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T B Strom

Publications and source records attributed to T B Strom.

At least 163 records · Page 9Linked to original sources
PubMed1990

Interleukin 2 receptor-targeted cytotoxicity. Receptor binding requirements for entry of a diphtheria toxin-related interleukin 2 fusion protein into cells.

The receptor binding requirements for entry of the NAD+ ADP-ribosyltransferase component of DAB486-IL 2 into target cells were examined. Experiments utilizing cell lines bearing either high-affinity or individual subunits of the interleukin 2 receptor (IL 2R) as well as human peripheral blood mononuclear cells with natural killer activity demonstrate that the high-affinity receptor facilitates delivery of fragment A from DAB486-IL 2 to the cytosol approximately 1000 times more efficiently than either the intermediate-(p75) or low-affinity (p55) forms of the IL 2R. We show that elongation factor 2 (EF-2) in these cells is not quantitatively or qualitatively altered indicating that the relative resistance to intoxication displayed by IL 2R variant cell lines cannot be attributed to an altered intracellular target of the hybrid toxin. We also demonstrate that an alteration in the binding of DAB486-IL 2 to the p75 subunit of the IL 2R may account for the selective cytotoxicity of DAB486-IL 2 for cells bearing the heterodimeric high-affinity IL 2R.

Amino Acid Sequence↗
PubMed1990

Physiologic signaling in normal human T-cells: mRNA phenotyping by northern blot analysis and reverse transcription-polymerase chain reaction.

Synergy between ionomycin and sn-1,2-dioctanoylglycerol (diC8) was shown at the level of lymphokine gene transcription. Transcriptional activation of interleukin-2 (IL-2), interferon-gamma (IFN-gamma), and the protooncogene H-ras was accomplished by signaling highly purified normal human resting T-lymphocytes (T-cells) with diC8, a physiologic regulator of protein kinase C, and the calcium ionophore, ionomycin. Northern blot analysis of mRNA for early T-cell activation genes demonstrated the synergism between diC8 and ionomycin at the gene induction level. To amplify very low levels of IL-2 mRNA, sequential reverse transcription and polymerase chain reaction (RT-PCR) of T cell mRNA were used to demonstrate the capacity of the calcium signal (ionomycin) to promote low-level IL-2 transcription in normal human T-cells without additional signals. Cyclosporine (CsA) prevented diC8 and ionomycin-induced expression of IL-2, IFN-gamma, and H-ras genes. The completeness of its inhibitory effect was evident by the absence of IL-2 transcripts in CsA-treated cultures screened by the RT-PCR technique. CsA also prevented IL-2 and IFN-gamma gene expression in accessory cell-depleted T-cells stimulated by cross-linking the CD2 and CD3 antigens on the cell surface. Our observations demonstrate that a physiologic regulator of PKC, diC8, and cell surface crosslinking of the CD2 and CD3 antigen, promote gene expression in normal human quiescent T-cells independently of accessory cells, and that CsA prevents gene expression via a direct effect on T-cells.

Antigens, Differentiation, T-Lymphocyte↗
PubMed1990

The cascading, interrelated roles of interleukin-1, interleukin-2, and interleukin-6 in murine anti-CD3-driven T cell proliferation.

T cell stimulation occurs following interaction of T cell receptor (TcR) with processed antigen presented by autologous accessory cells (AC). The effects of antigen stimulation on T cells are mimicked by monoclonal antibodies (Mabs) defining proteins of the TcR-CD3 complex. In this study, we examine the roles of T cell density, AC density, divalent and polyvalent forms of anti-CD3 Mab, and the cytokines interleukin (IL)-1, IL-2, and IL-6 in T cell activation and proliferation. Stringently AC-depleted T cells do not proliferate in response to Con A or divalent anti-CD3; however, polyvalent anti-CD3 provides a powerful signal for isolated resting T cell proliferation. Recombinant (r)IL-2 strongly amplifies T cell proliferation in response to anti-CD3, whereas rIL-1 exerts no direct effects on anti-CD3-stimulated T cells. In the presence of AC, however, rIL-1 augments T cell proliferation to anti-CD3. Recombinant IL-6 promotes T cell proliferation among T cells stimulated with polyvalent but not divalent anti-CD3. As deduced by Northern blot analysis, rIL-1 increases cytoplasmic levels of IL-6 mRNA in AC. Recombinant IL-6, in turn, amplifies the accumulation of stable IL-2 transcripts in purified T cells stimulated with polyvalent anti-CD3. Hence, IL-1, IL-6, and IL-2 support T cell proliferation through cascading effects at the level of gene transcription. The cytokines evaluated in this study, however, do not fully reconstitute AC functions in promoting anti-CD3 Mab T cell proliferation.

