDF) with filgrastim support in non-small cell lung cancer (NSCLC) patients treated with paclitaxel and carboplatin during multiple cycles of chemotherapy [abstract].
Proceedings ASCO.
17
1998
73a
101
Vadhan-Raj
S
Patel
S
Broxmeyer
HE
et al
Phase I-II investigtion of recombinant human thrombopoietin (rhTPO) in patients with sarcoma receiving high dose chemotherapy (CT) with adriamycin (A) and ifosfamide (I) [abstract].
Blood.
88
1996
448a
102
Vadhan-Raj
S
Murray
LJ
Bueso-Ramos
C
et al
Stimulation of megakaryocyte and platelet production by a single dose of recombinant human thrombopoietin in patients with cancer.
Ann Int Med.
126
1997
673
681
103
Vadhan-Raj
S
Verschraegen
C
McGarry
L
et al
Recombinant human thrombopoietin (rhTPO) attenuates high-dose carboplatin (C)-induced thrombocytopenia in patients with gynecological malignancy [abstract].
Blood.
90
1997
580a
104
Vadhan-Raj
S
Verschraegen
CF
Bueso-Ramos
C
et al
Recombinant human thrombopoietin attenuates carboplatin-induced severe thrombocytopenia and the need for platelet transfusions in patients with gynecologic cancer.
Ann Intern Med.
132
2000
364
368
105
Basser
RL
Rasko
JE
Clarke
K
et al
Thrombopoietic effects of pegylated recombinant human megakaryocyte growth and development factor (PEG-rHuMGDF) in patients with advanced cancer.
Lancet.
348
1996
1279
1281
106
Basser
RL
Underhill
C
Davis
I
et al
Enhancement of platelet recovery after myelosuppressive chemotherapy by recombinant human megakaryocyte growth and development factor in patients with advanced cancer.
J Clin Oncol.
18
2000
2852
2861
107
Moskowitz
C
Nimer
S
Gabrilove
J
et al
A randomized, double blind, placebo-controlled, dose finding, efficacy and safety study of PEG-rHuMGDF (M) in non-Hodgkin's lymphoma (NHL) patients (pts) treated with ICE (ifosfamide, carboplatin and etoposide) [abstract].
J Clin Oncol.
17
1998
76a
108
O'Malley
CJ
Rasko
JE
Basser
RL
et al
Administration of pegylated recombinant human megakaryocyte growth and development factor to humans stimulates the production of functional platelets that show no evidence of in vivo activation.
Blood.
88
1996
3288
3298
109
Vadhan-Raj
S
Patel
S
Broxmeyer
H
et al
Schedule-dependent reduction in thrombocytopenia by recombinant human thrombopoietin (rhTPO) in patients with sarcoma receiving high dose chemotherapy (CT) with adriamycin (A) and ifosfamide (I) [abstract].
J Clin Oncol.
18
1999
52a
110
Tornebohm
E
Lockner
D
Paul
C
A retrospective analysis of bleeding complications in 438 patients with acute leukaemia during the years 1972-1991.
Eur J Haematol.
50
1993
160
167
111
Anderlini
P
Luna
M
Kantarjian
HM
et al
Causes of initial remission induction failure in patients with acute myeloid leukemia and myelodysplastic syndromes.
Leukemia.
10
1996
600
608
112
Archimbaud
E
Ottmann
OG
Yin
JA
et al
A randomized, double-blind, placebo-controlled study with pegylated recombinant human megakaryocyte growth and development factor (PEG-rHuMGDF) as an adjunct to chemotherapy for adults with de novo acute myeloid leukemia.
Blood.
94
1999
3694
3701
113
Archimbaud
E
Ottmann
O
Lin
J
et al
A randomized, double-blind, placebo-controlled study using PEG-rHuMGDF as an adjunct to chemotherapy for adults with de-novo acute myeloid leukemia (AML): early results [abstract].
Blood.
99
1996
447a
114
Schiffer
CA
Miller
K
Larson
RA
et al
A double-blind, placebo-controlled trial of pegylated recombinant human megakaryocyte growth and development factor as an adjunct to induction and consolidation therapy for patients with acute myeloid leukemia.
Blood.
95
2000
2530
2535
115
Cripe
L
Neuberg
D
Tallman
M
et al
A pilot study of recombinant human thrombopoietin (rh-TPO) and GM-CSF following induction therapy in patients older than 55 years with acute myeloid leukemia (AML) [abstract].
Blood.
92
1998
616a
116
Glaspy
J
Vredenburgh
J
Demetri
GD
et al
Effects of PEGylated recombinant human megakaryocyte growth and development factor (PEG-rHuMGDF) before high dose chemotherapy (HDC) with peripheral blood progenitor cell (PBPC) support [abstract].
Blood.
