A Phase I and Pharmacokinetic Study of Subcutaneously-Administered Recombinant Human Interleukin-4 (rhuIL-4) in Patients with Advanced Cancer

Abstract

Purpose To investigate the pharmacokinetics and tolerability of recombinant human interleukin-4 (rhuIL-4), administered by daily subcutaneous injection, in patients with advanced cancer.

Patients and Methods Fourteen patients with advanced cancer treated with rhuIL-4 at escalating dose levels of 0.25, 1.0 and 5.0 μg/kg/day, on days 1, 8-17, and 28-57. The primary endpoints of the study were toxicity of rhuIL-4 and the determination of the pharmacokinetics of rhuIL-4 when given by subcutaneous injection. Secondary endpoints included effects on blood counts, hematopoietic cell precursors, and various immunologic parameters.

Results rhuIL-4 was well tolerated at all three dose levels. Detectable serum levels of IL-4 were found in patients at the 1.0 and 5.0 μg/kg/day dose levels. Peak serum IL-4 levels were achieved about 2 h after injection and IL-4 was still detectable 8 h after injection. No grade 4 toxicities were observed and grade 3 toxicities were confined to fever, headache and raised hepatic alkaline phosphatase. No consistent hematological or immunologic effects were observed. Although therapeutic efficacy was not an endpoint, one complete response (Hodgkin's disease) was observed. One patient with chronic lymphocytic leukemia progressed on therapy.

Conclusion rhuIL-4 up to 5.0 μg/kg/day is well tolerated when given by subcutaneous injection. Biologically relevant serum IL-4 levels can be achieved and sustained for at least 8 h after a single injection.     

Development of a recombinant growth factor and fusion protein: lessons from GM-CSF.

Several colony stimulating factors (CSFs) and cytokines have been successfully used to mobilize hematopoietic cells during myeloablative therapy, bone marrow failure, and transplantation and to provide supportive treatment during sepsis. The use of yeast-derived recombinant human granulocyte-macrophage CSF (rhuGM-CSF) and its interleukin-3 fusion protein, PIXY321, provides an example of issues associated with development programs for recombinant hematopoietic growth factors. Species specificity of rhuGM-CSF, different bioactivity of homologous molecules in mice, and production in laboratory animals of antibodies to human proteins limit preclinical evaluation of such molecules. In clinical trials, rhuGM-CSF was efficacious and well tolerated. The derivation of the recombinant molecule, optimal dosing, scheduling, and confounding effects of concurrent disease and treatments are factors that influence efficacy, adverse responses, and immunogenicity reported in patients treated with CSFs. In comparisons of yeast-derived with Escherichia coli-derived rhuGM-CS, the reduced severity and frequency of all adverse events, preponderance of low-grade adverse events, and similarity of positive clinical response versus adverse events reported for granulocyte CSF support safety and efficacy of yeast-derived rhuGM-CSE Enhanced pharmacoeconomic evaluations are beginning to limit and redirect clinical applications in this class of biological agents.     

Histomorphologic observations for cynomolgus monkeys after subchronic subcutaneous injection of recombinant human interleukin-4.

Recombinant human interleukin-4 (rhuIL-4) is a candidate for the treatment of refractory cancer based on its potential to enhance the function of the immune system. Total daily dosages of 0 (placebo control), 1, 5, or 25 micrograms/kg of rhuIL-4 were given as divided (b.i.d.) subcutaneous dosages to male and female cynomolgus monkeys (5/sex/group) for 1 month followed by a 2-week recovery. Histomorphologic evaluation of 3/sex/group at 1 month revealed vascular lesions, granulocytic hyperplasia, and seminiferous tubular atrophy attributed to treatment with rhuIL-4. Dosage-dependent proliferative and inflammatory vascular lesions with eosinophil infiltration affected principally the arterial tree. After 2 weeks of recovery, these lesions, including chronic endarteritis and chronic and/or obliterative arteritis, occurred with an overall lower incidence, and were not observed for monkeys from the 1.0 micrograms/kg/day group. Granulocytic hyperplasia in bone marrow observed for monkeys from all groups given rhuIL-4 at 1 month was not present after 2 weeks of recovery. Seminiferous tubular atrophy was observed for monkeys from the 5 and 25 micrograms/kg/day groups at 1 month and after 2 weeks of recovery.     