Animals↗
PubMed1990

Evidence that glucocorticosteroids block expression of the human interleukin-6 gene by accessory cells.

The mode of action of glucocorticosteroids as immunosuppressive and antiinflammatory agents is not fully understood. Glucocorticosteroids block synthesis of interleukin 1 by interfering with the transcription of the IL-1 beta gene. Glucocorticosteroids may also induce rapid degradation of IL-1 mRNA. In the presence of antigen, IL-1 is a potent accessory-cell-derived growth and differentiation co-factor for stimulating resting T lymphocytes. The recently defined interleukin 6 protein is even more powerful than IL-1 in promoting T cell growth and differentiation and acts synergistically with IL-1. Like IL-1, IL-6 is produced by accessory cells and exhibits pleiotropic functions. We herein describe the effects of glucocorticosteroids on IL-6 synthesis. We provide evidence that glucocorticosteroids prevent IL-6 gene transcription in human peripheral blood mononuclear cells.

Antigen-Presenting Cells↗
PubMed1990

Possible association of the immunosuppressive and B cell lymphoma-promoting properties of cyclosporine.

Central to the immunosuppressive properties of cyclosporine is a drug imposed blockade of the interleukin-2 gene activation. As IL-6 stimulates antigen-activated T cells to release IL-2, we examined the influence of CsA on IL-6 gene expression and IL-6-supported T cell proliferation. Northern blot analysis revealed that CsA failed to abolish IL-6 gene expression in mitogen-activated peripheral blood mononuclear cells. In fact, increased IL-6 gene transcription and increased release of IL-6 bioactivity were detected using mitogen-activated PBMCs cultured with CsA doses (200-800 ng/ml) only slightly in excess of the minimal antiproliferative dose. CsA completely abrogated the IL-6-stimulated proliferative responses of macrophage-depleted T cells stimulated with polyvalent anti-CD3 monoclonal antibodies. It is interesting that CsA-treated patients evidence an increased incidence of polyclonal lymphoproliferative disorders and B cell lymphomas. As IL-6 fosters B cell activation and growth of EBV-transformed B cells, excessive CsA doses may support development of EBV-transformed B cell lymphomas via superinduction of the IL-6 gene.

Antigens, Differentiation, T-Lymphocyte↗
PubMed1990

A kinetic analysis of the effects of interleukin-2 diphtheria toxin fusion protein upon activated T cells.

The interleukin-2 diphtheria toxin-related fusion protein (IL-2 toxin) inhibits protein and DNA synthesis IN rIL-2 (10(-10) M) stimulated T lymphoblasts in a dose-dependent fashion. However, prior to target cell death very low concentrations of rIL-2 and IL-2 toxin synergistically stimulate [3H] thymidine incorporation despite inhibition of [14C] leucine uptake. A sequential analysis of [3H] thymidine incorporation shows that high IL-2 toxin concentration (10(-9)-10(-7) M) stimulates DNA synthesis at 18 hr of culture and inhibits [3H] thymidine uptake after 24 hr, while low concentrations of IL-2 toxin (10(-12)-10(-10) M) exhibits stimulatory effects only after 24 hr of culture. Anti-Tac a monoclonal antibody directed against the p55 chain of the high affinity IL-2 receptor (IL-2R) blocks the stimulatory effects of high-dose IL-2 toxin, thereby proving that these effects are mediated through the IL-2 domain of the fusion protein. At 7 hr following interaction with IL-2R receptor (IL-2R)+ T cells, IL-2 toxin-treated cells evidence augmented transcription of the heat shock protein gene, an effect indistinguishable from those mediated by rIL-2. We conclude that interaction of IL-2 toxin with IL-2R+ T cells initially mimicks the stimulatory effects of IL-2 upon gene transcription and DNA synthesis yet concomitant inhibition of protein synthesis is evident.

Cells, Cultured↗
PubMed1990

Dose-related mechanisms of immunosuppression mediated by murine anti-CD3 monoclonal antibody in pancreatic islet cell transplantation and delayed-type hypersensitivity.

