We examined the effects of recombinant human thrombopoietin (rhTPO) on myelosuppressive chemotherapy-induced thrombocytopenia in cynomolgus monkeys. After treatment with nimustine (ACNU) on day 0, the monkeys intravenously received rhTPO at a dose of 0.04, 0.2 or 1 microgram/kg/d or monkey's serum once each day from day 1 to day 28. Administration of rhTPO reduced the severity of thrombocytopenia and accelerated the rate of platelet recovery in a dose-dependent fashion. Treatment with the highest rhTPO dose completely prevented thrombocytopenia and stimulated a marked increase in platelet counts over the normal values. Animals treated with ACNU also became neutropenic and slightly anaemic. Administration of rhTPO following ACNU treatment significantly improved neutropenia with increasing doses of rhTPO, but had no effect on anaemia. Compared to the control animals, rhTPO-treated animals exhibited no significant changes in several serum parameters. C-reactive protein concentration and some blood coagulation profiles within the study period. These results suggest a therapeutic efficacy of rhTPO in improving chemotherapy-induced thrombocytopenia.

Abstract. The in vivo effects of pegylated recombinant human megakaryocyte growth and development factor (PEG-rHuMGDF), a truncated molecule of recombinant human thrombopoietin modified with polyethylene glycol, were investigated in normal Balb/c mice. PEG-rHuMGDF was more potent in producing platelets and the dose-response curve was steeper compared with the case of the nonpegylated form of this molecule. Five consecutive injections with PEG-rHuMGDF caused a dose-dependent increase in peripheral platelet counts with a peak on day 8. There was a dose-dependent rise in platelet counts on day 8 at daily doses from 0.333 to 30 µg/kg. Intermediate doses of PEG-rHuMGDF (1.111 to 10 µg/kg/day) caused a significant decrease in mean platelet volume, and conversely, higher doses of PEG-rHuMGDF (30 to 270 µg/kg/day) induced a dose-dependent increase in mean platelet volume. There was a dose-dependent decrease in hemoglobin concentration with a minimum on day 8 but no significant reduction in reticulocyte counts following PEG-rHuMGDF administration. White blood cell counts were unchanged by PEG-rHuMGDF treatment. Marrow megakaryocyte size enlarged to 1.5-fold and the number of marrow megakaryocytes increased to sixfold by consecutive administration of PEG-rHuMGDF at 30 µg/kg/day. A twofold increase in the number of marrow megakaryocytic progenitor cells (colony-forming units-megakaryocyte) was also observed. Marrow erythroid progenitor (colony forming units-erythroid) counts decreased by splenic colony-forming units-erythroid, marrow and splenic erythro/myeloid progenitor cell counts, and splenic granulocyte/macrophage progenitor cell counts increased with PEG-rHuMGDF treatment. Marrow and splenic erythroid burst-forming cells were unchanged. These results indicate that PEGrHuMGDF, a truncated molecule of thrombopoietin, is a potent stimulator for megakaryopoiesis and thrombopoiesis, and also affects the development of other hematopoietic cells in normal mice.

Abstract. Thrombopoietin, the endogenous c-Mpl ligand, is a novel lineage-specific hematopoietic factor that plays a pivotal role in the regulation of megakaryocytopoiesis and thrombopoiesis. In this study, we examined the effects of pegylated recombinant human megakaryocyte growth and development factor (PEG-rHuMGDF), a truncated molecule of recombinant human c-mpl ligand derivatized with polyethylene glycol, on myelosuppressive chemotherapy-induced thrombocytopenia in mice. We developed a new murine model of thrombocytopenia induced by I.V. injections of mitomycin C (MMC) for two consecutive days. In control mice, platelet counts began to decrease on day 6, reached a nadir of less than 5% of basal level on day 14, and could not recover to basal level by day 26. Administration of PEG-rHuMGDF greatly enhanced recovery of the number of megakaryocyte progenitor cells and the megakaryocytes in bone marrow, and markedly reduced the severity of thrombocytopenia; it also accelerated platelet recovery in a dose-dependent manner in myelosuppressed mice. Mice receiving consecutive administration of higher doses of PEG-rHuMGDF showed no thrombocytopenia but rather had platelet counts being increased over basal level. Although absolute neutrophil counts and red cell counts also were decreased following MMC treatment, administration of PEG-rHuMGDF also improved neutropenia and anemia. Administration of PEGrHuMGDF on alternate days or once a week after chemotherapy was almost as effective as consecutive administration in improving thrombocytopenia. Combined administration of PEGrHuMGDF and rHuG-CSF had an addictive effect on improvement of thrombocytopenia and neutropenia. These results suggest that PEGrHuMGDF is a therapeutically effective agent in the treatment of thrombocytopenia associated with chemotherapy.

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Figure 3 Administration schedules of PEG-rHuMGDF following BMT in mice. Subcutaneous administration schedules of PEG-rHuMGDF were shown as follows: (1) consecutive treatment at a dose of 30 g/kg/day from the day after BMT (day 1) for 13 consecutive days ( ); (2) alternate-day treatment at a dose of 55.7 g/kg/day on days 1, 3, 5, 7, 9, 11 and 13 (`); and (3) at an interval of 3 days at a dose of 78 g/kg/day on days 1, 5, 9 and 13 ( ). The same volume of vehicle solution was injected as control from the day after BMT for 13 consecutive days ( ). Each bar indicates the mean Ϯ s.e. of five to six mice. Significant differences (*P Ͻ 0.05, **P Ͻ 0.01) compared with the vehicle-treated mice were tested by Dunnett's test.