A comparison of treatment of canine cyclic hematopoiesis with recombinant human granulocyte-macrophage colony-stimulating factor (GM-CSF), G-CSF interleukin-3, and canine G-CSF.

Cyclic hematopoiesis in gray collie dogs is a stem cell disease in which abnormal regulation of cell production in the bone marrow causes cyclic fluctuations of blood cell counts. In vitro studies demonstrated that recombinant human granulocyte-macrophage colony-stimulating factor (GM-CSF), interleukin-3 (IL-3), and granulocyte colony stimulating factor (G-CSF) all stimulated increases in colony formation by canine bone marrow progenitor cells. Based on these results, gray collie dogs were then treated with recombinant human (rh) GM-CSF, IL-3, or G-CSF subcutaneously to test the hypothesis that pharmacologic doses of one of these hematopoietic growth factors could alter cyclic production of cells. When recombinant canine G-CSF became available, it was tested over a range of doses. In vivo rhIL-3 had no effect on the recurrent neutropenia but was associated with eosinophilia, rhGM-CSF caused neutrophilia and eosinophilia but cycling of hematopoiesis persisted. However, rhG-CSF caused neutrophilia, prevented the recurrent neutropenia and, in the two animals not developing antibodies to rhG-CSF, obliterated periodic fluctuation of monocyte, eosinophil, reticulocyte, and platelet counts. Recombinant canine G-CSF increased the nadir neutrophil counts and amplitude of fluctuations at low doses (1 micrograms/kg/d) and eliminated all cycling of cell counts at high doses (5 and 10 micrograms/kg/d). These data suggest significant differences in the actions of these growth factors and imply a critical role for G-CSF in the homeostatic regulation of hematopoiesis.     

Clinical significance of recombinant human granulocyte colony-stimulating factor (rG-CSF) in the chemotherapy of patients with malignant lymphoma]

We examined the effects of recombinant human granulocyte colony-stimulating factor (rG-CSF) on neutropenia induced by chemotherapy in 10 patients with non-Hodgkin's lymphoma (NHL). The numbers of peripheral blood hematopoietic progenitors were also evaluated before and after administration of rG-CSF. Six patients received an administration of 2 micrograms/kg/body weight of rG-CSF subcutaneously for 14 days after 2nd chemotherapy. Four patients received intravenous infusion of rG-CSF (300 micrograms/body/day) for 4 days from nadir state after chemotherapy. Administration of rG-CSF from the termination of chemotherapy, markedly shortend the period of bone marrow hypoplasia induced by chemotherapy. On the other hand, administration of rhG-CSF from nadir state after chemotherapy have accelerated the recovery of neutrophil counts. In addition, this type of therapy induced 26 to 60 folds increase of peripheral blood hematopoietic progenitors. These results demonstrate the validity of administration of rhG-CSF not only in the chemotherapy of NHL, but also in peripheral blood stem cell transplantation (PBSCT).     

Clinical evaluation of recombinant human granulocyte colony-stimulating factor (rhG-CSF) in autologous bone marrow transplantation]

We administered recombinant human granulocyte colony-stimulating factor (rhG-CSF) at 5 micrograms/kg/day by intravenous drip infusion for 21 consecutive days in autologous bone marrow transplanted patients. The period of posttransplant neutropenia was markedly shortened by the rhG-CSF treatment; mean days required for neutrophil recovery (greater than 500/mm3) of 14.3 days in the rhG-CSF group (n = 21) versus 27.8 days in the historical control group (n = 11). More importantly, the numbers of febrile days between day 15 and day 28 were found to be fewer in the rG-CSF group than in control group. These effects were obtained without delay in the recovery of other blood cell series and without any side effect. We conclude that the posttransplant use of the rhG-CSF is beneficial for prevention and treatment of infectious complications after autologous bone marrow transplantation.     