Although anti-CD3 mAb therapy is used extensively in clinical transplantation, the dose-related effects and mechanisms of action are not clearly defined. We have examined the dose-related effects of an antimurine CD3 mAb, 145-2C11, in pancreatic islet cell allograft and the delayed type hypersensitivity reaction models of T-cell-dependent immunity. Low-dose anti-CD3 therapy (0.5 micrograms/day) administered over several days mediated superficially equal, effective clinical immunosuppression as a single high-dose intravenous injection (400 micrograms). T cells harvested from animals treated with high-dose anti-CD3 were unresponsive to in vitro restimulation. In contrast, T cells isolated from low-dose treated animals retained in vitro proliferative capacity when restimulated with polyvalent anti-CD3 mAb. The terminal complement components were not required to support in vivo immunosuppression mediated by anti-CD3 mAb as C5 deficient mice were immunosuppressed by the administration of this mAb. In some pancreatic islet cell allograft recipients, permanent engraftment, but not tolerance, was achieved. Replacement of donor leukocytes produced acute rejection in hosts bearing long-term, well-accepted grafts. Prolonged anti-CD3 mAb treatment may provide sufficient time for replacement or inactivation of donor leukocytes.

Animals↗
PubMed1990

Immunotoxins and cytokine toxin fusion proteins.

Paul Ehrlich first suggested the simple and elegant concept of creating specific cell toxins or 'magic bullets' through the fusion of cell specific antibodies and toxins. In practice it has proven difficult to create safe and effective 'magic bullets'. In the past several years, several immunotoxins have been applied to clinical testing. These immunotoxins have been created by the biochemical coupling of cell or lineage specific monoclonal antibodies to plant toxins or fragments thereof. These immunotoxins have been used to treat bone marrow transplant recipients and patients with autoimmune disorders. In recent years, another strategy has also been pursued to create hybrid toxins. Rather than use antibodies as the targeting moiety, cytokines have been used to target a select population of cells bearing a high copy number of receptors for the specific cytokine. Rather than biochemically couple a cytokine to the toxin, the cytokine and toxin are fused by a peptide bond established via genetic engineering. A prototype IL-2 diphtheria toxin-related fusion protein is now being tested in the clinic for treatment of hematopoietic malignancies and autoimmune disorders.

Animals↗
PubMed1989

Cytotoxicity of interleukin 2-toxin toward lymphocytes from patients with adult T-cell leukemia.

The inhibitory effect of a diphtheria toxin-related interleukin 2 fusion protein, IL-2-toxin, on protein synthesis in adult T-cell leukemia/lymphoma (ATL) cells was examined in vitro. Peripheral blood ATL cells from 12 patients (six acute type, four chronic type, and two smoldering type ATL) and the lymph node cells from three ATL patients (two acute type and one lymphoma type ATL) were examined. At a concentration of 10(-8) M, IL-2-toxin inhibited protein synthesis by 60 to 98% in lymph node ATL cells, whereas protein synthesis in peripheral blood ATL cells was inhibited from 20 to 57% in acute type, and from 3 to 13% in chronic type. In contrast, IL-2-toxin had no measurable effect on T-cells from either patients with smoldering type ATL or normal controls. The cytopathic effects of IL-2-toxin were blocked by the addition of anti-CD25 monoclonal antibody, suggesting that the inhibition of protein synthesis in target cells was mediated by the IL-2 receptor (IL-2R). The degree of inhibition of protein synthesis, however, was not closely correlated with expression of CD25 antigen (low-affinity Mr 55,000 glycoprotein, IL-2R, Tac antigen) on ATL cells. There was an apparent correlation between the degree of inhibition and the rate of protein synthesis in ATL cells. We demonstrate that ATL cells from patients with acute or lymphoma type disease were more sensitive to IL-2-toxin than cells from chronic or smoldering disease. These findings suggest that the high affinity IL-2R present on acute and lymphoma type ATL cells may serve as a target for therapy with this recombinant chimeric toxin.

Adult↗
PubMed1989

Antibody valence and induced signal transduction: the role of antibody valence in anti-CD3-induced signal transduction in isolated normal T cells.

This report provides direct evidence that protein kinase C (PKC) is activated in isolated, rigorously accessory cell (AC)-depleted T cells when the T cell antigen recognition complex is stimulated by divalent anti-CD3 monoclonal antibody. Anti-CD3 monoclonal antibody-stimulated PKC activation alone does not, however, directly stimulate T cell proliferation in the absence of AC. A rise in cytosolic calcium is the second signal believed to be of paramount importance in T cell activation. While mitogenic concentrations of some divalent anti-CD3 antibodies do not cause a rise in cytosolic calcium, polyvalent anti-CD3 does evoke increased intracellular Ca2+ in rigorously AC-depleted resting human T cells.