Measles as a potential oncolytic virus

The use of replicating viruses for cancer therapy is attracting increasing interest. Numerous viruses are now being considered as potential cancer therapeutics, including the vaccine strain of measles virus (MV). The attenuated strain of measles readily lyses transformed cells, whilst replication and lysis are limited in normal human cells. It has a number of features which make it highly suitable for further development as an oncolytic agent, among them stability and a long history of safety in human use. These features are being combined with its ready potential for genetic manipulations to generate recombinant MVs with desirable therapeutic attributes. This review summarises the pre-clinical studies of the oncolytic efficacy of MV to date. Promising developments in MV engineering--such as re-targeting MV entry to specific cell types and enhancing its utility as a therapeutic agent by expression of non-viral proteins--as well as outstanding issues, such as the role of anti-MV immunity, are highlighted.     

Expression and localization of rabbit B-cell activating factor (BAFF) and its specific receptor BR3 in cells and tissues of the rabbit immune system

Rabbits are widely used for vaccine development, and investigations of human infectious and autoimmune diseases such as Systemic Lupus Erythematosus (SLE). For these applications, we cloned, sequenced and expressed rabbit B-cell Activating Factor (BAFF), and localized BAFF in cells and tissues of the rabbit immune system. The rabbit homolog of the human BAFF binding site (miniBR3 peptide) within the BAFF-specific receptor BR3 was synthesized. This 26-residue core domain binds to recombinant rabbit BAFF protein. Flow cytometric analyses using purified recombinant rabbit BAFF combined with real-time PCR findings revealed that BAFF detected on peripheral blood B-cells from normal rabbits is probably complexed to BAFF receptors rather than produced by the B-cells. BAFF was detected in developing appendix of young rabbits by immunohistochemical staining suggesting that BAFF plays a role during the period following birth when rabbit B-cell development and pre-immune antibody repertoire diversification and selection is occurring.     

Immunoregulation in the common marmoset, Calithrix jaccus: functional properties of T and B lymphocytes and their response to human interleukins 2 and 4.

Non-human primates have been used to study immune function to a much lesser extent than readily available strains of inbred rodents. Nevertheless, in situations where it might be desirable, but impossible, to study human immune responses in vivo, lower primates could provide an acceptable alternative. In order to extent the knowledge of T- and B-lymphocyte function in lower primates, the common marmoset Callithrix jaccus was used as an experimental model. The functional similarities between this species and humans at the level of T-B co-operation in the antibody response were examined, and xenoreactive T-lymphocyte clones were obtained from marmoset spleen cells using Epstein-Barr virus (EBV)-transformed human B cells as stimulators. These clones could act as helper cells when co-cultured with human B lymphocytes, inducing the secretion of both IgM and IgG. Lymphokine production by mitogen-stimulated marmoset T-cell clones was also examined. Interleukins (IL) 2 and 4 activities were detected in clone supernatants using bioassays and interferon-gamma (IFN-gamma) was detected using a solid-phase ELISA system. However, SDS-PAGE analysis of biosynthetically labelled marmoset and human T-cell clone supernatant proteins revealed major differences between the soluble T-cell products of the two species. The proliferative responses of marmoset T and B cells to recombinant human IL-2 and IL-4 were also examined. Stimulation of [3H]thymidine uptake was detected in both T cell- and anti-IgM-stimulated B-cell cultures with both of the lymphokines. These results suggests that the key components of the antibody response are functionally conserved between lower primates and man and that the common marmoset may be useful as an in vivo model of immune function, particularly with regard to the role of interleukins such as IL-2 and IL-4.     

Lymphokine gene therapy for cancer

The plethora of recombinant lymphokines that have recently become available has led to renewed hope for an immunotherapeutic solution to cancer. Many lymphokines, either singly or in combination, have already shown promise in animal trials and in preliminary human trials. Systemic lymphokine toxicity has been the major constraint on this type of therapy, often precluding the use of doses sufficient to induce tumour regression. To realize the therapeutic potential of recombinant lymphokines against cancer, alternative modes of delivery are needed which maximally stimulate local anti-tumour responses whilst causing minimal systemic toxicity. In this article, Stephen Russell proposes that tumour cell targeted lymphokine gene therapy would optimize lymphokine delivery. Some of the practical difficulties likely to be encountered with such an approach are also discussed.     

