Hematopoietic growth-promoting effects of natural killer cells in mice in response to in vivo administration of human interleukin-2.

BACKGROUND: Interleukin-2 (IL-2) is currently being evaluated as an anti-neoplastic agent because of its stimulatory effects on the immune system. However, little is known about the effects of systemic IL-2 administration on hematopoietic parameters. PURPOSE: Recombinant human IL-2 (rHuIL-2) was administered to mice to evaluate its in vivo hematopoietic effects. The mechanism underlying the effects of rHuIL-2 administration was also determined. METHODS: Mice were given 5 x 10(4) IU of rHuIL-2 for 5 days, and their hematopoietic progenitor cell status was determined as measured by growth in soft agar. Antiserum to natural killer (NK) cell-specific markers was used to ascertain if NK cells were responsible for the hematopoietic effects of rHuIL-2. NK cells were purified and cultured in vitro with rHuIL-2 and then adoptively transferred into syngeneic mice to determine the effects on hematopoiesis. RESULTS: Treatment of mice with rHuIL-2 resulted in a significant increase in bone marrow and splenic hematopoietic progenitor cell content. Mice with severe combined immune deficiency (SCID), which lack T cells and B cells yet have NK cells, also responded to the myelostimulatory effects of rHuIL-2. However, removal of NK cells from the SCID mice with antiserum to NK cell-specific markers abrogated the myelostimulatory properties of rHuIL-2. Adoptive transfer of NK cells that were propagated in vitro with rHuIL-2 into mice also resulted in an increase in splenic hematopoietic progenitor cells. CONCLUSIONS: rHuIL-2 exerts significant myelostimulatory effects after in vivo administration, and NK cells are responsible for at least some of these effects. IMPLICATIONS: These results suggest that NK cells and rHuIL-2 may be of use clinically to promote hematopoiesis after bone marrow transplantation or in the face of other myelotoxic therapy in the treatment of cancer.     

Thrombopoietin, interleukin-11, and early-acting megakaryocyte growth factors in human myeloid leukemia cells.

In this study we report our data on effects of early-acting megakaryocyte growth factors, particularly the c-mpl ligand also known as thrombopoietin (TPO) and interleukin-11 (IL-11), on cell proliferation and apoptosis (Apo) of primary acute myeloid leukemia (AML) cells. A proliferative response to TPO was noticed in the majority of AML samples (17/19) with an average increase of S-phase cells from 7.8% +/- 1.5 to 14.5% +/- 2.1 (p=0.0006). Resulting cell cycle activation did not always correlate with expression of the c-mpl receptor, although it was coupled, in the majority of samples, by an average decrease of apoptotic cells from 13% +/- 0.7 to 8.8% +/- 1.8 (p=0.05). Clonogenic cell growth (CFU-L) was confirmed in 5/17 of the samples with a mean colony number of 21.4 +/- 9.6 x 10(5) cells plated. Conversely, effects of IL-11 on AML cells demonstrated that cell cycle changes (recruitment from G0 to S phase) were promoted only in a minority of samples (2/14) and there was little, if any, effect on CFU-L growth (mean colony number=17.5 +/- 9.5) or Apo (from 13% +/- 0.7 to 13.3 +/- 1.9). Combination of TPO with IL-11 induced a slight increase of clonogenic cell growth, while the addition of IL-3 or SCF to the c-mpl ligand significantly raised the mean colony numbers up to 119.2 +/- 68.3 and 52.9 +/- 22.1 x 10(5) cells plated, respectively. In summary, TPO shows activity on AML cells by stimulating their proliferation in a significant proportion of cases and generally protecting the majority of AML blast cells from induction of Apo. Conversely, IL-11 exerts little effect on the cell cycle activation and Apo. These data help to understand regulation of myeloid leukemia cell growth and should be considered in the clinical use of early-acting megakaryocyte growth factors in acute leukemia.     

