Recombinant Granulocyte-macrophage Colony Stimulating Research Articles

AGIG Chemo-Immunotherapy in Patients With Advanced Pancreatic Cancer: A Single-Arm, Single-Center, Phase 2 Study.

BackgroundTo date, chemotherapy remains the only effective treatment of unresectable pancreatic adenocarcinoma. In the past few years, the interest in immunological anticancer therapy rises sharply. AGIG is a novel chemo-immunotherapy regimen that combines nab-paclitaxel + gemcitabine chemotherapy with sequential recombinant interleukin-2 (IL-2) and granulocyte-macrophage colony stimulating factor (GM-CSF) therapy. We conducted a single-arm prospective phase II study to determine the efficacy and safety of the first-line treatment of advanced pancreatic cancer with AGIG regimen.MethodsNab-paclitaxel (125 mg/m2) and gemcitabine (1000 mg/m2) were administered intravenously to all patients on days 1 and 8 triweekly, interleukin-2 (1000000U) and GM-CSF (100 µg) were administered subcutaneously on days 3-5 after chemotherapy. The primary end point was ORR by the Response Evaluation Criteria in Solid Tumors, version 1.1. Secondary end points included safety profile, progression-free survival (PFS), overall survival (OS). Patients’ conditions along with the efficacy and safety were assessed every two cycles.ResultsBetween 11/2018 and 01/2020, sixty-four patients were enrolled. In the sixty-four evaluable patients, the disease control rate (DCR) and overall response rate (ORR) were 76.6% and 43.75%, respectively. The median follow-up time was 12.1 (range 7.1–22.4) months. The median PFS was 5.7 (range 1.63–15.8) months. The median OS was 14.2 (range 2.9–22.0) months. The most common adverse event was fever (75%). The incidence of III/IV grade neutropenia was 4.69%. In subgroup analyses, we found that eosinophil count in the blood elevated three times higher than baseline level predicted a longer survival.ConclusionsThe AGIG chemo-immunotherapy regimen has presented favorable ORR, OS, and manageable toxicities as first-line therapeutic strategy of advanced pancreatic cancer treatment. This regimen may be a novel reliable therapeutic option for patients with preserved performance status. The improvement of treatment efficiency may be related to the activation of non-specific immune response.Clinical Trial Registration https://clinicaltrials.gov/. identifier NCT03768687.

AGIG chemo-immunotherapy in patients with advanced pancreatic cancer: A single-arm, single-center, phase II study.

384 Background: To date, chemotherapy remains the only effective treatment of unresectable pancreatic adenocarcinoma. In the last few years, the interest in the use of immunological anticancer strategies is greatly increased. AGIG is a novel chemo-immunotherapy regimen that combines nab-paclitaxel + gemcitabine chemotherapy with sequential recombinant interleukin-2 and granulocyte-macrophage colony stimulating factor (GM-CSF) therapy. We conducted a single-arm prospective phase II study to determine the efficacy and safety of the first-line treatment of advanced pancreatic cancer with AGIG regimen. Methods: Nab-paclitaxel (120 mg/m2) and gemcitabine (1000 mg/m2) were administered intravenously to all patients on days 1 and 8 triweekly, interleukin-2 and GM-CSF (100 µg) were administered subcutaneously on days 3-5 after chemotherapy. The primary end point was ORR by the Response Evaluation Criteria in Solid Tumors, version 1.1. Secondary end points included safety profile, progression-free survival (PFS), overall survival (OS). Patients’ conditions and the efficacy and safety were assessed every 4 cycles. Results: Between 11/2018 and 01/2020, 64 patients were enrolled. In the 64 evaluable patients, the ORR and DCR were 43.75% and 76.6%, respectively. The median follow-up time was 12.1 (range 7.1–22.4) months, the median PFS was 5.7 (range 1.63–15.8) months, and the median OS was 14.2 (range 2.9–22.0) months. The most common adverse event was fever (75%). The incidence of grade 3/4 neutropenia was 4.69%. In subgroup analyses, we found that eosinophil count in the blood elevated three times higher than baseline level predicted a longer survival. Conclusions: The AGIG Chemo-immunotherapy regimen has presented encouraging ORR, OS, and manageable toxicities as first-line therapy for advanced pancreatic cancer. This regimen may be a novel reliable therapeutic option for patients with preserved performance status. The improvement of treatment efficiency may be related to the activation of non-specific immune response. Clinical trial information: NCT0376867. Research Sponsor: The Affiliated Nanjing Drum Tower Hospital of Nanjing University Medical School. [Table: see text]