Extension of the life span of pressure ulcer fibroblasts with recombinant human interleukin-1 beta.

Recombinant human interleukin-1 beta (rhuIL-1 beta) was investigated in a randomized, blinded placebo-controlled trial to evaluate its effect on the healing of chronic pressure ulcers. The influence of this topically applied cytokine to 26 pressure ulcer patients was correlated with tissue culture and electron microscopic evaluation. Cellular replication studies showed that low (0.01 micrograms/cm2/day) and medium (0.1 micrograms/cm2/day) concentrations of rhuIL-1 beta were not effective in extending replication in pressure ulcer fibroblasts, in vitro. Tissue culture measurements from pressure ulcer biopsies demonstrated that, after 29 days of a high level of rhuIL-1 beta treatment (1.0 micrograms/cm2/day), the cytokine was effective in extending the ability of pressure ulcer fibroblasts to replicate. Tissue culture and electron microscopy suggested that, although rhuIL-1 beta promoted increases in fibroblast numbers, the primary effect appeared to be development of the extracellular matrix. The possible direct and indirect influences of rhuIL-1 beta therapy on pressure ulcers are discussed.     

Human nonspecific suppressive lymphokines

Since the term lymphokine first appeared in print over 20 years ago, a tremendous number of these soluble mediators of the immune system have been described. Within the past few years, many human nonspecific suppressive lymphokines have been identified. This review discusses the historical basis of immunologic suppression and suppressor factors. Later reports describing suppressive human lymphokines are then grouped into four categories: primarily stimulatory lymphokines that also mediate certain suppressive activities, suppressive lymphokines produced during altered states of immunity, suppressive lymphokines produced by exogenously stimulated lymphocytes, and suppressive lymphokines produced by unstimulated lymphocytes. Recent work I have been involved in focusing on the human suppressive lymphokine soluble suppressor factor (SSF) is also discussed.     

Immunomodulators for asthma

New information regarding the molecular mechanisms of allergic disorders has led to a variety of novel therapeutic approaches. This article briefly reviews the pathogenesis of asthma and allergic diseases, discusses the rationale behind using immunomodulators in these diseases; and examines the therapeutic effects of immunomodulators on allergic diseases. There are a number of immunomodulators that have been developed for the treatment of allergic disorders. Some have looked very promising in pre-clinical trials, but have not shown significant benefits in human clinical trials thus indicating the disparity between mouse models and human asthma. This review focuses on immunomodulators that are in human clinical trials and not molecules in pre-clinical development.     

Lymphocyte-neutrophil interactions: opposite effects of interleukin-2 and tumour necrosis factor-beta (lymphotoxin) on human neutrophil adherence

Human neutrophil adherence was enhanced by recombinant human tumour necrosis factor-beta (TNF beta) but suppressed by recombinant human interleukin-2 (IL-2). The opposite effects of these two lymphokines were observed over a range of concentrations consistent with their other biological activities, occurred within 15 min of incubation, and were still evident after 60 min. Pretreatment of neutrophils with both IL-2 and TNF beta resulted in adherence values intermediate between the values obtained with the individual lymphokines. IL-2 suppressed the stimulatory effects of both the chemotactic peptide formyl-methionyl-leucyl-phenyl-alanine (FMLP) and the phorbol ester phorbol myristate acetate (PMA). The combination of TNF beta with either FMLP or PMA produced enhancement of neutrophil adherence which exceeded that of either agent alone. These effects of the lymphokines were not due to endotoxin contamination since their effects were sensitive to heating and insensitive to polymyxin B treatment. These experiments provide further evidence for the critical role of these lymphokines in the regulation of acute and chronic inflammatory processes.     

The production of lymphokines by primary alloreactive T-cell clones: a co-ordinate analysis of 233 clones in seven lymphokine assays.