Recombinant interleukin 2 toxicity, pharmacokinetics, and immunomodulatory effects in a phase I trial

Twenty-two patients with refractory malignancies were treated with four escalating weekly doses of recombinant interleukin 2 (IL2), given either i.v. by 2- or 24-h infusion, or s.c. A 1-wk washout period between each dose of IL2 was provided for the evaluation for pharmacokinetic and immunomodulatory effects. The maximum i.v. dose was 30 X 10(6) units; the dose-limiting toxicities were fever, flu-like symptoms, and hypotension. The maximum s.c. dose was 3 X 10(6) because of volume limitations with s.c. injection. No tumor regression was seen. During infusions of 3 X 10(6) units over 2 h or 24 h, serum IL2 levels greater than or equal to 223 units/ml or 16 units/ml were maintained, respectively; with s.c. injection of 3 X 10(6) units, levels greater than 20 units/ml were maintained for 9 h. Marked lymphopenia was observed 24 h after the initiation of IL2 doses which was completely reversible when measured prior to the next dose. The lymphopenia was nonselective; T- and B-lymphocytes decreased in an IL2 dose-dependent manner, without consistent change in the OKT4:OKT8 ratio. No change was detected in monocyte expression of HLA-Dr or T-cell expression of the IL2 receptor. The in vitro generation of lymphokine-activated killer cytotoxicity decreased sharply and transiently shortly after i.v. doses. Mitogen responsiveness, delayed-type hypersensitivity, natural killer cytotoxicity, and mixed-lymphocyte reactivity were unchanged or decreased transiently shortly after IL2 doses. These studies help define the bioavailability of IL2 by i.v. or s.c. routes, and they will aid in the design of studies utilizing daily doses of IL2.     

Arsenic trioxide and breast cancer: analysis of the apoptotic,differentiative and immunomodulatory effects

Arsenic trioxide (As₂O₃) is used clinically to treat acute promyelocytic leukemia and has activity in vitro against several solid tumour cell lines, where induction of differentiation and apoptosis are the prime effects. To investigate the potential therapeutic application of As₂O₃ to breast cancer, we analysed the effects of As₂O₃ on the growth of four human breast cancer cell lines: MCF7, MDA-MB-231, T-47D and BT-20. Cells were cultured in 0.5, 2 and 5M As₂O₃, a range of pharmacologically achievable concentrations of As₂O₃. At 2M, As₂O₃ rapidly induced cell death by apoptosis in MCF7 and MDA-MB-231 while T-47D and BT-20 were partially resistant. At 0.5M, As₂O₃ was subapoptotic but induced features of differentiation consisting in upregulation of ICAM-1 (CD54), a marker of mammary epithelial differentiation, and cell cultures appeared morphologically more orga-nized. Furthermore, we demonstrate by standard cytotoxicity assays that As₂O₃ treatment can augment breast cancer cell lysis by lymphokine-activated killer cells and demonstrate an important role of the ICAM-1/LFA-1 interaction in this process. This additional activity of As₂O₃ could translate into improved antitumour immunosurveillance in vivo. In conclusion, As₂O₃ induced varying degrees of differentiation, apoptosis and lysis in these model cell lines, and may be a promising adjuvant to current treatments of breast cancer by virtue of its triple apoptotic, differentiative and immunomodulatory effects.     

Cancer incidence in multiple sclerosis and effects of immunomodulatory treatments

Multiple sclerosis (MS) has been linked to reduced rates of cancer prior to the era of immunomodulating treatments. We assessed the incidence of cancer in a cohort of 1338 MS patients and evaluated the effect of exposure to immunomodulatory treatment. Cancer incidence in the MS population was compared with the expected age- and gender-matched incidence rates in the Israeli population for the period 1960–2003. Time-dependant Cox model analysis was used to estimate hazard ratios for glatiramer acetate, -interferons (1a and 1-b) and intravenous immunoglobulins (IVIg). Among 892 female MS patients, 15 (1.7%) developed breast cancer, and 31 (3.5%) developed cancers of any type. Seventeen of 446 (3.8%) male MS patients developed cancer. The standardized incidence ratios (SIRs) computed until the time of first immunomodulatory treatment were 0.60 (95% CI, 0.38–0.92, p = 0.02) for all female cancer, and 1.11 (95% CI, 0.64–1.91) for all male cancer. Time-dependent covariate analyses for female breast cancer yielded a relative risk for glatiramer acetate of 3.10 (95% CI, 0.86–11.1) and 0.52 (95% CI, 0.07–4.05) for -interferons. For IVIg, the analyses were uninformative. Our findings indicate that cancer incidence is significantly lower in female MS patients than in the general population. Female MS patients treated with glatiramer acetate showed an elevated rate of breast cancer and all MS patients treated with -interferons showed an elevated risk of non-breast cancers though not statistically significant (p = 0.122 and 0.072, respectively). Further study is needed to assess possible associations between long-term exposure to the novel immunomodulatory treatments in MS and rate of caner.     