Granulocyte–Macrophage Colony-Stimulating Factor Is Essential for Normal Wound Healing

Granulocyte-macrophage colony-stimulating factor (GM-CSF) is a multipotent growth factor, which plays an important role during the process of wound healing. In clinical settings it has occasionally been employed in the treatment of cutaneous wounds of diverse etiologies. In a previous study, we have shown the positive influence of GM-CSF on full thickness excisional wounds in transgenic mice overexpressing GM-CSF in the basal layer of the epidermis. Direct GM-CSF action as well as indirect processes through the induction of secondary cytokines were proposed to contribute towards the beneficial effects. In this study, we analyzed the process of wound healing in transgenic mice overexpressing a GM-CSF antagonist in the epidermis. These mice not only exhibited a delayed scab rejection and reepithelialization but also neovascularization was reduced. The newly formed tissue was of poor quality as exhibited by the presence of extensive fibrosis. We suggest that the presence of GM-CSF in the repair process is of basic importance and its absence leads not only to delayed wound healing but it is also detrimental for the quality of the newly formed tissue.

The influence of mycophenolate mofetil upon the maturation and allostimulatory activity of cultured dendritic cell progenitors and the effects of tolerance induction in allograft recipients

To investigate the influence of mycophenolate mofetil (MMF) on the maturation and allostimulatory activity of cultured dendritic cell progenitors (DCPs) and to evaluate the efficacy of pretreatment of donor dendritic cells (DCs) with MMF in tolerance induction in allograft recipients and its possible mechanism. DCPs of Balb/c mice were cultured in culture fluid containing recombinant mouse granulocyte-macrophage colony stimulating factor (GM-CSF), and then divided into 4 groups: control group, MMF group, lipopolysaccharide (LPS) group, and MMF + LPS group. Seven days later, flow cytometry was used to analyze the phenotypes of the DCs. ELISA was used to examine the level of IL-12 in the supernatant. T cells from the spleens of C57/BL6 mice were cultured together with inactivated DCs from Balb/c mice, and added with [(3)H]-TdR. Mixed lymphocyte reaction (MLR) was analyzed. The DCs and MMF-DCs cultured for 5 days were co-cultured with T hybridoma cells of the line MF2.2D9 in culture fluid containing ovalbumin (OVA). Twenty-four hours later, the supernatant was collected and ELISA was used to measure the level of interleukin (IL)-12 so as to reflect the antigen-presenting ability of the DCs. OVA immunized C57/BL6 mice for 4 times. 21 days after T cells were collected from the spleens and co-incubated with DCs and MMF-DCs, [(3)H]-TdR was added and the values of counts per minute (cpm) were calculated so as to analyze the antigen-specific proliferation. Twenty-four Balb/c mice were randomly divided into 3 groups: group A (without treatment), group B (treatment with intravenous injection of untreated DCs of Balb/c mice), and group C (treatment with intravenous injection of DCs of Balb/c mice treated with MMF), and then transplanted with the hearts of C57/BL6 mice. The functions of the transplanted hearts were evaluated by touching the arterial pulse and histological examination. ELISA was used to detect the levels of Th1 cytokines, such as IL-12, IL-4, IL-10, IL-2, and IFN-gamma, in the cultured DCs and in the sera of the recipients 7 and 14 days after culture or transplantation. The immunophenotypic analysis showed that in comparison with those in the control group and the LPS group the expressions of the costimulatory molecules, Ia(d), CD80, and CD86, of the DCPs were significantly weaker in the MMF-group and MMP + LPS group. The IL-12 levels in the supernatant of the MMF and MMF + LPS groups of DCPs were significantly lower than those in the other groups (P < 0.01). The IL-12 level of the MF2.2D9 cells treated with MMF-treated DCs was significantly lower than control group (P < 0.01). The ability to stimulate proliferation of T cells of the same genotype in the MMF-DC group was significantly inhibited (P < 0.01). The survival time of the transplanted heart was 30.50 +/- 3.25 days in the C57/BL6 mice injected with untreated DCPs of Balb/c mice (21.25 +/- 2.12, P < 0.01) and that in the control C57/BL6 mice (8.63 +/- 1.06 days, P < 0.01) and with a significant difference between the latter 2 groups too (P < 0.01). The levels of, such as IL-2 and IFN-gamma, 7 and 14 days after heart transplantation of the group B were all significantly lower than those of the group A (both P < 0.05). The IL-2 and IFN-gamma levels 7 days after the heart transplantation were similar to those in the group B (both P > 0.05) and even lower than those of the group C (both P < 0.05). The IL-10 level in the groups B and C were all higher than those in the group A (all P < 0.05) with a significant difference between the group B and group C. The level of IL-4 was not significantly different among the 3 groups. MMF has a significant suppressive effect on the maturation and function of DCs, which leads to a donor-specific tolerance in transplant recipients.