Clinical effects of recombinant human granulocyte colony-stimulating factor in leukemia patients: a phase I/II study

A phase I/II study of recombinant human granulocyte colony-stimulating factor (rhG-CSF) in 24 leukemia patients was conducted at our institute. Recombinant human G-CSF (50-200 micrograms/m2/day) was administered i.v. In seven allogeneic bone marrow transplantation (BMT) recipients, treatment with rhG-CSF was started 5 days after BMT. Neutrophils began to increase within 3 days after the start of rhG-CSF administration in five of seven patients. The mean duration necessary for recovery of neutrophils to greater than 500/microliters was 11.3 days after BMT with rhG-CSF; 26.8 days is the figure for recovery without rhG-CSF from Japanese historical data. In seven out of eight patients who received rhG-CSF administration after the first remission-induction chemotherapy, the neutrophil counts increased from less than 300/microliters to greater than 4000/microliters within 10 days. Blasts did not increase in all patients including four acute nonlymphocytic leukemia (ANLL) patients. Severe infections such as septicemia and pneumonia, which were unable to be controlled by antibiotics only, were successfully treated with rhG-CSF and antibiotics. rhG-CSF either stimulated or inhibited myeloid leukemic cells in some refractory cases. Mild bone pain occurred in one patient while receiving rhG-CSF i.v. rhG-CSF seems to have the ability to shorten the period of neutropenia, prevent infections after allogeneic BMT and remission-induction chemotherapy for acute leukemia, and support therapy for infections.     

Combination therapy for radiation-induced bone marrow aplasia in nonhuman primates using synthokine SC-55494 and recombinant human granulocyte colony-stimulating factor

Combination cytokine therapy continues to be evaluated in an effort to stimulate multilineage hematopoietic reconstitution after bone marrow myelosuppression. This study evaluated the efficacy of combination therapy with the synthetic interleukin-3 receptor agonist, Synthokine- SC55494, and recombinant methionyl human granulocyte colony-stimulating factor (rhG-CSF) on platelet and neutrophil recovery in nonhuman primates exposed to total body 700 cGy 60Co gamma radiation. After irradiation on day (d) 0, cohorts of animals subcutaneously received single-agent protocols of either human serum albumin (HSA; every day [QD], 15 micrograms/kg/d, n = 10), Synthokine (twice daily [BID], 100, micrograms/kg/d, n = 15), rhG-CSF (QD, 10 micrograms/kg/d, n = 5), or a combination of Synthokine and rhG-CSF (BID, 100 and 10 micrograms/kg/d, respectively, n = 5) for 23 days beginning on d1. Complete blood counts were monitored for 60 days postirradiation and the durations of neutropenia (absolute neutrophil count < 500/microL) and thrombocytopenia (platelet count < 20,000/microL) were assessed. Animals were provided clinical support in the form of antibiotics, fresh irradiated whole blood, and fluids. All cytokine protocols significantly (P < .05) reduced the duration thrombocytopenia versus the HSA-treated animals. Only the combination protocol of Synthokine + rhG-CSF and rhG-CSF alone significantly shortened the period neutropenia (P < .05). The combined Synthokine/rhG-CSF protocol significantly improved platelet nadir versus Synthokine alone and HSA controls and neutrophil nadir versus rhG-CSF alone and HSA controls. All cytokine protocols decreased the time to recovery to preirradiation neutrophil and platelet values. The Synthokine/rhG-CSF protocol also reduced the transfusion requirements per treatment group to 0 among 5 animals as compared with 2 among 5 animals for Synthokine alone, 8 among 5 animals for rhG-CSF, and 17 among 10 animals for HSA. These data showed that the combination of Synthokine, SC-55494, and rhG-CSF further decreased the cytopenic periods and nadirs for both platelets and neutrophils relative to Synthokine and rhG-CSF monotherapy and suggest that this combination therapy would be effective against both neutropenia and thrombocytopenia consequent to drug- or radiation- induced myelosuppression.     

Effect of stem cell factor with and without granulocyte colony-stimulating factor on neonatal hematopoiesis: in vivo induction of newborn myelopoiesis and reduction of mortality during experimental group B streptococcal sepsis.

