ELISA methodology for detection of modified osteoprotegerin in clinical studies.
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Animals↗
Biomedical subjects
H Gunn
Publications and source records attributed to H Gunn.
Immunogenicity of recombinant human interleukin-3.
Most recombinant human (rh) proteins elicit an immune response in experimental animals. Antibodies are usually detected within 1 to 2 weeks of protein administration. The immunogenicity of rh proteins is an important aspect for both experimental and clinical investigations. The formation of neutralizing antibodies-as shown here for rhIL-3-reduced the absolute bioavailability of the protein in Rhesus monkeys.
Animals↗
Phase I trial of subcutaneous interleukin 3 in patients with refractory malignancy: hematological, immunological, and pharmacodynamic findings.
We conducted a Phase I trial of s.c. recombinant human interleukin 3 (rhIL-3) to evaluate the toxicity, maximal tolerated dose, pharmacokinetics, and in vivo biological effects of this cytokine. Thirty-one patients with refractory cancer were entered into the study between November 1991 and June 1993. Therapy consisted of s.c. rhIL-3 daily for 15 days administered to cohorts of three to nine patients at dose levels of 60-4000 microgram/m2/day. Cycles were repeated at intervals of 28 days. Seventy-five cycles of rhIL-3 were administered (median, two per patient) and the maximal tolerated dose was 2000 microgram/m2/day. Toxicity was moderate, with most patients developing chills, fever, and myalgia. Dose-limiting toxicity consisted of diarrhea (two patients) and headache (one patient). Hematological effects of rhIL-3 included significant dose-related increases of WBC (P < 0.001), neutrophils (P < 0.001), and eosinophils (P < 0.001). Platelet counts and absolute lymphocyte numbers also increased. Various CD3(+) lymphocyte subsets increased; however, lytic activity (natural killer and lymphokine-activated killer) of peripheral blood lymphocytes was not enhanced. Serum levels of the soluble IL-2 receptor increased in a dose-related fashion, and IL-2-induced lymphocyte proliferation also was increased variably. Pharmacokinetic studies were performed in 13 patients, and area under the curve and maximal concentration values increased with increasing rhIL-3 dose levels (P < 0.001) and correlated with maximal changes from baseline in WBC, neutrophils, and eosinophils. rhIL-3 antibodies were detected in 8% of patients by day 29 of cycle 1 but were not neutralizing. rhIL-3 is well tolerated when administered s.c. and has reproducible hematological and immunological effects. The pleiotropic effects of this cytokine on various in vivo biological parameters were demonstrated clearly. Further studies of its immunoregulatory effects are warranted.
Adult↗
Pharmacodynamics of daily subcutaneous recombinant human interleukin-3 in normal volunteers.
Normal volunteers received subcutaneous injections of recombinant human interleukin-3 (rhIL-3) on 4 consecutive days to characterize toxicity, pharmacokinetics, and hematopoietic effects. Dosages were 2.5, 5.0, and 7.5 micrograms/kg/day (n = 6 subjects per group). Adverse effects consisted predominantly of flu-like symptoms such as fever and headache. Mean area under the serum concentration-time curve and maximum serum concentration were linearly related to dose. Serum clearance was not apparently related to dose. Clearance increased slightly but significantly between days 1 and 4. Rapid but modest elevations in neutrophil and eosinophil counts were observed during treatment. Mean platelet counts rose modestly, peaking on day 10. Increases of CD34+ cell counts were correlated with increases of colony-forming unit-granulocyte macrophage (peak, day 7).
Adult↗
Phase I multicenter trial of interleukin 6 therapy after autologous bone marrow transplantation in advanced breast cancer.
