UNIPROT:P04141 - FACTA Search

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Query: UNIPROT:P04141 (granulocyte-macrophage colony-stimulating factor)
6,790 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

In vivo, growth factors are currently investigated for their capacity to trigger leukemic stem cells into cycle and thus overcome kinetic drug resistance. In this study, the susceptibility of leukemic clonogenic cells to individual growth factors was related to cytosine-arabinoside (Ara-C) sensitivity. The effects of interleukin-3 (IL-3), granulocyte-macrophage colony-stimulating factor (G-CSF), granulocyte colony-stimulating factor (G-CSF), and combinations of these recombinant hematopoietic factors were tested on blast cells of nine acute myeloid leukemia (AML) patients. Growth factor responses were assessed in semi-solid clonogenic assay and in a 10-day liquid culture followed by clonogenic assay. Heterogeneity in growth factor response was observed in both test systems, resulting in a variable pattern for individual leukemias. In the majority of cases (six of nine) the response patterns in the semi-solid and liquid cultures were divergent. To test the Ara-C sensitivity, leukemic blasts were exposed in liquid to various concentrations of Ara-C in the absence and presence of preselected growth factors. After 10 days, the number of surviving leukemic colony-forming cells (CFU-L) was assessed. Exposure to Ara-C in the presence of optimal stimulatory factor(s) resulted in a 3- to 1000-fold increase of the Ara-C toxicity in seven patients. The Ara-C concentrations resulting in 50% inhibition of clonogenicity (ID50) were 0.48-123 x 10(-8) M Ara-C in the absence of stimulatory growth factors, versus only 0.12-0.40 x 10(-8) M Ara-C in the presence of these factors. In two patients, addition of one or more factors neither increased the number of CFU-L in liquid nor enhanced the Ara-C toxicity. Even in the absence of growth factors the ID50 values in these cases were as low as 0.20 and 0.28 x 10(-8) M Ara-C and in the same range as the ID50 values observed with maximum growth factor stimulation in the other seven patients. These results indicate that Ara-C cytotoxicity can be enhanced by individually selected, clonogenic cell growth-promoting hematopoietic factors.
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PMID:In vitro response of blasts to IL-3, GM-CSF, and G-CSF is different for individual AML patients: factors that stimulate leukemic clonogenic cells also enhance Ara-C cytotoxicity. 751 90

Stimulation of clonogenic acute myeloid leukemia (AML) cells by hematopoietic growth factors (HGF) is capable of enhancing cytosine-arabinoside (Ara-C) cytotoxicity in vitro. Until now it has not been known to what extent in vitro Ara-C cytotoxicity can be restored by HGF stimulation in samples from previously treated AML patients. Therefore, we studied the individual effects of the hematopoietic growth factors (HGF) granulocyte colony-stimulating factor (G-CSF), interleukin-3 (IL-3), and granulocyte-macrophage colony-stimulating factor (GM-CSF) stimulation on the Ara-C sensitivity of clonogenic leukemic cells from six patients with newly diagnosed AML. These results were directly compared with the outcome using AML samples from the same patients at relapse or primary refractory AML. In one patient, a sample of the second relapse was also studied. The results were expressed as IC50 values, and were used to calculate sensitivity ratios, defined as the ratio of the IC50 value with drug exposure alone and with HGF plus Ara-C. In AML samples treated with Ara-C alone and no HGF, IC50 values of Ara-C in relapsed/refractory AML were greater than IC50 values of AML cells at diagnosis. Addition of either HGF enhanced the Ara-C cytotoxicity for the relapsed samples significantly (for G-CSF p = 0.036, IL-3 p = 0.036, and for GM-CSF p = 0.036). The values of Ara-C sensitization of AML samples due to HGF at relapse did not significantly differ from those at diagnosis. However, enhancement of Ara-C cytotoxicity to AML progenitors by IL-3 or GM-CSF stimulation was significantly less in the cell specimens from AML recurrence patients as compared with the original diagnosis samples. In three AML samples at diagnosis and at their relapse, Ara-C incorporation into the DNA was determined. IL-3 stimulation enhanced Ara-C incorporation in all samples tested. Nevertheless, Ara-C incorporation in the relapsed samples was significantly less than that in the diagnosis samples of the same patients. A good correlation between Ara-C incorporation and bromodeoxyuridine (BrdU) incorporation was found. The results indicate that HGF stimulation in relapsed/refractory AML enhances Ara-C cytotoxicity, but not to the level that was observed with AML samples at diagnosis.
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PMID:Hematopoietic growth factor stimulation and cytarabine cytotoxicity in vitro: effects in untreated and relapsed or primary refractory acute myeloid leukemia cells. 752 89