Neonatal hematopoiesis and host defense are developmentally immature and under states of increased demand predispose the newborn to peripheral cytopenias and depletion of bone marrow storage pool reserves. We have previously demonstrated that recombinant human granulocyte colony-stimulating factor (rhG-CSF) can significantly modulate neonatal rat granulopoiesis and act synergistically with antibiotic therapy to reduce the mortality rate during experimental group B streptococcal sepsis. Stem cell factor (SCF) has been shown to stimulate early hematopoietic progenitor cells and, in the presence of lineage-specific CSFs, enhance committed progenitor cell proliferation. In the present study we examined the in vivo neonatal hematologic effects of recombinant rat (rr) SCF (14 days), simultaneous rrSCF + rhG-CSF (14 days), and sequential combination of rrSCF (7 days) + rhG-CSF (7 days). Sprague-Dawley newborn rats (less than or equal to 24 hours) were injected intraperitoneal (IP) x 14 days with the above combinations. rrSCF (0 to 200 micrograms/kg/d) had a negligible effect on the peripheral platelet count and absolute neutrophil count (ANC) but the diminution in the hematocrit during the first 10 days of treatment was less pronounced (P = .0001). However, the simultaneous use of rrSCF + rhG-CSF synergistically increased the circulating day 6 to 13 ANC (P = .001). Similarly, sequential rrSCF + rhG-SCF also had a synergistic significant effect during the second week of therapy on the circulating ANC (P = .01). The bone marrow neutrophil storage and proliferative pools were also significantly increased in newborn rats treated with rrSCF + rhG-CSF versus rhG-CSF (P = .02). The bone marrow and liver/spleen CFU-GM pool was unchanged; however, the CFU-GM proliferative rates were significantly increased in the rrSCF + rhG-CSF group (P = .04). rrSCF also induced a significant increase in the bone marrow and liver/spleen mast cell pool (P = .002). Lastly, rrSCF x 14 days +/- rhG-CSF significantly reduced the mortality rate at 48 and 120 hours after experimental group B streptococcus sepsis (P = .03 and .05, respectively). These data suggest that combination SCF + G-CSF therapy compared with G-CSF alone significantly increases the neonatal rat peripheral neutrophil count, bone marrow myeloid pools and proliferative rates, and induces a reduction in the mortality rate during experimental bacterial sepsis. SCF therapy may have future potential applications in the modulation of human neonatal hematopoiesis and host defense.     

Additive effects of human recombinant interleukin-11 and granulocyte colony-stimulating factor in experimental gram-negative sepsis

Recombinant human granulocyte colony-stimulating factor (rhG-CSF) is widely used to promote granulocyte recovery from a variety of pathologic states. Recombinant human interleukin-11 (rhIL-11) has recently become available clinically as a platelet restorative agent after myelosuppressive chemotherapy. Preclinical data has shown that rhIL-11 limits mucosal injury after chemotherapy and attenuates the proinflammatory cytokine response. The potential efficacy of combination therapy with recombinant human forms of rhIL-11 and rhG-CSF was studied in a neutropenic rat model of Pseudomonas aeruginosa sepsis. At the onset of neutropenia, animals were randomly assigned to receive either rhG-CSF at a dose of 200 micrograms/kg subcutaneously every 24 hours for 7 days; rhIL-11 at 200 micrograms/kg subcutaneously every 24 hours for 7 days; the combination of both rhG-CSF and rhIL-11; or saline control. Animals were orally colonized with Pseudomonas aeruginosa 12.4.4 and then given a myelosuppressive dose of cyclophosphamide. rhG-CSF resulted in a slight increase in absolute neutrophil counts (ANC), but did not provide a survival advantage (0 of 12, 0% survival) compared with the placebo group (1 of 12, 8% survival). rhIL-11 was partially protective (4 of 10, 40% survival); the combination of rhG-CSF and rhIL-11 resulted in a survival rate of 80% (16 of 20; P <.001). rhIL-11 alone or in combination with rhG-CSF resulted in preservation of gastrointestinal mucosal integrity (P <.001), lower circulating endotoxin levels (P <.01), and reduced quantitative levels of P. aeruginosa in quantitative organ cultures. These results indicate that the combination of rhIL-11 and rhG-CSF is additive as a treatment strategy in the prevention and treatment of experimental Gram-negative sepsis in immunocompromised animals. This combination may prove to be efficacious in the prevention of severe sepsis in neutropenic patients.     