Our purpose was to determine the maximum tolerated dosage of rhIL-6 after high-dose cytotoxic chemotherapy and autologous BMT in patients with advanced breast cancer. Twenty patients (median age 43.5 years) received either CY and thiotepa (n = 3) or CY, thiotepa and carboplatin (n = 17) for 4 days. Unpurged autologous BM was reinfused 72 h later. Daily rhIL-6 therapy began the day of marrow infusion and continued until recovery of neutrophils (> or = 1.5 x 10(9)/l) and platelets (> or = 50 x 10(9)/l) or for a maximum of 28 days at a dosage of 0.3 microgram/kg/day (n = 7), 1 microgram/kg/day (n = 6) or 3 micrograms/kg/day (n = 7). Two of the initial 4 patients given rhIL-6 at 0.3 mu/kg i.v. experienced grade 4 hyperbilirubinemia, so subsequent patients received s.c. rhIL-6. Most toxicities attributable to rhIL-6 were reversible or mild constitutional symptoms, but dose-limiting grade 4 hyperbilirubinemia also occurred in 3 of the 7 patients receiving the 3 micrograms/kg dose. At the 0.3 and 1 microgram/kg/day doses, 8 of 13 patients completed the study vs. only 2 of 7 at the 3 micrograms/kg/day dose. During rhIL-6 treatment, neutrophil recovery (> or = 500 x 10(6)/l) occurred in 12 patients and platelet recovery (> or = 20 x 10(9)/l) occurred in 6 patients, 5 of whom received the 0.3 or 1 microgram/kg/day s.c. dose. The maximal tolerated dose of rhIL-6 after autologous BMT appeared to be 1 microgram/kg/day s.c., a dose appreciably lower than the maximal tolerated dose after conventional cytotoxic therapy.
Adenocarcinoma↗
A phase I trial of intravenous interleukin-6 in patients with advanced cancer.
Eighteen patients were treated with escalating doses of recombinant, Escherichia coli-derived human interleukin-6 (IL-6) intravenously every 8 h. Therapy was given for two cycles of 7 days each separated by a week off therapy. Fevers and chills were observed in most patients. Mild renal and liver function abnormalities were noted at higher doses of IL-6. Dose-limiting toxicity was reached at 30 micrograms/kg i.v. every 8 h due to reversible neurotoxicity, but significant rapidly reversible anemia and hyperglycemia were seen at lower doses. Platelet counts, white blood cell counts, and acute phase reactant levels were substantially elevated. No antitumor responses were seen. A maximum tolerated dose of 10 micrograms/kg i.v. every 8 h for two 7-day cycles is recommended for future phase II trials.
Acute-Phase Proteins↗
Primate antibodies to components of the human immune system.
The feasibility to raise nonhuman primate antibodies against selected components of the human immune system was tested. The immunogens were whole cells (human T lymphocytes) or purified, recombinant human proteins (cytokines: TNF alpha or GM-CSF; soluble forms of cell surface antigens: sCD4 or sCD25). Significant immunizations, yielding functionally relevant antibodies, were readily achieved in rhesus monkeys, but, not surprisingly, may be less frequent in chimpanzees. The results suggest a general strategy for production of therapeutically useful MAB.
Animals↗
Hypothalamic-pituitary-adrenal axis activation and stimulation of systemic vasopressin secretion by recombinant interleukin-6 in humans: potential implications for the syndrome of inappropriate vasopressin secretion.
