Subcutaneous Granulocyte-macrophage Colony-stimulating Factor Research Articles

A phase II study of GM-CSF and rIFN-γ1b plus carboplatin for the treatment of recurrent, platinum-sensitive ovarian, fallopian tube and primary peritoneal cancer

Objective To evaluate the efficacy and toxicity of carboplatin, granulocyte-macrophage colony-stimulating factor (GM-CSF) and recombinant interferon gamma 1b (rIFN-γ1b) in women with recurrent, platinum-sensitive ovarian, fallopian tube and primary peritoneal cancer. Methods In this phase II study, patients with recurrent, platinum-sensitive ovarian, fallopian tube or primary peritoneal cancer were treated with subcutaneous GM-CSF and rIFN-γ1b before and after intravenous carboplatin until disease progression or unacceptable toxicity. All patients had measurable disease and a chemotherapy-free interval > 6 months. Response was determined using RECIST criteria and CA 125 levels. Results Between 2003 and 2007, 59 patients received a median of 6 cycles of therapy (range, 1 to 13 cycles). Median age at enrollment was 61 years (range, 35 to 79 years). Median time to progression prior to enrollment was 11 months (range, 6 to 58 months). Of 54 patients evaluable for response, 9 (17%) had a complete response, 21 (39%) had a partial response, and 24 (44%) had progressive disease. The overall response rate was 56% (95% CI: 41% to 69%). With a median follow-up of 6.4 months, median time to progression was 6 months. Myeloid derived cells and platelets increased on day 9 of each chemotherapy cycle. The most common adverse effects were bone marrow suppression, carboplatin hypersensitivity, and fatigue. Responders reported improved quality of life. Conclusion This pre and post-carboplatin cytokine regimen resulted in a reasonable response and a hematologic profile that could invite further evaluation of its components in the treatment of patients with ovarian cancer.

Chemoimmunotherapy of chemotherapy-refractory metastatic colorectal cancer with gemcitabine plus FOLFOX followed by subcutaneous granulocyte macrophage colony-stimulating factor and interleukin-2

Chemoimmunotherapy of chemotherapy-refractory metastatic colorectal cancer with gemcitabine plus FOLFOX followed by subcutaneous granulocyte macrophage colony-stimulating factor and interleukin-2

A Phase 1 Study of Granulocyte Macrophage Colony-stimulating Factor (Sargramostim) and Escalating Doses of Thalidomide in Patients With High-risk Malignant Melanoma

This phase 1 study evaluated the safety and tolerability of adjuvant treatment with subcutaneous granulocyte macrophage colony-stimulating factor (GM-CSF) administered in combination with escalating doses of thalidomide in patients with surgically resected stage II (T4), III, or IV melanoma at high risk for recurrence. Adjuvant treatment included GM-CSF 125 microg/m2 subcutaneously for 14 days and thalidomide at an initial dose of 50 mg/d, escalated in cohorts of 3 to 6 patients each to a maximum of 400 mg/d followed by 14 days of rest. Treatment was continued for up to 1 year in the absence of disease progression. Of 19 patients treated, the most common toxicities were grade 1/2 constipation (68%), fatigue (58%), neuropathy (42%), bone and joint pain (37%), and dyspnea, dizziness, injection site skin reaction, and somnolence (32% each). Thrombotic events in 3 of 19 patients (16%), including 1 treatment-related death, were the most serious adverse events and were thought to be due to thalidomide. With a median follow-up of 945 days (2.6 y), 8 (42%) patients were alive, including 1 with disease and 7 without evidence of disease. GM-CSF plus thalidomide as adjuvant therapy for patients with resected high-risk melanoma was associated with a high incidence of thrombotic events. Because life-threatening events are unacceptable in the adjuvant setting, up-front antithrombotic prophylaxis will be necessary for further evaluation of GM-CSF plus thalidomide as a viable regimen in this patient group.

Immunity Feedback and Clinical Outcome in Colon Cancer Patients Undergoing Chemoimmunotherapy with Gemcitabine + FOLFOX followed by Subcutaneous Granulocyte Macrophage Colony-Stimulating Factor and Aldesleukin (GOLFIG-1 Trial)