Seven-day administration of recombinant human granulocyte colony-stimulating factor to newborn rats: modulation of neonatal neutrophilia, myelopoiesis, and group B Streptococcus sepsis.

Single-pulse administration of rhG-colony-stimulating factor (CSF) to neonatal rats was previously demonstrated to induce peripheral neutrophilia and modulate bone marrow (BM) neutrophil storage and proliferative pools (NSP + NPP). In this study, we investigated the prolonged effects of 7 days of rhG-CSF therapy (5 micrograms/kg/per day). Sprague-Dawley newborn rats (less than or equal to 24 hours) were injected intraperitoneally (IP) (daily for 7 days) with rhG-CSF or phosphate-buffered saline/human serum albumin (PBS/HSA). RhG-CSF induced a significant early and late peripheral neutrophilia: 6,905 +/- 1,625 (day 1) and 9,223 +/- 515 microL (day 7) v 1,275 +/- 90/microL (P less than or equal to .0001). In addition, 7 days of rhG-CSF resulted in a significant increase in the BM NSP: 3,247 +/- 190/microL v 1,677 +/- 339/microL (P less than or equal to .001). There was, however, no depletion or significant change in the BM NPP. Seven days of rhG-CSF also induced a mild increase in BM CFU-GM colony formation (P less than or equal to .01). There was, however, no significant change in liver/spleen CFU-GM colonies or in the CFU-GM proliferative rate in either the BM or liver/spleen cultures. Finally, 7 days of prophylactic rhG-CSF therapy resulted in a synergistic response with antibiotic therapy and significantly modulated the mortality rate during experimental group B streptococcal sepsis (GBS) (100% v 50%) (GvsC) (P less than or equal to .001). Pulse rhG-CSF administered at 6 hours or 18 hours after GBS inoculation, however, failed to act synergistically with antibiotics to improve survival or prevent peripheral neutropenia. This study suggests that 7 days of prophylactic rhG-CSF therapy induces peripheral neutrophilia, myeloid maturation, increases neutrophil BM reserves and also may provide immunologic enhancement of neonatal host defense during experimental GBS in term neonatal rats.     

Results of a phase I/II trial of recombinant human granulocyte- macrophage colony-stimulating factor in very low birthweight neonates: significant induction of circulatory neutrophils, monocytes, platelets, and bone marrow neutrophils