Exposure to hemopoietic growth factors (HGFs) induces proliferation of clonogenic acute myeloid leukemia (AML) cells. Recruitment of quiescent, clonogenic blasts may improve the cytotoxic effects of cell-cycle-specific drugs like cytosine arabinoside (Ara-C). Because other studies have shown heterogeneous effects of HGF and Ara-C incubation, we analyzed the individual effects of granulocyte colony-stimulating factor (G-CSF), interleukin 3 (IL-3), and granulocyte-macrophage colony-stimulating factor (GM-CSF) combined with an S-phase and non-phase specific cytostatic agent on clonogenic blasts of 14 newly diagnosed AML patients under standardized, serum-free conditions. AML cells were incubated for 24 hours with titrated concentrations of Ara-C (0.01, 0.1, 1 microM) or mafosfamide (0, 1.0, 10, or 20 micrograms/ml) following preincubation for 48 hours with or without G-CSF, IL-3, or GM-CSF, starting at 24 hours prior to chemotherapy exposure. AML colony-forming cells (AML-CFU) were then determined in semi-solid culture in the presence of the same growth factor. The results showed significantly enhanced cytotoxicity of Ara-C to AML-CFU following stimulation by G-CSF (p < 0.002 at 0.01 microM, p < 0.002 at 0.1 microM, and p < 0.01 at 1 microM Ara-C), IL-3 (p < 0.002 at 0.01 microM, p = 0.001 at 0.1 microM, p < 0.01 at 1 microM, Ara-C), and GM-CSF (p = 0.01 at 0.01 microM, p < 0.01 at 0.1 microM, and p < 0.002 at 1 microM Ara-C). A moderate but significant enhancement of mafosfamide cytotoxicity by HGF was also observed (p < 0.05 at 1.0 microgram/ml mafosfamide by IL-3 and GM-CSF and p < 0.05 at 10 micrograms/ml mafosfamide by GM-CSF). Ara-C cytotoxicity to normal bone marrow progenitors was enhanced significantly only by G-CSF (p = 0.02 at 0.01 microM, p = 0.01 at 0.1 microM and p < 0.01 at 1 microM Ara-C), and by GM-CSF at 0.1 microM Ara-C (p = 0.045). However, the effect of HGF stimulation as studied by bromodeoxyuridine (BrdU) incorporation during the first or second 24 hours of HGF stimulation did not explain the difference between poor and good HGF enhanced Ara-C cytotoxicity, indicating that other cellular changes than cell-cycle activation as the consequence of growth factor stimulation are responsible for enhanced cytotoxicity. Our findings indicate that combining HGFs, and especially IL-3, with chemotherapy may be useful in the AML treatment.
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PMID:Enhanced chemosensitivity of clonogenic blasts from patients with acute myeloid leukemia by G-CSF, IL-3 or GM-CSF stimulation. 768 39