We recently demonstrated that sc administered interleukin-6 (IL-6) strongly stimulates the human hypothalamic-pituitary-adrenal (HPA) axis, with mild toxicity and no hypotensive effects. In this study, we evaluated the response of the human HPA axis to escalating iv doses of recombinant IL-6 in six patients with cancer and good performance status who received daily, every 8 h, three equal doses of 0.3-30 micrograms/kg IL-6. The plasma levels of IL-6 assayed by a specific enzyme-linked immunosorbent assay during the 4 h following the first IL-6 injection were elevated for 2-4 h, proportionally to the amount of injected IL-6. Administration of the cytokine was followed by marked elevations of plasma ACTH (53.0-98.6 pmol/L) and cortisol (824.9-1729.9 nmol/L) independently of the IL-6 dose administered, suggesting that the doses employed were at the top of the dose-response curve for these hormones. Interestingly, plasma arginine vasopressin (AVP) levels were also elevated during the 2 h after IL-6 injection in all patients who received a dose of 3 micrograms/kg or more, suggesting that IL-6 activated the magnocellular AVP-secreting neurons and that it might be involved in the syndrome of inappropriate AVP secretion. Cortisol elevations with peaks similar to those observed after the first injection of IL-6 were also detected in plasma sampled every 2 h after the second and third injections, suggesting that there was no rapid tachyphylaxis in response to IL-6 administration. Plasma IL-1 beta and tumor necrosis factor-alpha concentrations, assayed by specific enzyme-linked immunosorbent assays during the 4 h after the first IL-6 injection, were either within the normal range or undetectable, confirming in vitro observations that IL-6 does not stimulate IL-1 beta or tumor necrosis factor-alpha secretion and suggesting that it exerts its effect on the HPA axis and AVP secretion independently of them. We conclude that IL-6 is a potent stimulator of the human HPA axis and a secretagogue of magnocellular AVP secretion, which might be employed as a challenge test of the axis and the magnocellular AVP neuron.
Adrenocorticotropic Hormone↗
Interleukin-6 exacerbates glomerulonephritis in (NZB x NZW)F1 mice.
The ability of interleukin-6 (IL-6) to modulate immune parameters and mesangial cell function suggests a role for this cytokine in the development of autoimmune glomerulonephritis. This hypothesis was tested in 6-month-old female (NZB x NZW)F1 mice that were administered recombinant human IL-6 (rhIL-6) (50 and 250 micrograms/kg s.c.) for 12 weeks, resulting in an accelerated and severe form of membranoproliferative glomerulonephritis associated with marked upregulation of mesangial major histocompatibility complex class II antigen and glomerular ICAM-1 expression. To distinguish direct effects of rhIL-6 on the renal mesangium from those mediated through the immune system, (NZB x NZW)F1 mice were immunosuppressed with cyclosporin. Immunosuppression by cyclosporin inhibited the development of glomerulonephritis, decreased class II antigen expression, and abrogated IL-6-mediated effects. Administration of neutralizing anti-IL-6 antibody had no effect on the spontaneous development of glomerulonephritis in (NZB x NZW)F1 mice. This finding, together with undetectable IL-6 serum levels, makes a pathogenetic role of endogenously produced IL-6 in this disease model unlikely. In contrast to (NZB x NZW)F1 mice, parental NZW or BALB/c mice given high doses of rhIL-6 (500 micrograms/kg) or recombinant murine IL-6 (100 micrograms/kg) daily for 4 weeks failed to develop morphological or biochemical evidence of glomerulonephritis. Induction of acute phase proteins, anemia, thrombocytosis, and induction of renal class II antigen confirmed the biological activity of IL-6 in these mice. In conclusion, while non-nephritogenic in normal mice, IL-6 accelerates the development of the genetically determined glomerulonephritis of (NZB x NZW)F1 mice through effects mediated by a modulated immune system. Since neutralizing IL-6 antibody treatment did not prevent the development of glomerulonephritis, it is unlikely that increased IL-6 production plays a role in the pathogenesis of lupus nephritis.
Animals↗
Phase I trial of subcutaneous interleukin-6 in patients with advanced malignancies.