Neonates, especially those of very low birthweight (VLBW), have an increased risk of nosocomial infections secondary to deficiencies in development. We previously demonstrated that granulocyte-macrophage colony-stimulating factor (GM-CSF) production and mRNA expression from stimulated neonatal mononuclear cells are significantly less than that from adult cells. Recombinant murine GM-CSF administration to neonatal rats has resulted in neutrophilia, increased neutrophil production, and increased survival of pups during experimental Staphylococcus aureus sepsis. In the present study, we sought to determine the safety and biologic response of recombinant human (rhu) GM-CSF in VLBW neonates. Twenty VLBW neonates (500 to 1,500 g), aged < 72 hours, were randomized to receive either placebo (n = 5) or rhuGM-CSF at 5.0 micrograms/kg once per day (n = 5), 5.0 micrograms/kg twice per day (n = 5), or 10 micrograms/kg once per day (n = 5) given via 2-hour intravenous infusion for 7 days. Complete blood counts, differential, and platelet counts were obtained, and tibial bone marrow aspirate was performed on day 8. Neutrophil C3bi receptor expression was measured at 0 and 24 hours. GM-CSF levels were measured by a sandwich enzyme-linked immunosorbent assay at 2, 4, 6, 12, and 24 hours after the first dose of rhuGM-CSF. At all doses, rhuGM-CSF was well tolerated, and there was no evidence of grade III or IV toxicity. Within 48 hours of administration, there was a significant increase in the circulating absolute neutrophil count (ANC) at 5.0 micrograms/kg twice per day and 10.0 micrograms/kg once per day, which continued for at least 24 hours after discontinuation of rhuGM-CSF. When the ANC was normalized for each patient's first ANC, there was a significant increase in the ANC on days 6 and 7 at each dose level. By day 7, all tested doses of rhuGM- CSF resulted in an increase in the absolute monocyte count (AMC) compared with placebo-treated neonates. In those receiving rhuGM-CSF 5.0 micrograms/kg twice per day, there was additionally a significant increase in the day 7 and 8 platelet count. Tibial bone marrow aspirates demonstrated a significant increase in the bone marrow neutrophil storage pool (BM NSP) at 5.0 micrograms/kg twice per day and 10.0 micrograms/kg once per day. Neutrophil C3bi receptor expression was significantly increased 24 hours after the first dose of rhuGM-CSF at 5.0 micrograms/kg once per day. The elimination half-life (T1/2) of rhuGM-CSF was 1.4 +/- 0.8 to 3.9 +/- 2.8 hours.(ABSTRACT TRUNCATED AT 400 WORDS)     

Efficacy of delayed administration of post-chemotherapy granulocyte colony-stimulating factor: evidence from murine studies of bone marrow cell kinetics

The optimal schedule of post-chemotherapy granulocyte colony-stimulating factor (G-CSF) administration has not been determined. G-CSF is customarily started 24 hours after chemotherapy; however, clinical data demonstrated that delaying G-CSF until 5 days after completion of chemotherapy has not resulted in a longer duration of neutropenia. Here, we examined the optimal timing of post-chemotherapy G-CSF administration in a mouse model, to show that delayed administration does not postpone the appearance of mature granulocytes in the peripheral blood. We also investigated the mechanism of decreased efficacy of the early G-CSF application after chemotherapy by characterizing the changes in bone marrow cellular composition. To our knowledge, we demonstrate for the first time, that early after chemotherapy, the bone marrow is predominantly composed of mature residual granulocytes and very few progenitors and precursors, on which G-CSF would act to generate granulocytes. The point when immature progenitors reappear does not occur in murine bone marrow until 48 hours after a single dose of cyclophosphamide. Our results indicate that the bone marrow cellular composition early after discontinuation of chemotherapy is not optimal for G-CSF action on acceleration of myeloid recovery. Given the high cost of G-CSF prophylaxis, its delayed administration may potentially result in substantial economic benefits.     

Bilineage response in refractory aplastic anemia patients following long-term administration of recombinant human granulocyte colony-stimulating factor.

5 patients with refractory aplastic anemia (AA) received long-term administration (2-11 + months) of recombinant human G-CSF (rhG-CSF) in doses from 250-500 micrograms/body/day by intravenous infusion or 75-300 micrograms/body/d by subcutaneous injection. All 5 evaluable patients showed a substantial increase in absolute neutrophil count (ANC) with a recovery of myeloid components in the bone marrow after 1 to 2 months of treatment. Interestingly, 2 out of the 5 patients showed a dramatic improvement in severe anemia after 2 to 4 months of treatment accompanying a recovery of erythroid components in the bone marrow. In addition, there was no serious infection before or during therapy. Long-term administration of rhG-CSF was well tolerated because of its minimal toxicity. Clonal assay revealed a recovery of myeloid progenitors in all patients and a recovery of erythroid progenitors in 3 out of the 5 patients. These results suggest that long-term administration of rhG-CSF at least mobilizes residual myeloid as well as erythroid progenitor cells and induces a bilineage response in severe refractory AA.     