GOLFIG chemoimmunotherapy regimen proved to be a safe and very active chemoimmunotherapy regimen in advanced colon cancer patients. We have thus investigated the immunobiological feedback to the treatment and its possible correlation with the clinical outcome of these patients. This clinical and immunologic study involved 46 patients, 27 males and 19 females, enrolled in the GOLFIG-1 phase II trial who received gemcitabine (1,000 mg/m(2) on days 1 and 15), oxaliplatin (85 mg/m(2) on days 2 and 16), levofolinic acid (100 mg/m(2) on days 1, 2, 15, and 16), and 5-fluorouracil (400 mg/m(2) as a bolus, and 800 mg/m(2) as a 24-hour infusion on days 1, 2, 15, and 16) followed by s.c. granulocyte macrophage colony-stimulating factor (100 mug, on days 3-7) and interleukin 2 (0.5 x 10(6) IU twice a day on days 8-14 and 17-29). The regimen was confirmed to be safe and very active in pretreated patients with metastatic colorectal cancer. A subgroup analysis of these patients revealed a prolonged time to progression and survival in six patients who developed late signs of autoimmunity. A multivariate analysis validated the occurrence of autoimmunity signs as an independent predictor of favorable outcome. A parallel immunologic study detected in the peripheral blood mononuclear cells of these patients a progressive increase in lymphocyte and eosinophil counts, amplification in central memory, a marked depletion of immunosuppressive regulatory T cells, and activation of colon cancer-specific cytotoxic T cells. Our results suggest that immunity feedback to GOLFIG regimen and its antitumor activity are tightly correlated.

Interleukin-2 and Granulocyte-Macrophage Colony Stimulating Factor Is an Alternative to Donor Lymphocyte Infusion for Relapse after Allogeneic Stem Cell Transplantation and Is Associated with Increase Immune Effector Cells and Polarization to Type 2 Dendritic Cells.

Donor lymphocyte infusion (DLI) is use for relapse after allogeneic stem cell transplant (ASCT). Immune activation with cytokines maybe an alternative to DLI. We administered Granulocyte-Macrophage Colony Stimulating Factor (GM-CSF) and Interleukin-2 (IL-2) at the time of relapse after ASCT in patients (pts) with hematologic malignancies. Pts. received subcutaneous GM-CSF at 500 mcg/day on days 1–14 and IL-2 at 1 × 106 units/m2/day on days 8–14. Pts. were off immunosuppressive therapy and had no prior history of graft versus host disease (GVHD) at the start of treatment. Twelve pts. received IL-2/GM-CSF for treatment of relapse AML (7), ALL (2), CML (1), MDS (2). Median age was 55 (range 8–66). Stem cell sources included: peripheral blood = 9, bone marrow = 2, umbilical cord blood (UCB) = 1. Donor sources were: match-related sibling = 4 and match-unrelated donor = 8 (UCB=1). Nine pts. had resistant relapse or primary resistant disease at time of ASCT. Median time from transplant to relapse was 4 months (range = 1–14). Two pts. had failed DLI and 5 pts. had received reinduction chemotherapy prior to IL-2/GM-CSF. Eight pts. responded to IL-2/GM-CSF (CR = 7, PR = 1). Two pts. remain disease free at 18 and 26 months post IL-2/ GM-CSF. Six pts. developed GVHD and of these 4 were responders. Two pts. had GM-CSF discontinued due to increase in peripheral blood blasts. No other toxicities related to IL-2/GM-CSF except for mild flu-like symptoms. The table below summarizes quantitative analysis of immune activation. Values represent means +/− standard error at day 0 (first day of GM-CSF), day 8 (prior to start of IL-2) and day 14 (last day of IL-2 and GM-CSF). P-values are based on paired t-test analysis of day 8 versus day 0 and day 14 versus day 0, respectively. Flow cytometric analysis showed an increase in the numbers of T-lymphocytes (CD3) and T-cell subsets (CD3/CD8 and CD3/CD4) as well as an increase in natural killer cells (CD16/56). Although no differences were seen in the number of dendritic cell subsets, DC1/DC2 ratios decreased with the administration of GM-CSF/IL-2. Limited (n= 4) CD4/FoxP3 analysis did not show change in absolute numbers with administration of GM-CSF/IL-2 (data not shown). In conclusion, cytokine therapy with IL-2/GM-CSF is well tolerated and is an alternative to DLI for relapse after ASCT. Flow cytometry analysis demonstrated a quantitative increase in immune effector cells and polarization to DC2.IMMUNE ACTIVATION FLOW CYTOMETRY ASSAYSD0 (Mean +/− SE)D8 (Mean +/− SE)D14 (Mean +/− SE)P-Value (Day7–0)P-Value (Day 14–0)CD3 (K/uL)309 +/− 117535 +/− 1031306 +/− 4030.0340.027CD3/CD8 (K/uL)94 +/− 42174 +/− 33325 +/− 900.0210.029CD3/CD4 (K/uL)309 +/− 117404 +/− 102977 +/− 3100.2490.045CD16/CD56 (K/uL)124 +/− 60404 +/− 110496 +/− 1620.0290.044CD19 K/uL)68 +/−3989 +/− 36116 +/− 310.1830.044Total Lymphs (K/uL488 +/− 167942 +/− 1602353 +/− 5320.0160.013CD11(DC1) (K/uL)97.1 +/− 60.746.3 +/− 32.532.6 +/− 27.50.1080.101CD123(DC2) (K/uL)32.1 +/− 7.339.7 +/− 15.945.4 +/− 35.80.6940.628DC1/DC22.77 +/− 1.260.68 +/− 0.350.61 +/− 0.07SE=Standard Error; DC = dendritic cells