The MTT assay, a colorimetric assay, is found to be suitable for chemosensitivity testing. Recently, it has been suggested that hematopoietic growth factors (HGF) may enhance the effects of cytostatic drugs in acute myeloid leukemia (AML). We therefore studied the effects of granulocyte colony-stimulating factor (G-CSF), interleukin 3 (IL-3), and granulocyte-macrophage colony-stimulating factor (GM-CSF) combined with cytosine arabinoside (Ara-C), daunorubicin (DNR), mitoxantrone (MXT), or etoposide (VP-16) by using the MTT assay. The results were compared with in vitro clonogenic assays. Briefly, AML cells of nine patients were incubated in the presence or absence of G-CSF, IL-3, or GM-CSF under serum-free conditions for 24 hours. Next, for the MTT assay, Ara-C (final dilution range: 0.0024-240 micrograms/ml), DNR (final dilution range: 0.05-3.2 micrograms/ml), MXT (final dilution range: 0.05-3.2 micrograms/ml), or VP-16 (final dilution range: 0.1-100 micrograms/ml) were added and incubated for 48 hours. Cell survival was determined and IC75 values (75% reduction as compared to control cultures) were calculated. For clonogenic assays, the three lowest drug concentrations were used. After 48 hours, the clonogenic response was determined in serum-free, semi-solid cultures with G-CSF, IL-3, or GM-CSF. The results obtained by the MTT assay showed no significant enhancement of cytotoxicity by HGF on cytostatic drug preincubated cells compared to cytostatic drugs alone. The results obtained by the clonogenic assays showed increased cytotoxicity of Ara-C combined with G-CSF, IL-3, or GM-CSF. The median IC75 values of Ara-C decreased from 0.056 to 0.0168 microgram/ml with G-CSF (p = 0.01), from 0.108 to 0.0168 microgram/ml with IL-3 (p = 0.004) and from 0.12 to 0.0204 microgram/ml for GM-CSF (p = 0.02). Only moderate enhanced cytotoxicity was observed when VP-16 was combined with IL-3 (p = 0.036) or GM-CSF (p = 0.036), but not with G-CSF. No enhanced cytotoxicity of DNR and MXT to clonogenic AML cells was found when these agents were combined with HGF stimulation. The results indicate that the MTT assay underestimates HGF enhanced cytotoxicity of Ara-C or VP-16 to clonogenic cells. Therefore, the assay is not useful for accurately detecting differences of clonogenic response due to the proliferative status of cells. In this paper, the potential explanations for the failure of the MTT assay are discussed.
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PMID:Enhanced chemosensitivity in acute myeloid leukemia by hematopoietic growth factors: a comparison of the MTT assay with a clonogenic assay. 769 94

In preparation for a clinical trial using GM-CSF on days 4-10 of sequential high-dose cytarabine (ara-C) and asparaginase (ASNase) on days 1-3 and 8-10, potential interactions between the protein synthesis inhibitor ASNase and GM-CSF were evaluated. Granulocyte-macrophage colony-stimulating factor (GM-CSF) can stimulate acute myeloid leukemia (AML) cells to proliferate in vitro and in vivo. Log phase HL-60 cells were exposed to ara-C (10 microM x 3 h) and/or ASNase (10 U/ml during the last 2 h of ara-C). Ara-C and/or ASNase was removed and cells were incubated with or without GM-CSF (10 ng/ml). After 24, 48 and 72 h of GM-CSF there was no significant difference in the S phase fraction of cells exposed to ASNase prior to GM-CSF. Soft agar cloning efficiency was determined after retreatment with ara-C +/- ASNase 24 h into the GM-CSF incubation. GM-CSF enhanced cytotoxicity for all combinations, although this effect was of borderline significance (P = 0.0621); addition of ASNase to the treatment regimen significantly (P = 0.0229) enhanced cytotoxicity without any evidence of a negative interaction with GM-CSF. In addition, ara-C metabolism was assessed during simultaneous exposure to ara-C (10 microM x 3 h) +/- ASNase (10 U/ml the last 2 h) +/- GM-CSF (10 ng/ml beginning 24 h prior to ara-C). Ara-C incorporated into DNA (P = 0.0302) and ara-CTP formation (P = 0.0084 and P = 0.0003 at 2 and 3 h timepoints, respectively) were both increased significantly by GM-CSF, with modest non-significant increases with ASNase exposures. Neither ASNase nor GM-CSF inhibited the effects of the other in this in vitro model. Therefore, when appropriately scheduled, both GM-CSF and ASNase may potentiate ara-C cytotoxicity.
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PMID:GM-CSF and asparaginase potentiate ara-C cytotoxicity in HL-60 cells. 788 38