A total of 233 primary alloreactive T-cell clones have been tested for the production of interleukin-2 (IL-2), interleukin-3 (IL-3), immune(gamma) interferon (IFN) and granulocyte-macrophage colony-stimulating factor (CSF-2), B-cell growth factor I and II (BCGFI, BCGFII), and eosinophil differentiation factor (EDF). EDF was assayed by means of the eosinophil differentiation assay (EDA). Two principal correlations were observed: IL-3 was shown to be the major lymphokine detected in the bone marrow proliferation assay (BMPA) used to detect CSF-2, and there was a high correlation between the EDA and BCGFII. Subsequent work has suggested that this latter correlation is because a single factor is responsible for both activities. Apart from these two exceptions, and low level correlations probably due to the fact that different assays detect more than one lymphokine, there was no evidence for co-ordinate expression of lymphokines. There was a large variation in amounts of individual lymphokines produced. More clones produced multiple lymphokines than would be expected from independent control. Taken together, this pattern of regulation is consistent with the hypothesis that antigen stimulation of T cells results in the activation of all the lymphokine genes, but the amount of each produced is determined by secondary controlling mechanisms.     

Present and future of immune checkpoint blockade: Monotherapy to adjuvant approaches

Immune regulation of aggressive tumor growth is often outpaced by tumor up-regulation of ligands that inhibit effector immune responses through the activation of immune checkpoints. A few of such checkpoints include programmed death-1 (PD-1), cytotoxic T lymphocyte associated antigen-4 (CTLA-4), lymphocyte activation gene-3, T-cell immunoglobulin and mucin protein-3, Glucocorticoid-induced TNFR family-related receptor (GITR), and killer cell immunoglobulin like receptor. With the exception of GITR, after binding to their respective ligands these checkpoints induce down-modulation of immune responses to prevent autoimmunity. However, such immune mechanisms are co-opted by tumors to allow rapid tumor cell proliferation. Pre-clinical studies in antibody blockade of PD-1 and CTLA-4 have led to promising augmentation of effector immune responses in murine tumor models, and human antibodies against PD-1 and CTLA-4 alone or in combination have demonstrated tumor regression in clinical trials. The development of immune checkpoint blockade as a potential future immunotherapy has led to increasing interest in combining treatment modalities. Combination checkpoint blockade with chemotherapy and radiation therapy has shown synergistic effects in pre-clinical and clinical studies, and combination checkpoint blockade with bacterial vaccine vectors have produced increased effector immune responses in pre-clinical models. The future of immune checkpoint blockade may be as a powerful adjuvant alongside the current standard of care.     

自身免疫性疾病的基因治疗研究进展

自身免疫性疾病是危害人类健康的重要疾病。随着基因转移技术的发展,基因治疗在自身免疫性疾病中的研究取得了一定进展。通过选择合适的载体转导免疫调节分子、缺陷基因、激活因子、细胞凋亡受体以及诱导免疫耐受等方式来达到治疗目的。与传统的治疗方法相比,基因治疗具有高效性、靶向性、副作用小等优点。近几年来自身免疫性疾病的基因治疗在临床前实验和临床实验阶段取得了很多成果。     

自身免疫性疾病的基因治疗研究进展

自身免疫性疾病是危害人类健康的重要疾病.随着基因转移技术的发展,基因治疗在自身免疫性疾病中的研究取得了一定进展.通过选择合适的载体转导免疫调节分子、缺陷基因、激活因子、细胞凋亡受体以及诱导免疫耐受等方式来达到治疗目的 .与传统的治疗方法相比,基因治疗具有高效性、靶向性、副作用小等优点.近几年来自身免疫性疾病的基因治疗在临床前实验和临床实验阶段取得了很多成果.     

Synthetic bacterial lipopeptide analogs: structural requirements for adjuvanticity

Modern vaccines aim at conferring immune protection, independently of the nature of the etiological agent causing the disease. These new therapeutics are based on highly purified antigenic moieties that offer potential advantages over traditional vaccines, including a high degree of safety and the capacity of eliciting highly specific immune responses. In spite of these advantages however, subunit vaccines tend to be poorly immunogenic in vivo, and require the coadministration of adjuvants that indirectly enhance cellular immunity. Thus, recombinant vaccines development is dependent on the design of new molecules, non-immunogenic per se, but endowed with immune modulatory properties. Synthetic analogs of bacterial lipoproteins were described more than a decade ago, but their capacity to act as adjuvants has been only recently dissected. These low molecular weight non-immunogenic molecules can be reproducibly synthetized, are safe, and of easy handling and administration. Furthermore, new experimental data from our laboratory reveal their powerful adjuvant effect on human HLA-I/II restricted T cell responses and identify the molecular and cellular requirements for optimal adjuvanticity.     