PURPOSE: Based on preclinical evidence in murine models that interleukin-6 (IL-6) mediates regression of metastatic tumors, we performed a phase I study of recombinant human IL-6 in patients with refractory advanced malignancies to determine its pharmacokinetics, toxicities, and possible immunologic and antitumor effects. PATIENTS AND METHODS: Recombinant IL-6 was administered as a single subcutaneous dose daily for 7 days, with 7 days off therapy followed by another 7 days of IL-6. Doses were escalated in cohorts of three patients starting at 3 micrograms/kg/d, provided that toxicity at the preceding dose level was not dose-limiting. Dose-limiting toxicity was defined as grade III or IV major organ toxicity that did not resolve to grade II or less in 24 hours after stopping IL-6, using the National Cancer Institute Common Toxicity Criteria. Patients were treated with 3, 10, and 30 micrograms/kg/d IL-6 subcutaneously. RESULTS: Three patients each were treated at the 3- and 10-micrograms dose levels. Two of five patients treated with 30 micrograms/kg/d IL-6 subcutaneously had grade III major organ toxicity that required IL-6 therapy to be discontinued. All patients experienced fever, chills, and minor fatigue. Significant increases in C-reactive protein (CRP), fibrinogen, platelet counts, and lymphocyte IL-2 receptor levels were seen in patients at the 10- and 30-micrograms/kg dose levels. Decreases in albumin and hemoglobin were observed, particularly at the 30-micrograms/kg dose level. The half-life (T1/2 beta) was 4.2 hours, with a peak IL-6 level at 5 hours. No antitumor responses were seen. CONCLUSION: A safely tolerated dose of daily subcutaneous IL-6 is 10 micrograms/kg, with hepatotoxicity and cardiac arrhythmia being the dose-limiting toxicities at 30 micrograms/kg. Phase II trials of IL-6 administered subcutaneously daily for at least 7 days for two cycles with an intervening week of rest are recommended for phase II trials. However, patients with extensive replacement of liver by tumor and abnormal liver functions should receive IL-6 therapy with caution.
Acute-Phase Proteins↗
Graft-versus-host disease associated with intestinal transplantation in the rat. Host immune function and general histology.
Graft-versus-host disease was studied on the 10th and 14th postoperative days in Lewis x Brown Norway F1 rats (LBN-F1) receiving Lewis accessory heterotopic intestinal allografts. LBN-F1 isograft recipients and LBN-F1 rats were used as controls. The rats were injected with sheep erythrocytes five days before sacrifice. Rats with graft-versus-host disease had progressive loss of the normal architecture of the lymphoid organs. Skin, liver, colon, and salivary glands were infiltrated with immunoblasts and had patchy areas of necrosis. Concurrent with these changes, there were significant, progressive reductions in hemolytic titers, splenocyte plaque-forming counts, viable splenocytes, and the in-vitro splenocyte response to stimulation with concanavalin A. Graft-versus-host disease following intestinal allotransplantation damages the host's lymphoid tissues, producing profound immunosuppression. This finding has implications for clinical intestinal transplantation.
Animals↗
Graft-versus-host reactions induced by intestinal transplantation in the rat produce host immunosuppression.
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Animals↗
Functional evaluation of murine allogeneic T lymphoblasts separated by Vicia villosa lectin positivity.
Mixed-lymphocyte culture-stimulated cells have been fractionated by their ability to bind the lectin Vicia villosa (Vv) and assessed for their cytolytic and suppressor activity in vitro. Vv positive and negative cells were separated either by cell affinity chromatography using Vv-Sepharose 6MB chromatography or by electronic cell sorting with FITC-Vv. Both populations expressed marked cytolytic and suppressor cell activity. Thus this lectin cannot be used to discriminate between these and other functional lymphoid cell population of blastoid cells binding the FITC-Vv appears following allogeneic stimulation; treatment with the immunosuppressive drug cyclosporin A, which affects cytotoxic cells preferentially, results in a considerable reduction of the Vv positive blastoid cells.
Animals↗
Effect of in vitro cyclosporin. A treatment on human natural and antibody-dependent cell-mediated cytotoxicity.
Concentrations of 0.1 to 10.0 micrograms of cyclosporin A (Cy A) per ml do not directly inhibit the lysis of 51Cr-labeled K562 target cells by human peripheral blood natural killer (NK) cells nor of antibody-coated chicken red blood cells by antibody-dependent cellular cytotoxicity (ADCC) effector cells. Pretreatment of effector cells with Cy A for 18 hr still failed to affect NK or ADCC effector function. However, induction of NK cell activity by overnight pretreatment of effector cells by polyinosinate-polycytidylate (PolyIC) is inhibited by as little as 1 micrograms of Cy A per ml. The mechanism of inhibition may involve a direct effect of Cy A on "induced" or "activated" NK cells or inhibition of the release of Poly IC-induced interferon from lymphocytes.