An Open-Label Trial of Granulocyte Macrophage Colony Stimulating Factor Therapy for Moderate Symptomatic Pulmonary Alveolar Proteinosis

Pulmonary alveolar proteinosis (PAP) is a rare idiopathic autoimmune lung disease in adults characterized by the accumulation of lipoproteinaceous material within the alveoli of the lung. The natural history of this disease is poorly defined. Current therapy of bilateral whole-lung lavage (WLL) under general anesthesia is invasive and has its limitations. Data suggest that relative granulocyte macrophage colony stimulating factor (GM-CSF) deficiency may be involved in the pathogenesis of this disease. There have been several case series that have described clinical improvement with exogenous GM-CSF therapy in a subset of patients with PAP. We describe the results of a prospective, open-label clinical trial of daily subcutaneous GM-CSF therapy in a group of adult patients with idiopathic PAP. In this series of 25 patients, the largest reported to date, administration of GM-CSF improved oxygenation as assessed by a 10 mm Hg decrease in alveolar-arterial oxygen gradient, as well as improvement in other clinical and quality of life parameters in 12 of 25 patients (48%) with moderate symptomatic disease who completed the trial. In addition, the serum anti-GM-CSF antibody titer correlated with lung disease activity and was a predictor for responsiveness to therapy. These data indicate that subcutaneous GM-CSF therapy is a promising alternative to WLL for symptomatic patients with PAP.

Granulocyte-macrophage colony-stimulating factor stimulates arteriogenesis in a pig model of peripheral artery disease using clinically applicable infusion pumps

A growing number of patients suffer from peripheral artery disease (PAD). Current therapies are often limited by the extent of vascular pathology and the occurrence of restenosis after angioplasty. The stimulatory effect of growth factor administration on collateral vessel formation (arteriogenesis) has evolved as a potential new treatment for this patient group. Granulocyte-macrophage colony-stimulating factor (GM-CSF) was shown to stimulate arteriogenesis in small-animal models and in a pilot study in patients with coronary artery disease. Although a recent clinical study demonstrated disappointing results after subcutaneous GM-CSF application in patients with PAD, we hypothesized that intra-arterial cytokine application using implantable infusion pumps might well stimulate arteriogenesis in a large-species model of peripheral vascular disease. We also aimed to compare continuous and intermittent infusion regimens and to validate experimental and clinically available measurements of collateral artery growth. Twenty-four pigs underwent unilateral occlusion of the right femoral artery and received either GM-CSF continuously, GM-CSF intermittently, or phosphate-buffered saline (PBS). After 1 week, collateral conductance was determined under maximal vasodilatation with adenosine and by using a pump-driven extracorporal shunt system. Conductance showed a significant stimulatory effect of GM-CSF on arteriogenesis (collateral conductance [mL/min/mm Hg]: PBS, 37.7 +/- 5.4; GM-CSF continuous, 69.2 +/- 12.5; GM-CSF intermittent, 71.5 +/- 11.1). Flow measurements under reactive hyperemia were consistent with these results (flow occluded/non-occluded hind limb: PBS, 40.5% +/- 9.1%; GM-CSF continuous, 48.9% +/- 3.9%; GM-CSF intermittent, 48.7% +/- 4.4%). Measurements of ankle/brachial indices were not sensitive enough to detect the differences in collateral growth between the three groups. These results demonstrate the proarteriogenic properties of GM-CSF in larger animal species, revealing comparable efficacy of continuous and intermittent intra-arterial infusion. Furthermore, we provide evidence that implantable pumps offer a possible means for the intra-arterial application of growth factors. Intra-arterial application of GM-CSF might be a future treatment option for vascular occlusive disease. Finally, we show that in the peripheral circulation, pressure measurements alone have a low sensitivity to determine the effects of proarteriogenic therapy compared with flow or combined flow-pressure measurements.

Neovasculogenic Therapy to Augment Perfusion and Preserve Viability in Ischemic Cardiomyopathy