Recombinant human granulocyte-macrophage colony-stimulating factor (rhGM-CSF) stimulates the growth of myeloid leukemic cells and increases their susceptibility to cell-cycle specific agents. We treated a patient with acute myelogenous leukemia (AML) in a state of second resistant relapse, with high-dose chemoradiotherapy combined with rhGM-CSF (total body irradiation: TBI 3Gy x 4, on days -8 & -7; cytosine arabinoside: Ara-C 3g/m2, iv, q12h, on days -5-2; rhGM-CSF 250 micrograms/m2/day, cont.iv, on days -5-2) followed by autologous peripheral blood stem cell transplantation (PBSCT). In this case, rhGM-CSF enhanced the proliferation of leukemic cells in vitro. The test dose of rhGM-CSF (84 micrograms/m2 over 8 hours) also promoted leukemic cell proliferation in vivo, resulting in an increase in the percentage of leukemic cells in the peripheral blood and reappearance of chromosomal aberrations in the bone marrow. The toxicity of rhGM-CSF-combined conditioning regimen included fever and mild liver damage. The patient achieved a complete remission lasting for 2 months, then relapsed. The rhGM-CSF-combined conditioning regimen was tolerated by this patient, but further studies will be required to confirm not only its safety but also its effectiveness in the treatment of refractory AML.
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PMID:[High-dose chemoradiotherapy combined with recombinant human granulocyte-macrophage colony-stimulating factor (rhGM-CSF) followed by autologous peripheral blood stem cell transplantation (PBSCT) in a case of refractory acute myelogenous leukemia (AML)]. 810 15

The effect of recombinant human granulocyte-macrophage colony-stimulating factor (GM-CSF) on the intracellular metabolism of cytosine arabinoside (Ara-C) was comparatively analyzed in normal bone marrow mononuclear cells (NBMMC) from eight healthy volunteers and in leukemic blasts from 50 patients with acute myeloid leukemia (AML). Pretreatment with GM-CSF (100 U/ml) for 48 h resulted in a significant enhancement of DNA synthesis in both cell types: 21 of 35 AML specimens were found to be responsive to GM-CSF as defined by an increase of 3H-TdR incorporation into the DNA > 1.5-fold while NBMMC from normal donors were responsive in all cases. In GM-CSF responsive AML blasts, overall DNA polymerase and DNA polymerase alpha activity increased from a median of 84.4 to 96.1 and from 3.45 to 5.2 pmol/min x mg as compared to a median of 96.7 to 189.9 and 1.2 to 2.2 pmol/min x mg in NBMMC (P < 0.05). Median Ara-C-mediated inhibition of DNA synthesis was significantly more effective in AML blasts as compared to NBMMC (76.5 vs 55.0% at 0.05 microM and 99.0 vs 96.0% at 5.0 microM Ara-C, P < 0.01) but was not influenced by GM-CSF pretreatment. Similarly, intracellular Ara-CTP levels were higher in AML blasts as compared to NBMMC (median of 46.9 vs 18.7 at 1 microM, 167.8 vs 48.0 at 10 microM and 337.5 vs 59.5 ng/10(7) cells at 100 microM extracellular Ara-C, P < 0.01) but showed no enhancement in the presence of GM-CSF. Median deoxycytidine (DCK) and thymidine kinase (TK) activity were only slightly increased in AML blasts after GM-CSF priming. In contrast, NBMMC revealed a significant increase in TK activity after GM-CSF pretreatment (from a median of 1.9 to 3.6 pmol/min x mg, P = 0.039). At low; intermediate and high extracellular Ara-C concentrations GM-CSF pretreatment resulted in a significant enhancement of the 3H-Ara-C incorporation into the DNA in both GM-CSF responsive AML blasts and NBMMC (median of 1.3 to 2.1- and 1.4 to 1.6-fold, P < 0.05). GM-CSF non-responsive AML blasts showed no change in 3H-Ara-C incorporation into the DNA in response to GM-CSF at low Ara-C concentrations but significant increases at intermediate and high extracellular Ara-C concentrations (median increases of 1.63-fold at 1.06 microM with P = 0.01 and 1.37-fold at 10 microM extracellular Ara-C with P = 0.0+005). NBMMC revealed significantly lower GM-CSF-induced increases of the 3H-Ara-C incorporation into the DNA as compared to the effect of GM-CSF priming on DNA synthesis (median increases of 1.4 to 1.7-fold vs 2.6-fold, P < 0.05). These data reveal a different effect of GM-CSF priming on the metabolism of Ara-C in normal vs leukemic cells which may cause a preferential increase in the antileukemic cytotoxicity of Ara-C in the presence of GM-CSF.
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PMID:Differential effect of GM-CSF pretreatment on intracellular Ara-C metabolism in normal bone marrow mononuclear cells vs acute myeloid leukemia (AML) blasts. 909 97