Effect of inclusion of serum and granulocyte - macrophage colony stimulating factor on secretion of interferon-τ during the in vitro culture of ovine embryos

In each of three experiments, in vitro-matured and -fertilised zygotes were cultured to Day 7 post insemination in synthetic oviductal fluid (SOF). In Experiment 1, zygotes were cultured in groups in either SOF plus albumin (SOFA) or serum (SOFS) and then blastocysts were cultured individually for a further 24 h without a change of media. In Experiment 2, zygotes were cultured in groups using a 2 x 2 factorial design in SOFA or SOFS, with or without recombinant ovine granulocyte-macrophage colony stimulating factor (GM-CSF; 5 ng mL(-1)). Blastocysts were then cultured individually using a split-plot design in SOFA or SOFS with or without GM-CSF. In Experiment 3, zygotes were cultured in SOFA in which GM-CSF was absent (A) or present (P) during Days 1-3, Days 3-5 or Days 5-7 of IVC in six combinations as follows: AAA, AAP, APP, PPP, PPA and PAA. Serum or GM-CSF increased secretion of interferon (IFN)-tau in Experiments 1 and 2 both between Days 5 and 7 of group culture and during individual culture. Secretion of IFN-tau during individual culture was determined by the medium in which embryos were group cultured and the effects of GM-CSF and serum were not additive. In Experiment 3, the presence of GM-CSF between Days 1 and 3 of culture was responsible for stimulation of secretion of IFN-tau between Days 5 and 7; IFN-tau secretion was detected as early as Day 3 post insemination.

Recombinant granulocyte macrophage colony stimulating factor (rhu-GM-CSF) in the treatment of extensive leg ulcers: A case report

Surgery 2000;127:589–92.

Analysis of a recombinant granulocyte macrophage colony stimulating factor dosage form by capillary electrophoresis, capillary isoelectric focusing and high-performance liquid chromatography

The analysis of a recombinant granulocyte macrophage colony stimulating factor (GM-CSF) dosage form by free solution capillary electrophoresis (FSCE), capillary isoelectric focusing (cIEF), and high-performance liquid chromatography (HPLC) is described. The quantitative use of capillary electrophoresis, whether FSCE or cIEF, will always be prone to special problems, such as sensitivity to differences in salt concentrations between the standard and sample, and can not match the ruggedness of HPLC. The usable quantitative linear range for both HPLC and FSCE surpass that achieved for cIEF methods by a factor of 10 or greater. The FSCE system, utilizing an octyl bonded/Brij-35 coated capillary, did not work for all proteins examined. This is probably due to an interaction of the protein with the bonded phase or the absorbed Brij-35. In contrast, the cIEF method worked well for all proteins tested thus far, yielding high efficiency and resolution comparable to slab gel isoelectric focusing. This paper addresses the potential for using free solution capillary electrophoresis and capillary isoelectric focusing as a quantitative analytical tool. Also, the effect of salt in the dosage form on quantitation, reproducibility, and efficiency of capillary electrophoresis methods is also discussed.

Divergent regulation of cell surface protease expression in HL-60 cells differentiated into macrophages with granulocyte macrophage colony stimulating factor or neutrophils with retinoic acid.