Ischemic cardiomyopathy is a global health concern with limited therapy. We recently described endogenous revascularization utilizing granulocyte-macrophage colony stimulating factor (GMCSF) to induce endothelial progenitor cell (EPC) production and intramyocardial stromal cell-derived factor-1alpha (SDF) as a specific EPC chemokine. The EPC-mediated neovascularization and enhancement of myocardial function was observed. In this study we examined the regional biologic mechanisms underlying this therapy. Lewis rats underwent left anterior descending coronary artery (LAD) ligation and developed ischemic cardiomyopathy over 6 weeks. Three weeks after ligation, the animals received either subcutaneous GMCSF and intramyocardial SDF injections or saline injections as control. Six weeks after LAD ligation circulating EPC density was studied by flow cytometry. Quadruple immunofluorescent vessel staining for mature, proliferating vasculature was performed. Confocal angiography was utilized to identify fluorescein lectin-lined vessels to assess perfusion. Ischemia reversal was studied by measuring myocardial adenosine triphosphate (ATP) levels. Myocardial viability was assayed by terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick-end labeling detection of apoptosis and quantitation of myofilament density. The GMCSF/SDF therapy enhanced circulating leukocyte (13.1 +/- 4.5 x 10(6) vs 3.1 +/- 0.5 x 10(6)/cc, p = 0.001, n = 6) and EPC (14.2 +/- 6.6 vs 2.2 +/- 2.1/cc, p = 0.001, n = 6) concentrations. Tetraimmunofluorescent labeling demonstrated enhanced stable vasculature with this therapy (39.2 +/- 8.1 vs 25.4 +/- 5.1%, p = 0.006, n = 7). Enhanced perfusion was shown by confocal microangiography of borderzone lectin-labeled vessels (28.2 +/- 5.4 vs 11.5 +/- 3.0 vessels/high power field [hpf], p = 0.00001, n = 10). Ischemia reversal was demonstrated by enhanced cellular ATP levels in the GMCSF/SDF borderzone myocardium (102.5 +/- 31.0 vs 26.9 +/- 4.1 nmol/g, p = 0.008, n = 5). Borderzone cardiomyocyte viability was noted by decreased apoptosis (3.2 +/- 1.4% vs 5.4 +/- 1.0%, p = 0.004, n = 10) and enhanced cardiomyocyte density (40.0 +/- 5.6 vs 27.0 +/- 6 myofilaments/hpf, p = 0.01, n=10). Endogenous revascularization for ischemic cardiomyopathy utilizing GMCSF EPC upregulation and SDF EPC chemokinesis upregulates circulating EPCs, enhances vascular stability, and augments myocardial function by enhancing perfusion, reversing cellular ischemia, and increasing cardiomyocyte viability.

Aerosol granulocyte-macrophage colony-stimulating factor for pulmonary alveolar proteinosis

Recently, granulocyte-macrophage colony-stimulating factor (GM-CSF) auto-antibodies have been found in many patients with pulmonary alveolar proteinosis (PAP). The present study reports a retrospective case series of patients who used aerosolised GM-CSF in the treatment of idiopathic PAP. Between 1999 and 2003, 12 patients elected to receive aerosolised GM-CSF (250 microg b.i.d. every other week) in lieu of whole-lung lavage or observation. Patient characteristics, pulmonary function tests, arterial blood gas analysis, laboratory values and chest radiographs were extracted from the patient's medical records. Of the six patients tested, all had GM-CSF neutralising antibodies. Additionally, abnormalities in GM-CSF gene expression (one patient), receptor expression (two patients) and ability to upregulate adhesion molecules (one patient) were found. All patients except one had a positive response (mean improvements in arterial oxygen tension, alveolar-arterial oxygen gradient, carbon monoxide diffusing capacity of the lung and forced vital capacity were 17.1 mmHg, 18.4 mmHg, 16.6% pred and 13.5% pred, respectively). Two patients made a complete recovery and were disease free 1 and 2 yrs after discontinuing treatment. Four patients showed complete response to both the initial course or when treated again for recurrence after discontinuation of treatment. One patient required dose escalation (500 microg b.i.d.) with complete response. GM-CSF was well tolerated without late toxicity after median (range) follow-up of 30.5 (3-68) months. In conclusion, aerosolised granulocyte-macrophage colony-stimulating factor is safe and effective in treating pulmonary alveolar proteinosis providing an alternative to whole-lung lavage or subcutaneous granulocyte-macrophage colony-stimulating factor.

A Phase 2 Study of Moderate Dose Interleukin-2 and Granulocyte-Macrophage Colony-Stimulating Factor in Patients With Metastatic or Unresectable Renal Cell Carcinoma

Interleukin-2 (IL-2) has been shown to produce durable complete remission in patients with renal cell carcinoma (RCC). A phase 2 study was conducted to evaluate the potential therapeutic synergy as well as the toxic side effects of the concurrent administration of IL-2 and granulocyte-macrophage colony-stimulating factor (GM-CSF) in patients with advanced stage disease. Twenty-one patients with unresectable or metastatic RCC having an Eastern Oncology Cooperative Group performance status of 0 or 1 were enrolled. Six patients had received prior immunotherapy with interferon (IFN)-alpha, IFN-gamma, and IL-12, whereas the remaining 15 subjects were previously untreated. Thirteen patients were assigned to a moderate-dose bolus of IL-2 at 72,000 IU/kg every 8 hours on days 1 through 5 and days 15 through 19, whereas 8 patients were given IL-2 as an intravenous continuous infusion at a dose of 5 MU/m2/d on days 1 through 5 and days 15 through 19. Subcutaneous GM-CSF at 125 microg/d on days 1 through 21 was administered concomitantly with IL-2. The median number of IL-2 bolus doses was 23 of a scheduled 28 (85%), whereas with the continuous infusion, 93% of planned IL-2 was given. All patients received 100% of GM-CSF doses. There were no complete or partial responses in this study. Of 13 patients treated in the bolus IL-2 arm, 10 had systemic progression of disease at 4 to 8 weeks, 1 developed metastasis in the brain at 4 weeks, and 2 had stable disease for 4 and 17 months. Among the 8 subjects treated with continuous infusion IL-2, 3 progressed with brain lesions at 3 to 8 weeks and 5 had stable disease at 6+, 7, 8+, 15+, and 17+ months. The median survival for the whole group was 10 months, with a range of 0.5 to 40+ months. There were no regimen-related deaths, and most of the observed toxicities were grade 1 and 2. Serious toxicities (grade 3 and 4) included anemia, atrial fibrillation, oliguria, abnormal liver function, and neurologic events like agitation or confusion. The combination of recombinant IL-2 and GM-CSF administered in the designed schedule and doses was not effective in patients with metastatic RCC and may even interfere with the therapeutic potential of moderate-dose IL-2 and increase its adverse events.