The present study was undertaken to assess the predictive value of pretherapeutic determinants of ara-C metabolism and proliferative activity of leukemic blasts for early response to antileukemic therapy in the setting of granulocyte-macrophage colony-stimulating factor (GM-CSF)-based priming before and during TAD-9 induction in 36 consecutive patients with de novo acute myeloid leukemia (AML). Ara-C metabolism was assessed by the activities of deoxycytidine kinase (DCK), deoxycytidine deaminase (DCD), DNA polymerase alpha (Poly alpha), and overall polymerase (overall Poly). The fraction of cells in S phase (%S phase) and thymidine kinase (TK) activity were determined as a measure of proliferative activity. Early response to therapy was defined by the percentage of leukemic blasts in the bone marrow 5 to 7 days after completion of TAD-9 with less than 5% signaling an adequate response and greater than 5% indicating an inadequate early reduction, respectively. While neither %S phase, DCK, nor overall Poly activity were predictive for early response, TK and Poly alpha activities were significantly higher for cases with adequate blast cell clearance. The respective median values were for TK 3.8 versus 1.85 pmol/min/mg protein (P = .012), and for Poly alpha 1.9 versus 0.69 pmol/min/mg protein (P = .014). An inverse relation was detected for DCD activity which was significantly lower in responding patients with a median of 0.33 nmol/min/mg protein (range, 0.0 to 29.5) as compared to a median of 5.1 nmol/min/mg protein (range, 0.11 to 8.45) in early nonresponders, (P = .009). Taking the respective median values as arbitrary cut-points for high or low enzyme activities, responders and nonresponders could be discriminated prospectively. Hence, 14 of 16 cases (88%) with DCD activities below the median of 1.56 nmol/min/mg protein responded as compared to only 3 of 14 (22%) patients with higher DCD activities (P = .0004). From the 15 patients with TK activity above the overall median of 3.2 pmol/min/mg protein, 11 cases (73%) achieved an adequate blast cell clearance while only 6 of 17 cases (35%) with lower values responded (P = .035). Similarly, 12 of 15 patients (80%) with high Poly alpha levels (>1.22 pmol/min/mg protein) responded to induction therapy as compared to only 5 of 14 patients (36%) with lower enzyme activities (P = .02). By logistic regression analysis of enzyme activities, DCD activity was found to be the most sensitive parameter to predict an adequate blast cell clearance (P = .032). Activities of DCD and TK were not only associated with initial response but were also found predictive for remission duration. Hence, from 11 patients with low TK levels 8 (73%) relapsed within 1 year, whereas only 2 of 11 (18%) patients with high TK activity experienced a recurrence of their disease (P = .015). Six of 9 (66%) patients with higher than median DCD levels relapsed within 1 year, whereas 10 of 14 patients (71%) with lower DCD levels had a longer remission duration (P = .085). Analysis of DCD gene expression at the mRNA level by a semi-quantitative reverse transcriptase-polymerase chain reaction method showed that a high transcription rate of the DCD gene was associated with high enzyme activities and vice versa. Hence, the observed intraindividual differences in DCD activity are a reflection of differences in gene activity and transcription rate rather than of variants in translation. Although further analyses are needed to elucidate the molecular mechanisms that determine the variation of enzyme activities in individual patients, the present study strongly suggests that pretherapeutic determination of TK and Poly alpha as well as of DCD allows to predict response to TAD-9 + GM-CSF induction therapy and may provide the means for the development of a risk adapted treatment strategy.
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PMID:Activity of thymidine kinase and of polymerase alpha as well as activity and gene expression of deoxycytidine deaminase in leukemic blasts are correlated with clinical response in the setting of granulocyte-macrophage colony-stimulating factor-based priming before and during TAD-9 induction therapy in acute myeloid leukemia. 929 31