Antibody-Dependent Cell Cytotoxicity↗
Direct staining of mouse T lymphoblasts with fluoresceinated Vicia villosa lectin.
Fluorescein conjugated Vicia villosa (Vv) lectin was used for direct staining of the surface of viable cells of various mouse lymphocyte populations. A varying proportion of polyclonally-activated T cells expressed Vv receptor although less strongly than cells of an influenza virus-specific cytotoxic T lymphocyte (CTL) clone and blasts generated in the mixed lymphocyte reaction. Cyclosporin A (CyA) treatment markedly reduced the expression of Vv receptor following concanavalin A (Con A) stimulation by between 66% and 93% with a concurrent inhibition of blastogenesis and complete abrogation of cytolytic function. Resting mouse lymphocytes and B-cell blasts were always Vv-negative. However, non-specific suppressor factor producing non-cytotoxic T-cell lines also expressed Vv receptor as shown by the weak, but specific, surface fluorescence of EL4 and BW5147 cells stained with Vv. Vv-positive cells were not restricted to a particular Ly phenotype, Vv-positive cells being found among both Lyt 1+ and Lyt2+ MLC stimulated lymphocytes. Our data suggest that the receptor for Vv lectin cannot be regarded as an exclusive differential marker for CTL.
Animals↗
Phase 1 trial of subcutaneous IL-6 in patients with refractory cancer: clinical and biologic effects.
The authors evaluated the clinical and biologic effects of human recombinant interleukin-6 (rhIL-6) in patients with refractory cancer. A phase 1 trial using escalating doses of rhIL-6 (1-50 microg x kg(-1) x d(-1), Monday through Friday for 4 weeks) was performed in 30 patients. Toxicity was moderate and the maximum tolerated dose was determined to be 25 microg x kg(-1)x d(-1) based on cardiac and neurocortical toxicity in one patient each and thrombocytosis (platelets > 800,000/microL) in three patients. One patient with non-small-cell lung cancer had a partial response after three cycles of therapy. The biologic effects of rhIL-6 included anemia and dose-related thrombocytosis. Various proinflammatory activities were induced and included dose-related cyclical increases in peripheral blood monocytes and the CD14+/CD45RB+ +/- CD16C+ mononuclear cell populations. These increases were accompanied by increased levels of C-reactive protein, serum neopterin, and type I soluble tumor necrosis factor receptor. In contrast, rhIL-6 did not affect lymphocyte numbers or function (cytotoxicity, cytokine levels, immunoglobulin levels), with the possible exception of IL-2Ralpha mRNA induction in peripheral blood lymphocytes. rhIL-6 has pleiotropic proinflammatory actions in vivo and moderate toxicity when administered as long-term therapy.
Acute-Phase Reaction↗
Hypermethylation of HpaII recognition sequences within the 5' coding region of the estrogen receptor gene is not associated with estrogen receptor negativity in primary breast tumours.
Estrogen receptor negative tumours are unable to respond to antiestrogen therapy. The underlying molecular mechanisms of estrogen receptor negativity are poorly understood. Cytosine mechylation is one of the mechanisms of gene control and previous studies, particularly on breast tumour derived cell lines have suggested that hypermethylation of HpaII recognition sequences within the 5' coding region of estrogen receptor may be responsible for gene inactivity. This study has examined the methylation status of HpaII recognition sequences in estrogen receptor positive and negative breast tumours taking into account a polymorphic HpaII site in the 5' coding region of the estrogen receptor gene. It is concluded that hypermethylation of one or more of the 4 or 5 HpaII recognition sequences in the 5' coding region of estrogen receptor is not associated with ER negativity in primary breast tumours.
Adenocarcinoma↗