Myeloid cell kinetics in mice treated with recombinant interleukin-3, granulocyte colony-stimulating factor (CSF), or granulocyte-macrophage CSF in vivo.

Myeloid cell kinetics in mice treated with pure hematopoietic growth factors have been investigated using tritiated thymidine labeling and autoradiography. Mice were injected subcutaneously with 125 micrograms/kg granulocyte colony-stimulating factor (G-CSF) (in some cases 5 micrograms/kg), or 10 micrograms/kg of granulocyte-macrophage CSF (GM-CSF), or interleukin-3 (IL-3) every 12 hours for 84 hours. 3HTdR labeling was performed in vivo after 3 days of treatment. G-CSF increased the peripheral neutrophil count 14-fold and increased the proportion and proliferation rate of neutrophilic cells in the marrow, suppressing erythropoiesis at the same time. Newly produced mature cells were released into the circulation within 24 hours of labeling, compared with a normal appearance time of about 96 hours. By contrast, GM-CSF and IL-3 had little effect on either marrow cell kinetics or on the rate of release of mature cells, although GM-CSF did stimulate a 50% increase in peripheral neutrophils. Monocyte production was also increased about eightfold by G-CSF and 1.5-fold by GM-CSF, but their peak release was only slightly accelerated. While the peripheral half-lives of the neutrophilic granulocytes were normal, those of the monocytes were dramatically reduced, perhaps due to sequestration in the tissues for functional purposes. The stimulated monocyte production in the case of G-CSF required an additional five cell cycles, a level that might have repercussions on the progenitor compartments.     

Clinical application of recombinant human colony-stimulating factor (rhG-CSF) in bone marrow transplantation]

Human granulocyte colony-stimulating factor (G-CSF) specifically stimulates granulocyte production and enhances functions of mature granulocytes. It also proliferates myeloid leukemic cells. The molecule was purified and molecularly cloned in 1986. In this study, 51 patients received single daily injections with recombinant human (rh) G-CSF (2-20 micrograms/kg) after bone marrow transplantation were compared with control 36 patients. Blood neurophilic recovery was accelerated remarkably by rhG-CSF administration without any adverse effects including delay in reticulocyte and platelet recoveries. Furthermore, the duration for the management in laminar airflow room and febrile days greater than or equal to 38 degrees C were shortened in rhGS-CSF treated group. In 6 patients with myeloid leukemia in relapse, we also tested the effects of the rhG-CSF-combined conditioning regimen. In all the patients tested, the leukemic cells responded to rhG-CSF. Clinically no relapse was observed and 3 patients are well on day 224-357 for the time being. These findings indicate that rhG-CSF is very useful in bone marrow transplantation.     

Evidence for a novel in vivo control mechanism of granulopoiesis: mature cell-related control of a regulatory growth factor

As part of phase I/II clinical trials of granulocyte colony-stimulating factor (G-CSF), the pharmacokinetics was studied. To determine the optimal way of abrogating the neutropenia caused by melphalan, patients received G-CSF and melphalan on several schedules. The half-life (t 1/2) of elimination of G-CSF was in the range 1.3 to 4.2 hours and was prolonged at higher doses, suggesting that one clearance mechanism becomes saturated at doses greater than 10 micrograms/kg, When a continuous subcutaneous (SC) infusion was administered for five days, a rapid reduction in serum G-CSF levels occurred during the last two days of the infusion, indicating that an additional clearance mechanism was induced. When a continuous infusion of G-CSF was administered after melphalan, serum G-CSF levels remained constant for a longer period of time but did decrease during the second phase of a biphasic neutrophil response. In another clinical trial, G-CSF was administered after high-dose chemotherapy and autologous bone marrow transplantation (ABMT). In these patients, the G-CSF levels did not decrease while the patients were neutropenic. These results show that increased neutrophil levels are associated with increased clearance of G-CSF. This may be one of the negative feedback mechanisms involved in maintaining neutrophil homeostasis in normal and disease states.     