Silymarin and skin cancer prevention: anti-inflammatory, antioxidant and immunomodulatory effects (Review)

Several environmental and genetic factors are involved in skin cancer induction, however exposure to chemical carcinogens and solar ultraviolet (UV) radiation are primarily responsible for several skin diseases including skin cancer. Chronic exposure of solar UV radiation to the skin leads to basal cell and squamous cell carcinoma, and melanoma. Chemoprevention of skin cancer by consumption of naturally occurring botanicals appears a practical approach and therefore world-wide interest is considerably increasing to use these botanicals. Sunscreens are useful but their protection is not ideal because of inadequate use, incomplete spectral protection and toxicity. Silymarin, a plant flavonoid isolated from the seeds of milk thistle (Silybum marianum), has been shown to have chemopreventive effects against chemical carcinogenesis as well as photocarcinogenesis in various animal tumor models. Topical treatment of silymarin inhibited 7,12-dimethylbenz(a)anthracene-initiated and several tumor promoters, like 12-O-tetradecanoylphorbol-13-acetate, mezerein, benzoyal peroxide and okadaic acid, induced skin carcinogenesis in mouse models. Similarly, silymarin also prevented UVB-induced skin carcinogenesis. Wide range of in vivo mechanistic studies indicated that silymarin possesses antioxidant, anti-inflammatory and immunomodulatory properties which may lead to the prevention of skin cancer in in vivo animal models. The available experimental information suggests that silymarin is a promising chemopreventive and pharmacologically safe agent which can be exploited or tested against skin cancer in human system. Moreover, silymarin may favorably supplement sunscreen protection and provide additional anti-photocarcinogenic protection.     

Repetitive weekly cycles of interleukin-2. II. Clinical and immunologic effects of dose, schedule, and addition of indomethacin

Clinical trials with high doses of interleukin 2 (IL-2) have shown antitumor responses, but many of the patients have experienced severe and occasionally life-threatening toxic effects. Preclinical studies indicate that modifications in IL-2 dose, route, and schedule can influence both immune activation and antitumor effects. This study evaluated the clinical tolerance to and immunologic modifications induced by four repetitive weekly cycles of IL-2, with two dose levels (1 X 10(6) and 3 X 10(6) U/m2 per day) of IL-2 and three different daily administration schedules [bolus, continuous, or combined (bolus and continuous)], with and without indomethacin treatment. Patients in all treatment groups experienced acceptable, non-life-threatening toxic effects and immune system stimulation characterized by rebound lymphocytosis with increased numbers of natural killer and lymphokine-activated killer cells and enhanced direct cytolytic function. These immune changes were significantly enhanced by the repetition of IL-2 cycles beyond the first week of therapy. At an IL-2 dose of 3 X 10(6) U/m2 per day, bolus IL-2 was less immunostimulatory than continuous-infusion IL-2. The combined regimen (with half of each daily dose given as a bolus and half as a 24-hr infusion) was as stimulatory as continuous-infusion IL-2 and also induced antitumor effects. Finally, the addition of indomethacin to this regimen did not significantly modify in vitro or in vivo immune response parameters but appeared to worsen the systemic toxic effects of renal dysfunction and capillary leakage. These results suggest that continuous or combined infusion of IL-2 at 3 X 10(6) U/m2 per day on this schedule should be considered for further testing in phase II trials or in combination with other therapeutic modalities.     

Pilot study of low-dose interleukin-11 in patients with bone marrow failure.

PURPOSE: Interleukin-11 (IL-11) is a thrombopoietic cytokine that attenuates postchemotherapy thrombocytopenia at doses of 50 microg/kg/d subcutaneously. Very little is known about the activity of IL-11 in patients with bone marrow failure states. PATIENTS AND METHODS: Our preliminary experience with IL-11 at doses of 50 microg/kg/d suggested that patients with bone marrow failure developed significant peripheral and pulmonary edema after the prolonged dosing necessary for treating these conditions. We, therefore, initiated a study of low-dose IL-11 (starting dose, 10 microg/kg/d). RESULTS: Sixteen patients were assessable for response. Six patients had diploid cytogenetics; the others had a variety of chromosomal abnormalities. Six (38%) of 16 patients showed a platelet response to IL-11, and two had a multilineage response (to IL-11 alone, n = 1; to IL-11 plus G-CSF and erythropoietin, n = 1). The median increase in peak platelet counts was 95 x 10(9)/L above baseline in the responders (range, increase of 55 x 10(9)/L to 130 x 10(9)/L above baseline). Responders included five of 11 patients with myelodysplasia and one of four patients with aplastic anemia. Response durations were 12, 13, 14+, 25, 30, and 30+ weeks. Side effects of IL-11 were mild (peripheral edema, n = 7; conjunctival injection, n = 7; myalgia, n = 1; all grade 1). Seven patients had no side effects. CONCLUSION: Our pilot study suggests that administration of low-dose IL-11 (10 microg/kg/d) can raise platelet counts without significant toxicity in selected thrombocytopenic patients with bone marrow failure.     