Taking advantage of the recently demonstrated presence of N-aminopeptidases and the serine protease dipeptidyl aminopeptidase IV (DPP IV) at the surface of human myeloblastic HL-60 cells, the regulation of these protease activities in HL-60 cell differentiation has been assessed using combined spectrophotometric and flow cytometric assays. Addition of human recombinant granulocyte macrophage colony stimulating factor (rHu-GM-CSF) to HL-60 cells to induce differentiation into macrophages led to a time- and dose-dependent increase in both cell surface N-aminopeptidase and DPP IV activities. Protease up-regulation was due to an enhancement in cell surface protease number, associated with a slight rise in apparent affinities of the enzymes for their substrates. In contrast, in HL-60 cells induced to differentiate into neutrophils in the presence of retinoic acid, expression of cell surface N-aminopeptidases was almost completely abolished in a time- and dose-dependent fashion, and this down-regulation was accompanied by a weak but significant decrease in affinity. However, no noticeable difference was seen in serine DPP IV expression between retinoic acid-treated and untreated HL-60 cells. Retinoic acid treatment also reduced soluble protease activity in vitro indicating that down-regulation of membrane aminopeptidases was not due to their proteolytic clip. No modulation in the activity of any of the enzymes tested was seen with human recombinant tumor necrosis factor-alpha or retinol which do not induce HL-60 cell differentiation.(ABSTRACT TRUNCATED AT 250 WORDS)

Production of epithelial cell growth factors by lamina propria mononuclear cells.

The effects of lamina propria mononuclear cell culture supernatant on epithelial cell DNA synthesis were studied using cells isolated from patients with inflammatory bowel disease and normal controls. Supernatants from resting and phytohaemagglutinin stimulated cells were studied and supernatants that strongly promoted DNA synthesis were pooled, and growth factor activity partially characterised. The effects of recombinant interleukins-1 beta,2,3,interferon-gamma, and granulocyte macrophage colony stimulating factor were tested in the same system. Resting lamina propria mononuclear cells produce factors that increase DNA synthesis. Production of these factors is increased by phytohaemagglutinin stimulation. No significant differences were found in production of these factors between patients with inflammatory bowel disease and normal controls. The molecular weight of the active factor(s) lies in the region 31-48 kD. Chromatofocusing produced two peaks of activity, one in the region pk 5.5 and one around pk 6.4. The activity was heat and acid pH labile. Activity was not destroyed, however, by 0.05% trypsin. Recombinant granulocyte macrophage colony stimulating factor was a weak stimulus to epithelial DNA synthesis, interleukin-1 beta was weakly inhibitory but other cytokines tested did not have any effect. Granulocyte macrophage colony stimulating factor is probably important in controlling epithelial cell growth.

In vitro activation of rGM‐CSF derived bone marrow macrophages by cisplatin and lipopolysaccharide

Treatment of fresh non-adherent bone marrow cells (NABMC) with cisplatin or lipopolysaccharide (LPS) did not render them tumoricidal. NABMC incubated in medium alone or medium containing recombinant granulocyte macrophage colony stimulating factor (rGM-CSF) for 4 days matured to macrophages that were positive for non-specific esterase staining. Bone marrow-derived macrophages cultured with medium alone did not respond to cisplatin or LPS by the induction of tumoricidal activity, whereas rGM-CSF derived bone macrophages showed significantly enhanced cytotoxicity after treatment with cisplatin or LPS. Culturing of NABMC with rGM-CSF enhanced cell survival compared to the cells incubated in medium alone. These results suggest that bone marrow cells not only mature in rGM-CSF, but are also primed by rGM-CSF for induction of tumoricidal activity.

Therapy of human ovarian cancer xenografts with intraperitoneal liposome encapsulated muramyl-tripeptide phosphoethanolamine (MTP-PE) and recombinant GM-CSF

Three intraperitoneal human ovarian cancer xenografts (OS, HU, and LA) were used to assess the antitumour activity of intraperitoneal therapy with liposome encapsulated MTP-PE. MTP-PE led to significant prolongation of survival in all three xenograft models, but with varying efficacy. In one tumour model (OS), 80% of mice were cured of tumour by twice weekly therapy for 4 weeks, whereas in another xenograft model (LA), the median survival time was approximately doubled compared to PBS injected and placebo liposome injected controls (median survivals: 30 vs 62.5 days respectively). The antitumor efficacy of MTP-PE did not correlate with the extent of peritoneal neutrophil infiltration after intraperitoneal therapy. Combined therapy with liposome encapsulated MTP-PE and recombinant murine granulocyte-macrophage colony stimulating factor led to increased survival of mice bearing the LA and HU xenografts, compared to tumour bearing mice treated with either agent singly.