Changes in neutrophil counts and functions after the administration of recombinant human granulocyte colony-stimulating factor in a patient with chronic idiopathic neutropenia]

The cause of chronic idiopathic neutropenia (CIN) is unknown. Recently recombinant human granulocyte colony-stimulating factor (rhG-CSF) has been purified. Many studies of effects of rhG-CSF on the patients with neutropenia have been undertaken. We examined changes in neutrophil counts and functions after the administration of rhG-CSF in a patient with CIN. Six hours after the intravenous administration of 40 micrograms of rhG-CSF, neutrophil counts were raised from 90 to 1570/microliters, and the increased neutrophils functioned normally; chemotaxis, phagocytosis and O2(-) generation. It is suggested that rhG-CSF is beneficial for the treatment of infection in patients with CIN.     

A comparison of hematopoiesis in normal and splenectomized mice treated with granulocyte colony-stimulating factor.

Recombinant human granulocyte colony-stimulating factor (rhG-CSF) induces leukocytosis in vivo in both intact and splenectomized mice. Full dose response data showed a plateau in this effect at doses over 500 micrograms rhG-CSF/kg body weight/d in intact mice. The effect is magnified in splenectomized mice, where leukocyte numbers reach 100 x 10(6) mL after 4 days' treatment at 250 micrograms/kg/d. Further hematopoietic precursor populations are also affected in both marrow and the spleen; in general, marrow parameters were depressed, while splenic populations were enlarged. In splenectomized mice, both blood-borne stem cells were enhanced, and foci of extramedullary hematopoiesis were enlarged in addition to the effects seen in intact mice. In the marrow of splenectomized and intact mice treated with a high dose of G-CSF, erythroid suppression in the marrow was confirmed with radioactive iron. Our studies confirm and extend previous work on the mode of action of G-CSF, and indicate that side effects of high dose G-CSF therapy might include erythroid suppression in the bone marrow.     

A phase II trial of recombinant human granulocyte colony-stimulating factor in the myelodysplastic syndromes

We conducted a phase II study of the intravenous administration of a glycosylated recombinant human granulocyte colony-stimulating factor (rhG-CSF) for 7-14 d in 41 patients with the myelodysplastic syndromes (MDS). Administration of rhG-CSF elicited striking rises in both leucocyte and neutrophil counts in the majority of the patients irrespective of the FAB subtypes of MDS. The rises in neutrophil counts were dose dependent and 5 micrograms/kg/d of rhG-CSF yielded approximately an 8-fold increase in neutrophil counts. Leucocytes and neutrophil counts started to increase shortly after the first injection of 5 micrograms/kg, was maintained at significantly elevated levels during 14 d of treatment, and returned to the pretreatment levels within several days following discontinuation of rhG-CSF. The action of rhG-CSF was specific for neutrophils since leucocytosis was due exclusively to neutrophilic increase associated with an increased marrow myeloid maturation. There were no consistent changes in the monocyte, eosinophil, lymphocyte, platelet or reticulocyte counts. After treatment, the percentage of marrow blast cells was reduced in eight of 13 evaluable patients with refractory anaemia with an excess of blasts (RAEB) or RAEB in transformation (RAEB-t). No patients developed acute leukaemia during the treatment or in the immediate follow-up period. The treatment was well tolerated with only minimal toxicity. The results suggest that rhG-CSF is a safe and effective way to promptly improve neutropenia in MDS patients.     