Despite significant advances in the treatment of acute myeloid leukemia (AML), the majority of patients will succumb to drug-resistant AML. To overcome this resistance, we have developed a novel fusion toxin consisting of the catalytic and translocation subunits of diphtheria toxin (DT388) linked to human granulocyte-macrophage colony-stimulating factor (GM-CSF). In vitro, DT388-GM-CSF demonstrated significant activity against numerous AML cell lines and fresh AML blasts. To determine its in vivo efficacy, we developed an in vivo model of human AML in severe combined immunodeficiency (SCID) mice injected intravenously with 1 x 10(7) HL-60 cells (AML-M2 cell line). The SCID mice developed abdominal masses, infiltration of the liver and bone marrow, and peripheral blasts with a median survival of 42.5 days. We tested DT388-GM-CSF, ara-C, human GM-CSF, and DAB389IL-2, which were injected intraperitoneally on days 2-6 in this model. DT3-GM-CSF significantly improved survival of the SCID mice over Ara-C, DAB389IL-2, or control (P < 0.001). DT388-GM-CSF-treated mice who developed leukemia exhibited no difference in the number of GM-CSF receptors (P = 0.39), ligand affinity (P = 0.77), or sensitivity (P = 0.56) to DT388-GM-CSF as compared to the controls. Frank leukemia in DT388-GM-CSF-treated mice may be due to incomplete penetration of drug into tissues rather than cellular resistance. DT388-GM-CSF is an active therapeutic agent in our SCID mouse model of AML with a unique mechanism of action and differing toxicities than current cytotoxic agents.
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PMID:DT388-GM-CSF, a novel fusion toxin consisting of a truncated diphtheria toxin fused to human granulocyte-macrophage colony-stimulating factor, prolongs host survival in a SCID mouse model of acute myeloid leukemia. 1021 72

Human granulocyte-macrophage colony-stimulating factor fused to truncated diphtheria toxin (DT388-GM-CSF) sensitized wild-type and Bcl2-overexpressing HL60 human leukemia cells to intoxication by Ara-C based on proliferation and clonogenic assays. The toxin/drug combination showed dramatic synergistic toxicity with combination indices of < 0.1. Synergy was not seen with two other protein synthesis inhibiting drugs--ricin and cycloheximide nor with GMCSF alone. No changes in Ara-C incorporation into cellular DNA or cell cycle occupancy were seen. As compared to exposure to DT388-GM-CSF or Ara-C alone, co-treatment produced significant increases in cytosolic accumulation of cytochrome c, a higher percentage of cells with loss of mitochondrial membrane potential and an increase in reactive oxygen species and morphologic changes of apoptosis, and a greater induction of poly(ADP-ribose) polymerase (PARP) and DNA fragmentation factor 45 (DFF45) cleavage activities of caspase 3. Co-treatment did not significantly alter Bcl2, Bcl-xL, Bax or Fas receptor (FasR), but modestly increased Fas ligand (FasL) protein. These finding suggest that co-treatment with DT388-GM-CSF may lead to a lowered apoptotic threshold and clonogenic survival of human AML blasts due to Ara-C. These observations also suggest that clinical trials of combination therapy may be warranted in patients with AML.
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PMID:Diphtheria toxin fused to granulocyte-macrophage colony-stimulating factor and Ara-C exert synergistic toxicity against human AML HL-60 cells. 1037 46

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