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REVIEW ARTICLE
From the Hematology/Oncology Unit, Massachusetts
General Hospital, Harvard Medical School, Boston, MA; and the Centre
for Child Health Research, The University of Western Australia and
Western Australian Institute for Medical Research, Perth, Western
Australia.
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 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.1 Multiagent 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).3-6
The 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
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).16-26
Early 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-1 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.
Isolation of c-Mpl ligand
Structure and biologic properties of TPO
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.58
Direct evidence to support the relative contribution of this mechanism to platelet production is lacking; but in experiments in which livers
from TPO 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 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 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
B Recombinant TPO
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
Nonmyeloablative treatments The stimulatory effects of PEG-rHuMGDF and rhTPO on megakaryocyte and platelet production have been demonstrated in several clinical trials (Table 2).98-107 PEG-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
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.106 Patients 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.102 This 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 (Figure
3). The need for platelet transfusions
decreased by 75% (Figure
4).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 (Tables 3 and 4).112-125
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.130Human 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.60,61 Within 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 (Figure
5). 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 (Table
4).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.
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 (Figure
6). 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.
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.
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 (Figure 9).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.
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
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.
Submitted November 8, 2001; accepted July 24, 2002.
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.
Reprints: David J. Kuter, Hematology/Oncology Unit, Massachusetts General Hospital, Harvard Medical School, 100 Blossom St, Boston, MA 02114; e-mail: kuter.david{at}mgh.harvard.edu.
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Abstract
Introduction
recombinant human thrombopoietin (rhTPO) and
pegylated recombinant human megakaryocyte growth and development factor
(PEG-rHuMGDF)
before or after carboplatin therapy increased
platelet counts and was effective at attenuating thrombocytopenia
associated with chemotherapy.