90
1997
580a
117
Bolwell
B
Vredenburgh
J
Overmoyer
B
et al
Safety and biological effect of pegylated recombinant megakaryocyte growth and development factor (PEG-rHuMGDF) in breast cancer patients following autologous peripheral blood progenitor cell transplantation (PBPC) [abstract].
Blood.
90
1997
171a
118
Beveridge
R
Schuste
RM
Waller
E
et al
Randomized, double-blind, placebo-controlled trial of pegylated recombinant megakaryocyte growth and development factor (PEG-rHuMGDF) in breast cancer patients following autologous bone marrow transplantation [abstract].
Blood.
90
1997
580a
119
Nash
R
Kurzrock
R
DiPersio
J
et al
Safety and activity of recombinant human thrombopoietin (rhTPO) in patients (pts) with delayed platelet recovery (DPR) [abstract].
Blood.
90
1997
262a
120
Nash
RA
Kurzrock
R
DiPersio
J
et al
A phase I trial of recombinant human thrombopoietin in patients with delayed platelet recovery after hematopoietic stem cell transplantation.
Biol Blood Marrow Transplant.
6
2000
25
34
121
Somlo
G
Sniecinski
I
ter Veer
A
et al
Recombinant human thrombopoietin in combination with granulocyte colony-stimulating factor enhances mobilization of peripheral blood progenitor cells, increases peripheral blood platelet concentration, and accelerates hematopoietic recovery following high-dose chemotherapy.
Blood.
93
1999
2798
2806
122
Gajewski
J
Korbling
M
Donato
M
et al
Recombinant human thrombopoietin (rhTPO) for mobilization of peripheral blood progenitor cells (PBPC) for autologous transplantation in breast cancer: preliminary results of a phase I trial [abstract].
Blood.
90
1997
97a
123
Linker
C
Anderlini
P
Herzig
R
et al
A randomized, placebo-controlled, phase II trial of recombinant human thrombopoietin (rhTPO) in subjects undergoing high dose chemotherapy (HDC) and PBPC transplant [abstract].
Blood.
92
1998
682a
124
Linker
C
Thrombopoietin in the treatment of acute myeloid leukemia and in stem-cell transplantation.
Semin Hematol.
37
2000
35
40
125
Bolwell
B
Vredenburgh
J
Overmoyer
B
et al
Phase 1 study of pegylated recombinant human megakaryocyte growth and development factor (PEG-rHuMGDF) in breast cancer patients after autologous peripheral blood progenitor cell (PBPC) transplantation.
Bone Marrow Transplant.
26
2000
141
145
126
Fibbe
WE
Heemskerk
DP
Laterveer
L
et al
Accelerated reconstitution of platelets and erythrocytes after syngeneic transplantation of bone marrow cells derived from thrombopoietin pretreated donor mice.
Blood.
86
1995
3308
3313
127
Liu Yin
JA
Adams
JA
Brereton
ML
Hann
A
Harrison
BD
Briggs
M
Megakaryopoiesis in vitro in myelodysplastic syndromes and acute myeloid leukaemia: effect of pegylated recombinant human megakaryocyte growth and development factor in combination with other growth factors.
Br J Haematol.
108
2000
743
746
128
Fontenay-Roupie
M
Dupont
JM
Picard
F
et al
Analysis of megakaryocyte growth and development factor (thrombopoietin) effects on blast cell and megakaryocyte growth in myelodysplasia.
Leuk Res.
22
1998
527
535
129
Nichol
JL
Thrombopoietin levels after chemotherapy and in naturally occurring human diseases.
Curr Opin Hematol.
5
1998
203
208
130
Komatsu
N
Okamoto
T
Yoshida
T
et al
Pegylated recombinant human megakaryocyte growth and development factor (PEG-rHuMGDF) increased platelet counts (plt) in patients with aplastic anemia (AA) and myelodysplastic syndrome (MDS) [abstract].
Blood.
96
2000
296a
131
Harker
LA
Marzec
UM
Novembre
F
et al
Treatment of thrombocytopenia in chimpanzees infected with human immunodeficiency virus by pegylated recombinant human megakaryocyte growth and development factor.
Blood.
91
1998
4427
4433
132
Harker
LA
Carter
RA
Marzec
UM
et al
Correction of thrombocytopenia and ineffective platelet production in patients infected with human immunodeficiency virus (HIV) by PEG-rHuMGDF therapy [abstract].
Blood.
92(suppl 1)
1998
707a
133
Dan
K
Nomura
S
Hotta
T
Fujimura
K
Ikeda
Y
Biological effect of pegylated recombinant megakaryocyte growth and development factor (PEG-MGDF) in patients with idiopathic thrombocytopenic purpura [abstract].
Blood.
98
2001
299a
134
Rice
L
Nichol
JL
Delavari
M
Roskos
L
Bacille
MH
McMillan
R
Cyclic thrombocytopenia with platelet auto-antibodies: response to PEG-rHu megakaryocyte growth and development factor [abstract].
Blood.