Enhanced superoxide generation by eosinophils from asthmatic children.

In 21 asthmatic children, aged 2-19 years, and 11 age-matched controls purified (purity greater than 75%) peripheral blood eosinophils and neutrophils (purity greater than 95%) were stimulated with phorbol myristate acetate (44 ng/ml) for 30 min at 37 degrees C. The superoxide generation was determined as well as the superoxide dismutase inhibitable cytochrome c reduction. Eosinophils obtained from asthmatic patients displayed a significantly higher O2- generation (p less than 0.01) when compared to neutrophils from the same patients or to eosinophils and neutrophils from healthy controls. Preincubation with human recombinant granulocyte macrophage colony stimulating factor for 30 min or platelet-activating factor for 10 min resulted in augmented O2- generation by eosinophils from control, but not from asthmatic children, indicating a possible in vivo priming of eosinophils in asthma.

Hypoalbuminaemia after prolonged treatment with recombinant granulocyte macrophage colony stimulating factor.

Hypoalbuminaemia after prolonged treatment with recombinant granulocyte macrophage colony stimulating factor.

Enhanced resistance of bone marrow transplanted mice to bacterial infection induced by recombinant granulocyte-macrophage colony- stimulating factor

The in vivo effect of recombinant murine granulocyte-macrophage colony stimulating factor (rGM-CSF) on the resistance of mice to bacterial infection and on the number and function of neutrophils was studied in lethally irradiated mice transplanted with syngeneic bone marrow cells. Bone marrow transplanted (BMT) mice were injected intraperitoneally with 150 ng rGM-CSF or buffer solution (diluent) twice daily for 18 consecutive days. Total neutrophil recovery from the peripheral blood and the number of neutrophils mobilized into the peritoneal cavity were accelerated in rGM-CSF-treated recipients. Peritoneal neutrophils isolated from mice treated with rGM-CSF exhibited primed superoxide generation (O2-) after in vitro stimulation with suboptimal concentrations of phorbol myristate acetate (PMA), as compared with control mice (treated with diluent). No additional increase in O2- production occurred upon in vitro incubation of these cells with rGM- CSF. The protective activity of rGM-CSF was examined in mice injected with Salmonella typhimurium. There was a 44- and 9-fold increase in the number of S typhimurium at 96 hours postinfection in the spleen and liver, respectively, of control mice, as compared with rGM-CSF-treated mice, after a single injection of the bacteria (3 X 10(7) per mouse). All the untreated control mice died within 14 days postinoculation (1 X 10(7) bacteria per mouse), whereas 35% of the mice treated with rGM-CSF remained alive for more than 30 days postinfection. These findings support the concept that increased granulopoiesis and enhanced functional activity of phagocytic cells is induced by rGM-CSF and is responsible for enhanced resistance of BMT mice to bacterial infection.

Enhancement of methotrexate cytotoxicity by modulation of proliferative activity in normal and neoplastic T lymphocytes and in a myeloid leukemia cell line.

Recent advances in the modulation of cell kinetics with growth factors suggest that the effect of cycle-specific cytostatic drugs can be enhanced by combination with such factors. The truth of this hypothesis was investigated by studying the effect of phytohemagglutinin and/or interleukin 2 on the sensitivity to methotrexate (MTX) of normal T lymphocytes and of lymphoblasts of a patient with acute T-cell lymphoid leukemia. In both cases, inhibition of proliferation by MTX was increased from less than 30% in resting cells or those suboptimally stimulated, in the case of leukemic blasts, to 68-83% in maximally stimulated cells. Similar results were observed when the AML 193 human myeloid leukemia line was stimulated with human recombinant granulocyte macrophage colony stimulation factor (GM-CSF). Under basal proliferation conditions, the addition of 1 microgram/ml and 10 micrograms/ml MTX was followed by 48% and 72% inhibition respectively. When 1 ng/ml GM-CSF (40 I.U./ml) was present, these figures rose to 89% and 91%. It is thus clear that growth factor-induced cell proliferation increases sensitivity to cycle-specific cytostatic agents. There is thus a biological premise for new perspectives in antineoplastic therapy.