Safety and Efficacy of Subcutaneous-Only Granulocyte-Macrophage Colony-Stimulating Factor for Collateral Growth Promotion in Patients With Coronary Artery Disease

This study was designed to investigate the safety and efficacy of a short-term subcutaneous-only granulocyte-macrophage colony-stimulating factor (GM-CSF) protocol for coronary collateral growth promotion. The safety and efficacy of an exclusively systemic application of GM-CSF in patients with coronary artery disease (CAD) and collateral artery promotion has not been studied so far. In 14 men (age 61 +/- 11 years) with chronic stable CAD, the effect of GM-CSF (molgramostim) on quantitatively assessed collateral flow was tested in a randomized, double-blind, placebo-controlled fashion. The study protocol consisted of an invasive collateral flow index (CFI) measurement in a stenotic as well as a normal coronary artery before and after a two-week period with subcutaneous GM-CSF (10 microg/kg; n = 7) or placebo (n = 7). Collateral flow index was determined by simultaneous measurement of mean aortic, distal coronary occlusive, and central venous pressure. Collateral flow index in all vessels changed from 0.116 +/- 0.05 to 0.159 +/- 0.07 in the GM-CSF group (p = 0.028) and from 0.166 +/- 0.06 to 0.166 +/- 0.04 in the placebo group (p = NS). The treatment-induced difference in CFI was +0.042 +/- 0.05 in the GM-CSF group and -0.001 +/- 0.04 in the placebo group (p = 0.035). Among 11 determined cytokines, chemokines, and their monocytic receptor concentrations, the treatment-induced change in CFI was predicted by the respective change in tumor necrosis factor-alpha concentration. Two of seven patients in the GM-CSF group and none in the placebo group suffered an acute coronary syndrome during the treatment period. A subcutaneous-only, short-term protocol of GM-CSF is effective in promoting coronary collateral artery growth among patients with CAD. However, the drug's safety regarding the occurrence of acute coronary syndrome is questionable.

Chemo-Immunotherapy of Metastatic Colorectal Carcinoma With Gemcitabine Plus FOLFOX 4 Followed by Subcutaneous Granulocyte Macrophage Colony-Stimulating Factor and Interleukin-2 Induces Strong Immunologic and Antitumor Activity in Metastatic Colon Cancer Patients

Tumor cell killing by anticancer drugs may be supported by their immuno- and pharmacologic effects. Chemotherapy is in fact able to (A) upregulate tumor-associated antigen expression, including carcinoembryonic antigen (CEA) or other target molecules such as thymidylate synthase (TS); and (B) downregulate tumor cell resistance to the death signals induced by tumor antigen-specific cytotoxic T lymphocytes. This provides the rationale for combining chemo- and immunotherapy. We describe the results of a translational phase II trial designed to evaluate the toxicity, antitumor activity and immunologic effects of gemcitabine + FOLFOX-4 (oxaliplatin, fluorouracil, and folinic acid) polychemotherapy followed by the subcutaneous administration of granulocyte macrophage colony-stimulating factor and low-dose interleukin-2 in colorectal carcinoma patients. The study involved 29 patients (16 males and 13 females with a mean age of 69 years), 21 of whom had received a previous line of treatment, and 19 had liver involvement. The treatment was well tolerated and induced very high objective response (68.9%) and disease control rates (96.5%), with an average time to progression of 12.5 months. An immunologic study of peripheral blood mononuclear cells (PBMCs) taken from 20 patients showed an enhanced proliferative response to colon carcinoma antigen and a significant reduction in suppressive regulatory T lymphocytes (CD4+CD25T-reg+). A cytofluorimetric study of the PBMCs of five HLA-A(*)02.01+ patients who achieved an objective response showed an increased frequency of cytolytic T lymphocyte precursors specific for known CEA- and TS-derived epitopes. The results show that our regimen has strong immunologic and antitumor activity in colorectal cancer patients and deserves to be investigated in phase III trials.