Novel thalidomide analogues display anti-angiogenic activity independently of immunomodulatory effects

The anti-tumour effects of thalidomide have been associated with its anti-angiogenic properties. Second generation thalidomide analogues are distinct compounds with enhanced therapeutic potential. Although these compounds are beginning to enter trials for the treatment of cancer there is very little information regarding the anti-angiogenic activity of these clinically relevant compounds. Furthermore, it is not known how the various immunomodulatory activities of these compounds relate to anti-angiogenic activity. In this study we assessed the anti-angiogenic activity of compounds from both IMiD and SelCID classes of analogues using a novel in vitro multicellular human assay system and the established rat aorta assay. Our results show that both the IMiDs and SelCIDs tested are significantly more potent than thalidomide. The anti-angiogenic potency of the analogues was not related to inhibition of endothelial cell proliferation, nor their TNF-alpha/PDE type 4 inhibitory properties. However, anti-migratory effects in vitro and inhibition of tumour growth in vivo was observed with the analogue IMiD-1 (clinically known as REVIMID). Our results show that anti-angiogenic activity spans both currently defined classes of thalidomide analogue and is not related to their previously described immunomodulatory properties. Identification of the differential effects of these compounds will enable targeting of such compounds into the appropriate clinical setting.     

Immunopharmacological and antitumor effects of second-generation immunomodulatory oligonucleotides containing synthetic CpR motifs

Bacterial and synthetic DNAs containing CpG dinucleotides in specific sequence contexts (CpG DNA) activate the vertebrate immune system and produce potent Th1 immune responses. Recently, we reported immunomodulatory oligonucleotides (IMOs) containing 3'-3'-attached novel structures (immunomers) and synthetic immunostimulatory CpR (R=2'-deoxy-7-deazguanosine) dinucleotides show potent stimulatory activity with distinct cytokine secretion profiles. In the present study, we evaluated in vivo immunopharmacological and antitumor properties of second-generation immunomodulatory oligonucleotides (IMOs) either alone or in combination with chemotherapeutic agents. Repeated peritumoral administration of IMOs at 1 mg/kg to mice bearing established subcutaneous CT26 colon tumor or B16.F0 melanoma resulted in complete regression or strong inhibition of tumor growth. Direct peritoneal injection of IMOs at 2.5 mg/kg to mice bearing peritoneally implanted ascites CT26 or B16.F0 tumors completely eradicated or inhibited tumor growth. Treatment of mice bearing beta-gal expressing CT26.CL25 tumor with IMOs resulted in a significant tumor-specific CTL responses compared with treatment with a control non-CpG DNA or PBS. These responses correlated with IFN-gamma, but not IL-4 secreted in IMO, treated mice. A 5-fold increase in beta-gal specific IgG2a antibodies was found in mice, significantly increasing the IgG2a/IgG1 ratio. IMOs showed similar antitumor activity in both wt and IL-6 knockout (ko) C57BL/6 mice but failed to elicit activity in IL-12 ko C57BL/6 mice. Tumor-free mice from the IMO treatment group rejected the same tumor cell rechallenge, suggesting an adaptive immune response against these cells. Moreover, na?ve mice quickly developed specific antitumor response without IMO treatment following adoptive transfer of splenocytes obtained from tumor free mice from the IMO treated group. Additionally, the co-administration of IMOs with chemotherapeutic agents, docetaxel and doxorubicin, resulted in synergistic antitumor effects in both B16.F0 melanoma and 4T1 breast carcinoma models. These results demonstrate potent antitumor activity of second-generation IMO compounds containing a synthetic CpR stimulatory motif in a broad spectrum of tumor models through induction of strong Th1 immune responses. IMO treatment resulted in the development of tumor-specific memory immune responses. No treatment-related toxicity was observed in mice at the doses and treatment schedules studied.     