Anorexia nervosa with neutropenia--response of neutrophils to G-CSF]

A 50-year-old woman with anorexia nervosa was admitted for evaluation of neutropenia (WBC 1,600/microliters). Her bone marrow was gelatinous, and myeloid cells had decreased. Homogeneous substance deposited in the marrow, stained by alcian blue (pH 2.5), indicative of acid mucopolysaccharides. CFU-G and CFU-GM were decreased in number and myeloid pool in the bone marrow also decreased. Anti-neutrophilic antibody was negative. Neutropenia may be related to myeloid hypoplasia, due to increase of acid mucopolysaccharides replacing adipose cells in the bone marrow under long-term mal-nutritional state. Neutrophils markedly increased by administration of rhG-CSF 5.0 micrograms/kg/day for 14 days without the first peak. Serum G-CSF level did not increase (less than 60 pg/ml). It is effective to administer G-CSF to anorexia nervosa with neutropenia.     

Hematopoietic progenitors in cyclic neutropenia: effect of granulocyte colony-stimulating factor in vivo.

The number and growth factor requirements of committed progenitor cells (colony-forming units-granulocyte/macrophage and burst-forming units-erythroid) in three patients with cyclic neutropenia (two congenital, one acquired) were studied before and during therapy with recombinant human granulocyte colony-stimulating factor (G-CSF; 3 to 10 micrograms/kg/d). When the patients with congenital disease were treated with G-CSF, the cycling of blood cells persisted, but the cycle length was shortened from 21 days to 14 days, and the amplitude of variations in blood counts increased. There was a parallel shortening of the cycle and increase of the amplitude of variations (from two- to three-fold to 10- to 100-fold) in the number of both types of circulating progenitor cells in these two patients. In the patient with acquired cyclic neutropenia, cycling of both blood cells and progenitors could not be seen. In cultures deprived of fetal bovine serum, erythroid and myeloid bone marrow progenitor cells from untreated patients and from normals differed in growth factor responsiveness. As examples, maximal growth of granulocyte/macrophage (GM) colonies was induced by granulocyte/macrophage (GM)-CSF plus G-CSF in the patients, whereas a combination of GM-CSF, G-CSF and interleukin-3 (IL-3) was required in the normals, and erythropoietin alone induced fourfold more erythroid bursts from cyclic neutropenic patients than from normal donors (46% versus 11% of the maximal colony number, respectively). The growth factor responsiveness of marrow progenitor cells slightly changed during the treatment toward the values observed with normal progenitors. These results indicate that treatment with G-CSF not only ameliorated the neutropenia, but also increased the amplitude and the frequency of oscillation of circulating progenitor cell numbers. These data are consistent with the hypothesis that G-CSF therapy affects the proliferation of the hematopoietic stem cell.     

Abnormal responses of myeloid progenitor cells to recombinant human colony-stimulating factors in congenital neutropenia.

The effects of recombinant human interleukin-3 (IL-3) and recombinant human granulocyte colony-stimulating factor (G-CSF) on the growth of myeloid progenitor cells (CFU-C) in semisolid agar culture were studied in two patients with Kostmann-type congenital neutropenia. CFU-C growth in bone marrow cells from patients was significantly reduced in response to various concentrations of either IL-3 or G-CSF alone, compared with that from normal subjects. There was no inhibitory effect of bone marrow cells from patients on normal CFU-C formation supported by IL-3 or G-CSF. However, the simultaneous stimulation with IL-3 and G-CSF induced the increase of CFU-C formation in patients with congenital neutropenia. Furthermore, CFU-C growth in both patients was supported when bone marrow cells were preincubated with IL-3 in liquid culture followed by the stimulation with G-CSF in semisolid agar culture. In contrast, that was not supported by the preincubation with G-CSF and the subsequent stimulation with IL-3. This evidence suggests that the hematopoietic progenitor cells in patients with congenital neutropenia have the potential for developing CFU-C in the combined stimulation with IL-3 and G-CSF, and that this growth may be dependent on the priming of IL-3 followed by the stimulation with G-CSF. The level of mature neutrophils in peripheral blood was not fully restored to normal levels by the daily administration of G-CSF in doses of 100 to 200 micrograms/m2 of body surface area for 20 to 25 days in both patients. These observations raise the possibility that the combination of IL-3 and G-CSF might have a potential role for the increase of neutrophil counts in patients with congenital neutropenia.