Treatment of Complete Spinal Cord Injury Patients by Autologous Bone Marrow Cell Transplantation and Administration of Granulocyte-Macrophage Colony Stimulating Factor

Transplantation of bone marrow cells into the injured spinal cord has been found to improve neurologic functions in experimental animal studies. However, it is unclear whether bone marrow cells can similarly improve the neurologic functions of complete spinal cord injury (SCI) in human patients. To address this issue, we evaluated the therapeutic effects of autologous bone marrow cell transplantation (BMT) in conjunction with the administration of granulocyte macrophage-colony stimulating factor (GM-CSF) in six complete SCI patients. BMT in the injury site (1.1 x 10(6) cells/microL in a total of 1.8 mL) and subcutaneous GM-CSF administration were performed on five patients. One patient was treated with GM-CSF only. The follow-up periods were from 6 to 18 months, depending on the patients. Sensory improvements were noted immediately after the operations. Sensory recovery in the sacral segment was noted mainly 3 weeks to 7 months postoperatively. Significant motor improvements were noted 3 to 7 months postoperatively. Four patients showed neurologic improvements in their American Spiral Injury Association Impairment Scale (AIS) grades (from A to C). One patient improved to AIS grade B from A and the last patient remained in AIS grade A. No immediate worsening of neurologic symptoms was found. Side effects of GMCSF treatment such as a fever (>38 degrees C) and myalgia were noted. Serious complications increasing mortality and morbidity were not found. The follow-up study with magnetic resonance imaging 4-6 months after injury showed slight enhancement within the zone of BMT. Syrinx formation was not definitely found. BMT and GM-CSF administration represent a safe protocol to efficiently manage SCI patients, especially those with acute complete injury. To demonstrate the full therapeutic value of this protocol, long-term and more comprehensive case-control clinical studies are required.

Phase I/II study of oral etoposide plus GM-CSF as second-line chemotherapy in platinum-pretreated patients with advanced ovarian cancer

The aim of this phase I/II study was to determine the maximum tolerated dose (MTD) and the dose-limiting toxicities of chronic oral etoposide given on days 1–10 followed by rescue with subcutaneous (s.c.) granulocyte-macrophage colony-stimulating factor (GM-CSF) on days 12–19 as second-line chemotherapy in platinum-pretreated patients (pts) with advanced ovarian carcinoma. Cohorts of three to six pts were treated with doses of oral etoposide from 750 mg m−2 cycle−1 escalated to 1250 mg m−2 cycle−1 over 10 days, every 3 weeks. Subcutanous GM-CSF, 400 μg once daily, days 12–19, was added if dose-limiting granulocytopenia was encountered. In total, 18 pts with a median Karnofsky index of 80% (range, 70–100%) and a median time elapsed since the last platinum dose of 10 months (range, 1–24 months), 30% of whom showed visceral metastases, were treated at four dose levels (DLs) of oral etoposide on days 1–10 of each cycle as follows: DL 1, 750 mg m−2 cycle−1, without GM-CSF, three pts; DL 2, 1000 mg m−2 cycle−1, without GM-CSF, three pts; DL 3, 1000 mg m−2 cycle−1, with GM-CSF, six pts; and DL 4, 1250 mg m−2 cycle−1, with GM-CSF, six pts. All pts were assessable for toxicity and 16 pts for response. Dose-limiting toxicity (DLT) was reached at DL 4 by three of six pts, showing World Health Organization (WHO) toxicity grade 4. One patient died from gram-negative sepsis associated with granulocytopenia grade 4. Two more pts developed uncomplicated granulocytopenia grade 4. Thus, we recommend that DL 3 can be used for further phase II evaluation (i.e. oral etoposide 1000 mg m−2 cycle−1, days 1–10, followed by s.c. GM-CSF 400 μg, days 12–19). The clinical complete or partial responses in each patient cohort were: DL 1, one of three pts; DL 2, one of three pts; DL 3, three of five pts; and DL 4, two of five pts. In conclusion, in this phase I/II study, we defined the MTD and the dose recommended for the therapy with oral etoposide given over 10 days followed by s.c. GM-CSF in platinum-pretreated patients with advanced ovarian cancer. Our data demonstrate encouraging activity of this regimen and strongly support its further investigation in a phase II study.