Anti-tumor effects of interleukin-4 and interleukin-5 against mouse B cell lymphoma and possible mechanisms of their action.

We investigated the anti-tumor effects of recombinant mouse interleukin (IL)-4 and IL-5 by using a transplantable B cell lymphoma 38C13 cell line as a model. Daily local administration of either IL-4 or IL-5 produced moderate but significant inhibition of the rate of local tumor growth and prolongation of mean survival time (MST) in syngeneic C3H/HeJ mice; these anti-tumor effects appeared to plateau at low doses. Histopathologic and immuno-histochemical examination revealed necrotic changes in the cytokine-treated tumors, associated with infiltration of inflammatory cells such as eosinophils, macrophages, and lymphocytes. The infiltrating lymphocytes were found to be Thy-1.2+ T cells. To elucidate the importance of T cells, the rate of tumor growth and the MSTs were compared between athymic T cell-deficient BALB/c nude mice and immunocompetent C3H/HeJ mice. In the nude mice the transplanted tumor grew more rapidly and the MST was shorter than in the normal mice, suggesting a significant contribution of infiltrating T cells in the anti-tumor effects of the interleukins. Lastly, in vitro, growth inhibition of the 38C13 cells was observed in a dose-dependent manner at relatively high concentrations of either cytokine. Therefore, we conclude that both IL-4 and IL-5 have moderate anti-tumor effects against 38C13 B cell lymphoma both in vivo and in vitro, and that the observed in vivo anti-tumor effects are probably mediated both by tumoristatic action of infiltrating cells, such as eosinophils, macrophages and T lymphocytes, and by direct anti-proliferative action of the recombinant cytokines.     

Hematopoietic growth factors and leukemia.

Hematopoietic growth factors, particularly granulocyte colony-stimulating factor and granulocyte-macrophage colony-stimulating factor, can be used as supportive agents to enhance bone marrow recovery after the administration of myelosuppressive chemotherapy given for nonmyeloid neoplasms. The use of these agents in the treatment of acute myeloid leukemia and myelodysplastic syndrome poses novel opportunities and challenges due to their direct effects on the neoplastic cells, which represent the transformed counterparts of normal hematopoietic stem cells. The interaction between hematopoietic growth factors and leukemic progenitor cells bearing a specific receptor for a given agent would be expected to result in proliferation, although maturation induction could occur. Hematopoietic growth factors have been employed as primary differentiating agents in myelodysplastic syndrome and as supportive agents after chemotherapy in acute myeloid leukemia. In either case, close monitoring for evidence of leukemic stimulation is required. Alternatively, pretreatment with colony-stimulating factors could induce cell cycling, thereby making the leukemic cells more susceptible to S-phase-specific chemotherapeutic agents, such as cytarabine.     

重组人白细胞介素-11在16例急性髓系白血病化疗中的应用

 目的　观察重组人白细胞介素-11（rhIL-11）对急性髓系白血病化疗所致血小板减少的疗效和患者不良反应。方法　新诊断的16例急性髓系白血病患者分别接受DA 或TA 化疗方案,采用随机空白对照、自身交叉的研究方法,观察rhIL-11对化疗后血小板下降恢复的影响。化疗结束后24 h开始使用rhIL-11,皮下注射,连用7～14 d,血小板稳定在30×109／L且无明显的出血倾向时停药。结果　rhIL-11明显提高患者化疗后血小板计数,减少输注血小板的次数,且rhIL-11注射后患者不良反应轻微,主要表现为乏力、肌肉酸胀或低热等。结论　rhIL-11可有效促进化疗后外周血小板恢复,缩短血小板减少持续时间,减少输血相关并发症的发生,患者耐受性好。     

Phase I and immunomodulatory study of a muramyl peptide, muramyl tripeptide phosphatidylethanolamine.