92
1998
180b
135
Siemensma
NP
Bathal
PS
Penington
DG
The effect of massive liver resection on platelet kinetics in the rat.
J Lab Clin Med.
86
1975
817
833
136
Kuter
DJ
The use of PEG-rhuMGDF in platelet apheresis.
Stem Cells.
16
1998
231
242
137
Nakamura
M
Toombs
CF
Duarte
IG
et al
Recombinant human megakaryocyte growth and development factor attenuates postbypass thrombocytopenia.
Ann Thorac Surg.
66
1998
1216
1223
138
Rasko
JE
Basser
RL
Boyd
J
et al
Multilineage mobilization of peripheral blood progenitor cells in humans following administration of PEG-rHuMGDF.
Br J Haematol.
97
1997
871
880
139
Murray
LJ
Luens
KM
Estrada
MF
et al
Thrombopoietin mobilizes CD34+ cell subsets into peripheral blood and expands multilineage progenitors in bone marrow of cancer patients with normal hematopoiesis.
Exp Hematol.
26
1998
207
216
140
Yagi
M
Ritchie
KA
Sitnicka
E
Storey
C
Roth
GJ
Bartelmez
S
Sustained ex vivo expansion of hematopoietic stem cells mediated by thrombopoietin.
Proc Natl Acad Sci U S A.
96
1999
8126
8131
141
Piacibello
W
Sanavio
F
Garetto
L
et al
Extensive amplification and self-renewal of human primitive hematopoietic stem cells from cord blood.
Blood.
89
1997
2644
2653
142
Kuter
DJ
Goodnough
LT
Romo
J
et al
Thrombopoietin therapy increases platelet yields in healthy platelet donors.
Blood.
98
2001
1339
1345
143
Goodnough
LT
Kuter
DJ
McCullough
J
et al
Prophylactic platelet transfusions from healthy apheresis platelet donors undergoing treatment with thrombopoietin.
Blood.
98
2001
1346
1351
144
Neelis
KJ
Visser
TP
Dimjati
W
et al
A single dose of thrombopoietin shortly after myelosuppressive total body irradiation prevents pancytopenia in mice by promoting short-term multilineage spleen-repopulating cells at the transient expense of bone marrow-repopulating cells.
Blood.
92
1998
1586
1597
145
Kaushansky
K
Lin
N
Grossmann
A
Humes
J
Sprugel
KH
Broudy
VC
Thrombopoietin expands erythroid, granulocyte-macrophage, and megakaryocytic progenitor cells in normal and myelosuppressed mice.
Exp Hematol.
24
1996
265
269
146
Kaushansky
K
Thrombopoietin: more than a lineage-specific megakaryocyte growth factor.
Stem Cells.
15(suppl 1)
1997
97
103
147
Mouthon
MA
Van der Meeren
A
Gaugler
MH
et al
Thrombopoietin promotes hematopoietic recovery and survival after high-dose whole body irradiation.
Int J Radiat Oncol Biol Phys.
43
1999
867
875
148
Molineux
G
Hartley
C
McElroy
P
McCrea
C
Kerzic
P
McNiece
I
An analysis of the effects of combined treatment with rmGM-CSF and PEG- rHuMGDF in murine bone marrow transplant recipients.
Stem Cells.
15
1997
43
49
149
Yang
C
Xia
Y
Li
J
Kuter
DJ
The appearance of anti-thrombopoietin antibody and circulating thrombopoietin-IgG complexes in a patient developing thrombocytopenia after the injection of PEG-rHuMGDF [abstract].
Blood.
94
1999
681a
150
Basser
RL
O'Flaherty
E
Green
M
et al
Development of pancytopenia with neutralizing antibodies to thrombopoietin after multicycle chemotherapy supported by megakaryocyte growth and development factor.
Blood.
99
2002
2599
2602
151
F-D-C Reports
In Brief: Amgen Megagen.
The Pink Sheet.
60
1998
27
152
Rebulla
P
Finazzi
G
Marangoni
F
et al
The threshold for prophylactic platelet transfusions in adults with acute myeloid leukemia.
N Engl J Med.
337
1997
1870
1875
153
Rebulla
P
Trigger for platelet transfusion.
Vox Sang.
78(suppl 2)
2000
179
182
154
Wandt
H
Frank
M
Ehninger
G
et al
Safety and cost effectiveness of a 10 × 10(9)/L trigger for prophylactic platelet transfusions compared with the traditional 20 × 10(9)/L trigger: a prospective comparative trial in 105 patients with acute myeloid leukemia.
Blood.
91
1998
3601
3606
155
Kuter
DJ
Whatever happened to thrombopoietin?
Transfusion.
42
2002
279
283

Author notes

David J. Kuter, Hematology/Oncology Unit, Massachusetts General Hospital, Harvard Medical School, 100 Blossom St, Boston, MA 02114; e-mail:kuter.david@mgh.harvard.edu.