Subcutaneous Granulocyte-Macrophage Colony-Stimulating Factor in Mucositis Induced by an Adjuvant 5-Fluorouracil plus Leucovorin Regimen

Objective: To test the activity of subcutaneous granulocyte-macrophage colony-stimulating factor (GM-CSF) administration in patients with colorectal cancer treated with adjuvant 5-fluorouracil (5-FU) plus leucovorin (LV) and suffering from mucositis. Methods: Thirty-one patients treated with adjuvant 5-FU 370 mg/m² and LV 100 mg/m² for 5 days every 4 weeks and reporting grade ≧2 mucositis participated in the study. Subcutaneous GM-CSF 4 µg/kg/day from days 6 to 10 was administered without chemotherapy dose reductions in the cycle that followed the cycle during which mucositis was reported. The disappearance of mucositis or a decrease by ≧1 grade was recorded as a therapeutic success. Baseline toxicity was: grade 2 stomatitis in 12 patients and grade 3 in 9; grade 4, 3 and 2 diarrhoea in 1, 4 and 8 patients, respectively. Results: Seventy-seven GM-CSF cycles were administered. A success was achieved in 20 (64.5%) patients. The efficacy was assessable in 5 (16.1%) patients with grade 2 and in 8 (25.8%) with grade 3 stomatitis, respectively, as well as in 1 (3.2%) and 5 (16.1%) patients with grade 3 and 2 diarrhoea, respectively. Success (3.2%) was reported in 2 patients suffering from both grade 2 stomatitis and diarrhoea. In 7 (22.5%) patients there was no evidence of efficacy. In 4 (12.9%) patients the treatment was stopped after the first administration of GM-CSF due to a grade 2 allergic reaction. Conclusions: The subcutaneous administration of GM-CSF relieved patients from symptoms of 5-FU/LV-induced mucositis with acceptable side effects permitting the maintenance of full-dose chemotherapy.

Immunotherapy of melanoma.

Over the past decade, tumour immunology has progressed enormously, as a result of the application of a range of new technologies. Screening of tumour gene expression libraries using cancer patients' sera and cancer-specific T lymphocytes has defined a large number of tumour-specific antigenic proteins. The repertoire of known tumour antigens is rapidly expanding as a consequence of these advances. In addition, over the last few years the analysis of human T-cell responses specific for several tumour antigens in cancer patients has been made possible by the use of novel staining reagents (i.e. tetramers). This has allowed the characterization and monitoring of specific tumour responses, and has provided an opportunity to greatly accelerate the development of new anticancer vaccines. Recent results have demonstrated that the immune system can detect the presence of malignant cells in cancer patients, and have raised hopes for the use of antigenic cancer proteins as vaccines to induce tumour-specific cell-mediated immunity. Tumour immunology is now on solid technical and conceptual footing. However, the challenge remains to apply to patients the results obtained in the laboratories.

Idiotype Vaccination in Human Myeloma: Generation of Tumor-Specific Immune Responses After High-Dose Chemotherapy

Igs contain unique portions, collectively termed idiotypes (Id), that can be recognized by the immune system. Id expressed by tumor cells in B-cell malignancies can be regarded as tumor-specific antigens and a target for vaccine immunotherapy. We have started a vaccination trial in multiple myeloma (MM) using Id-specific proteins conjugated to keyhole limpet hemocyanin (KLH) as immunogens and low doses of subcutaneous granulocyte-macrophage colony-stimulating factor (GM-CSF) or interleukin-2 (IL-2) as immunoadjuvants. Twelve patients who had previously been treated with high-dose chemotherapy followed by peripheral blood progenitor cell (PBPC) transplantation entered this study from August 1995 to January 1998. All patients were in first remission at the time of vaccination. They received subcutaneous injections of Id vaccines and immunoadjuvants in an outpatient setting. The generation of Id-specific T-cell proliferative responses was documented in 2 patients, whereas a positive Id-specific delayed-type hypersensitivity (DTH) reaction was observed in 8 of the 10 patients studied. DTH specificity was confirmed in 1 patient by investigating the reactivity to synthetic peptides derived from the VDJ sequence of the tumor-specific Ig heavy chain. None of the patients generated soluble immune responses to Id, whereas the generation of soluble and cellular immune responses to KLH was observed in 100% and 80%, respectively. Eleven patients completed the treatment, whereas 1 patient failed to finish owing to progression of disease. Freedom from disease progression (FFDP), measured from the date of first Id/KLH injection to the date of first treatment after vaccination or last follow-up, ranged from 9 to 36 months. These data indicate that the immune competence status of MM patients is still susceptible to specific immunization after high-dose chemotherapy and PBPC transplantation. It remains to be determined whether generation of Id-specific immune responses can reduce the relapse rate of patients with minimal residual disease.

Idiotype Vaccination in Human Myeloma: Generation of Tumor-Specific Immune Responses After High-Dose Chemotherapy