Muramyl tripeptide phosphatidylethanolamine (MTP-PE; CGP 19835A from Ciba Geigy) is a synthetic muramyl tripeptide structurally related to bacterial cell wall constituents. MTP-PE activates monocytes in vitro to a tumoricidal state and has in vivo antitumor effects in animal models. We studied the toxicity and immunomodulatory effects of once weekly i.v. administration of liposomal-encapsulated MTP-PE for 8 weeks in 27 patients with advanced malignancies. Doses ranged from 0.1 to 2.7 mg/m2. No major tumor responses were seen; 11 patients had stable disease after 8 weeks of therapy and 3 continued on maintenance therapy because of minor tumor regressions and/or clinical improvement. MTP-PE at these doses was well tolerated. Shaking chills and fevers were the most common toxicities and occurred at all dose levels. There was no treatment-induced loss of performance status. Immunomodulatory studies revealed evidence of a biological effect on monocytes. C-reactive protein levels rose in the majority of patients with end-of-treatment values 2 to 10 times higher than baseline. Serum neopterin levels were consistently increased 24 h after MTP-PE administration and significant decreases in expression of two different types of Fc receptors on peripheral blood monocytes were noted 6 h after treatment. Although no major tumor responses were seen in this group of patients with advanced malignancies, MTP-PE was well tolerated and exerted biological effects on monocytes. Serum neopterin levels may be a useful marker for the biological effects of MTP-PE.     

The biological basis for the attenuation of mucositis: the example of interleukin-11.

Oral mucositis is common, painful, dose-limiting toxicity of drug and radiation therapy for cancer. In granulocytopenic patients, the ulcerations which accompany mucositis are frequent portals of entry for indigenous oral bacteria often leading to bacteremias or sepsis. The complexity of mucositis as a biological process has only recently been appreciated. The condition appears to represent a sequential interaction of the oral mucosal cells and tissues, pro-inflammatory cytokines, and local environmental factors in the mouth such as microorganisms and saliva. The recognition that the pathophysiology of mucositis is a multifactorial process has presented opportunities for intervention based on biological attenuation. Interleukin-11, a pleotropic cytokine, has a range of activities which is potentially relevant to mucositis. Consequently, it has been used successfully to modify the development, severity and course of mucositis in an animal model which closely mimics the equivalent human condition.     

Aggressive non-Hodgkin's lymphoma: concomitant evaluation of interleukin-2, soluble interleukin-2 receptor, interleukin-4, interleukin-6, interleukin-10 and correlation with outcome

The purpose of this study was to assess the prognostic value of a large panel of cytokines in aggressive non-Hodgkin's lymphoma (NHL) and to confront it to parameters of the International Prognostic Index (IPI). It investigated the concomitant determination of interleukin-2 (IL-2), soluble interleukin-2 receptor (sIL-2R), interleukin-4 (IL-4), interleukin-6 (IL-6) and interleukin-10 (IL-10) on a uniform population of 116 previously untreated patients. Commercially available enzyme-linked immunoassay kits were used for cytokines measurements. Results were correlated with complete remission (CR), overall survival (OS) and failure free survival (FFS). In univariate analysis, sIL-2R and IL-6 demonstrated prognostic significance for CR (p = 0.016 and p = 0.048), OS (p = 0.0011 and p = 0.0387) and FFS (p = 0.0001 and p = 0.0363), but multi-variate analysis failed to demonstrate an independent prognostic significance. In the intermediate group risk defined by IPI, patients presenting high level of sIL-2R or IL-6 demonstrated lower CR rate and survival than those with low level. In conclusion, sIL-2R and IL-6 serum levels are elevated in high grade NHL and are correlated to CR, OS and FFS, but this study did not support their independent prognostic value. However, sIL-2R and IL-6 measurements may improve risk assignment by IPI and allow a better prognostic evaluation of patients with intermediate prognosis NHL.     

The effects of interleukin-3, GM-CSF, and G-CSF on the growth kinetics of colony-forming cells in myelodysplastic syndromes.

In order to obtain more insight into the nature of the abnormal in vitro colony formation in myelodysplastic syndromes (MDS), we investigated the kinetics of the colony formation of 23 MDS cases in response to recombinant human interleukin-3 (IL-3), Granulocyte-macrophage colony-stimulating factor (GM-CSF), granulocyte colony-stimulating Factor (G-CSF), and giant cell tumor cell line conditioned medium (GCT-CM). The kinetics of GCT-CM-induced colony formation were comparable to that of G-CSF-induced colony growth, both in MDS and in normal bone marrow cultures. Colony formation was found to be delayed in MDS. The delay in colony formation was most apparent in the GCT-CM (G-CSF) responsive progenitor cell compartment. In MDS cases with clinical features of high risk disease, this delay was more pronounced as compared with low risk cases (7 and 3 days, respectively, in response to GCT-CM). The delay in colony formation was found to be caused by an increase in the time interval before progenitor cells had begun to divide. These results suggest that a prolongation of the time spent in G0 of myeloid progenitor cells in MDS may be the cause of the indolent in vitro colony formation observed in this disease.