Regulation of IgA B-cell development in the mucosal immune system

The factors regulating the differentiation of IgA B cells have been of great interest to mucosal immunologists as well as those generally interested in B-cell differentiation. It is now clear that such differentiation involves two major steps: first, isotype switch differentiation of surface IgM-bearing B cells into surface IgA-bearing B cells and, second, terminal differentiation of IgA B cells into IgA-producing plasma cells. Both of these steps are regulated processes that are under the influence of various cytokines and lymphokines. This paper presents data that define the role of cytokines and lymphokines in the regulation of IgA B-cell differentiation. A model of IgA B-cell differentiation is described in which the first step involves activation of the C alpha gene, while the latter is in germline configuration and thus the induction of surface IgM-bearing B cells partially committed to IgA expression. This occurs in Peyer's patches as a result of as yet incompletely defined signals from patch “switch cells.” The second step consists of conversion of the partially committed B cells to fully IgA-committed B cells and thus the completion of isotype switch differentiation. This step may be under the control of interleukin-4 (IL-4). The last step of the model involves the activation of IgA B cells (by antigen or mitogen) followed by the appearance on the cell surface of receptors which allow the cell to interact with cytokines or lymphokines (particularly IL-5). Such interaction results in cells capable of secreting IgA. Evidence is presented that an important adjuvant for mucosal immune responses, cholera toxin, acts to augment IgA production by promoting IgA B-cell differentiation at the isotype switch step.     

Inducible lymphokines of T cell tumors

T lymphocytes comprise a major class of lymphocytes and are themselves functionally heterogeneous. Some T lymphocyte functions are mediated by soluble products called lymphokines. Different lymphokines promote the activation, growth and differentiation of T and B lymphocytes, macrophages, and hemopoietic cells. Lymphokine production is associated with, but not limited to, helper T cells, and usually follows antigenic or mitogenic stimulation. The recognition that some lymphokines are produced after stimulation of neoplastic T cells has proved advantageous in the study of these molecules. T cell tumors are monoclonal, grow easily in vitro, and may produce fewer lymphokines than normal T cells. Thus, the purification and biochemical and biological characterization of some lymphokines have been facilitated by the availability of these tumors. More recently, T cell tumors have been used for evaluating the molecular structure of lymphokine-encoding genes. They have also provided information relevant to our understanding of the nature of T cell neoplasia.     

Hyper-IgM immunodeficiency syndrome: Influence of lymphokines onin vitro maturation of peripheral B cells

Peripheral B cells from six patients affected with the hyper-IgM immunodeficiency syndrome, characterized by an absence of IgG and IgA in serum with a concomitant elevated level of IgM, were analyzed for phenotypic and functional characteristics. We report that although the membrane antigenic pattern expression was characteristic of mature B cells, B cells from most patients exhibited an impairment in theirin vitro response to several lymphokines, such as recombinant interleukin 2 (rIL-2) and low molecular weight B-cell growth factor (BCGF), that induce proliferation of anti--activated B cells. This impairment was also found in response to a lymphokine mixture from a CD2-activated T-cell clone. The decrease in lymphokine-induced B-cell proliferation was accompanied by a low B-cell differentiation, whether patients' B cells were stimulated by the T-cell clone supernatant or rIL-2 and rIL6, lymphokines able to support differentiation ofStaphylococcus aureus Cowan I (SAC)-activated B cells. In addition, none of the lymphokines tested were able to induce patients' B cells to switch from IgM-secreting cells to IgG- and IgA-secreting cells. We conclude that this syndrome is associated with a defect in lymphokine-dependent maturation of B lymphocytes, although the T- or the B-cell origin of the defect still cannot be determined.