AbstractIgs contain unique portions, collectively termed idiotypes (Id), that can be recognized by the immune system. Id expressed by tumor cells in B-cell malignancies can be regarded as tumor-specific antigens and a target for vaccine immunotherapy. We have started a vaccination trial in multiple myeloma (MM) using Id-specific proteins conjugated to keyhole limpet hemocyanin (KLH) as immunogens and low doses of subcutaneous granulocyte-macrophage colony-stimulating factor (GM-CSF) or interleukin-2 (IL-2) as immunoadjuvants. Twelve patients who had previously been treated with high-dose chemotherapy followed by peripheral blood progenitor cell (PBPC) transplantation entered this study from August 1995 to January 1998. All patients were in first remission at the time of vaccination. They received subcutaneous injections of Id vaccines and immunoadjuvants in an outpatient setting. The generation of Id-specific T-cell proliferative responses was documented in 2 patients, whereas a positive Id-specific delayed-type hypersensitivity (DTH) reaction was observed in 8 of the 10 patients studied. DTH specificity was confirmed in 1 patient by investigating the reactivity to synthetic peptides derived from the VDJ sequence of the tumor-specific Ig heavy chain. None of the patients generated soluble immune responses to Id, whereas the generation of soluble and cellular immune responses to KLH was observed in 100% and 80%, respectively. Eleven patients completed the treatment, whereas 1 patient failed to finish owing to progression of disease. Freedom from disease progression (FFDP), measured from the date of first Id/KLH injection to the date of first treatment after vaccination or last follow-up, ranged from 9 to 36 months. These data indicate that the immune competence status of MM patients is still susceptible to specific immunization after high-dose chemotherapy and PBPC transplantation. It remains to be determined whether generation of Id-specific immune responses can reduce the relapse rate of patients with minimal residual disease.

Effects of continuous localized infusion of granulocyte-macrophage colony-stimulating factor and inoculations of irradiated glioma cells on tumor regression.

Glioblastoma multiforme (GBM) is a malignant tumor of the central nervous system that directly suppresses immunological defenses in vitro and in vivo. The authors used the peripheral delivery of continuously infused granulocyte-macrophage colony-stimulating factor (GM-CSF) in the presence of irradiated tumor antigens as a tumor-specific stimulant to dendritic cells to initiate an immune response to GBM in rats. The 9L gliosarcoma tumors were established in the flanks of syngeneic Fischer 344 rats. Osmotic minipumps implanted in the animals' contralateral flanks continuously delivered recombinant GM-CSF (0, 0.1, 1, or 10 ng/day) for 28 days. Irradiated gliosarcoma cells were intermittently injected at the site of the GM-CSF infusion. Animals in the saline control group (0 ng/day GM-CSF) died on Day 59 with average tumor volumes greater than 30,000 mm3. This control group was significantly different from the GM-CSF-treated animals, which all survived with average tumor volumes that peaked on Day 23 and later regressed completely. Tumor growth as well as peak tumor volumes (5833+/-2284 mm3, 3294+/-1632 mm3, and 1979+/-1142 mm3 for 0.1, 1, and 10 ng/day GM-CSF, respectively) in the different treatment groups reflected a significant dose-response relationship with the GM-CSF concentrations. All animals treated with GM-CSF and irradiated cells were resistant to additional challenges of peripheral and intracerebral gliosarcoma, even when they were inoculated 8 months after initial immunotherapy. The colocalization of GM-CSF and inactivated tumor antigens was required to stimulate immunoprotection. To test the efficacy of a peripherally administered immunological therapy on intracerebral brain tumors the authors transplanted 10(6) gliosarcoma cells into the striatum of treated and control animals. Subcutaneous pumps that released GM-CSF (10 ng/day) and irradiated gliosarcoma cells were placed in the treated animals. The control animals all died within 31 days after intracerebral tumor implantation. In contrast, 40% of the animals receiving GM-CSF-irradiated cell vaccinations survived beyond 300 days. These long-term survivors showed no evidence of gliosarcoma at the injection site on evaluation by magnetic resonance imaging. These results suggest that the continuous localized delivery of subcutaneous GM-CSF in conjunction with inactivated tumor antigens can initiate a systemic response that leads to the regression of distant peripheral and intracerebral tumors. The success of this treatment illustrates the feasibility of tumor-specific peripheral immunological stimulation after tumor resection to prevent the recurrence of malignant brain tumors.

Granulocyte-macrophage colony-stimulating factor as infection prophylaxis in high-risk oncologic surgery

A method of augmenting host defenses against bacterial pathogens could result in a decrease in postoperative infections. Given its effects on leukocyte proliferation and function, it is possible that prophylactic granulocyte-macrophage colony-stimulating factor (GM-CSF) could reduce the incidence and severity of infections in high-risk surgical patients. The current study was undertaken to determine the safety and hematologic effects of perioperative GM-CSF. Cancer patients undergoing operations with a high risk of postoperative infection were treated perioperatively for 10 days with subcutaneous GM-CSF. Cohorts were treated with GM-CSF at 125 micrograms/m2/day (12 patients) and 250 micrograms/m2/day (11 patients). There were no severe or life-threatening toxicities associated with GM-CSF. Mean maximum neutrophil counts during the first 5 postoperative days were 16.3 +/- 9.14 and 24.5 +/- 7.60 at 125 and 250 micrograms/m2, respectively (P = 0.04). Only one wound infection was diagnosed during this study. GM-CSF may be safely administered perioperatively at doses that augment neutrophil number and function. An ongoing randomized clinical trial will determine the impact of GM-CSF on postoperative infection.