Antibodies, Monoclonal↗
PubMed1989

Toward more selective therapies to block undesired immune responses.

Our results provide important evidence that IL-2 receptor bearing cells are required for undesired immune reactions involved in autoimmunity, allograft rejection and nephritogenic processes. Administration of anti-IL-2 receptor monoclonal antibodies prolonged vascularized heart allograft survival across major histocompatibility barrier in mice and rats and renal monkey grafts. Indeed, several grafts survived indefinitely, although the antibody was administered only for the first 10 days post-transplantation. Rejection of the remaining grafts may well reflect inadequate dosage of antibody; dose-response studies have not been performed to date. In addition to preventing rejection, delayed treatment with anti-IL-2R monoclonal antibody was shown to reverse ongoing rejection in other recipients of heart allografts. Such long-term engraftment following cessation of therapy makes it unlikely that anti-IL-2R treatment prolongs graft survival by pharmacologic blockade of the IL-2R. Furthermore, exogenous IL-2 does not diminish the beneficial effects of anti-IL-2R antibody therapy in rodents. Whether or not such prolonged graft survival represents deletion of the responding T cell clones is a subject of current investigation. Results in a delayed-type hypersensitivity model indicate that complement fixation, is required to achieve optimal immunosuppression. Moreover, only anti-receptor antibodies that block IL-2 binding mediate optimal immunosuppression. Passive transfer experiments clearly prove that immediate post-transplant courses of anti-IL-2R monoclonal antibody spares suppressor T cells. In rodent models, delayed type hypersensitivity and lupus and diabetic autoimmunity are prevented by anti-IL-2R treatment. Finally, the availability of monoclonal antibodies directed against the human IL-2R provides an opportunity to extend this principle to clinical transplantation.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1989

Current stage of interleukin 2 receptor targeted therapy.

The overview summarizes a new and selective form of immunosuppressive therapy by using anti-Interleukin-2 receptor monoclonal antibodies, which may soon become an important form of recipient treatment in organ/tissue transplantation or/and of patient treatment with auto-immune diseases.

Animals↗
PubMed1989

Sequential effects of interleukin 2-diphtheria toxin fusion protein on T-cell activation.

The interleukin 2-diphtheria toxin-related fusion protein (IL-2-toxin) rapidly inhibits protein synthesis in IL-2 receptor (IL-2R)-bearing phytohemagglutinin-activated T cells but transiently stimulates DNA synthesis. At 7 hr after interaction with IL-2R+ phytohemagglutinin-activated T cells, IL-2-toxin-treated cells bear augmented steady-state levels of c-myc, interferon gamma, and IL-2R mRNA; these effects are indistinguishable from those produced by recombinant IL-2. Amplification of IL-2 sequences by the polymerase chain reaction reveals an increased level of IL-2 mRNA in cell cultures treated with recombinant IL-2, IL-2-toxin, and cycloheximide. These results suggest that IL-2-toxin can affect de novo IL-2 gene transcription/mRNA stabilization through independent mechanisms exerted by both the IL-2R binding domain and ADP-ribosyltransferase activity of the fusion protein. After 20 hr of culture, IL-2R mRNA was markedly decreased in both IL-2-toxin- and cycloheximide-treated phytohemagglutinin-activated T cells. Although interaction of IL-2-toxin with IL-2R+ T cells initially mimics the stimulatory effects of IL-2 upon c-myc, interferon gamma, IL-2R, and IL-2 gene expression, the consequences of inhibition of protein synthesis mediated by the ADP-ribosyltransferase activity of the toxin dominate after 7 hr and are indistinguishable from those effects mediated by cycloheximide.

Base Sequence↗
PubMed1989

Prolongation of primate renal allograft survival by anti-Tac, an anti-human IL-2 receptor monoclonal antibody.

In an effort to produce specific immunosuppression through the targeting of those lymphocytes expressing cell surface interleukin 2 receptors in response to an allograft, the anti-human IL-2 receptor monoclonal antibody anti-Tac was administered to cynomolgus monkeys receiving renal transplants. The data demonstrate that anti-Tac produces a significant delay in renal allograft rejection and prolongs host survival in cynomolgus monkeys. Though higher doses of anti-Tac produce modest delays in rejection, there was a surprising finding of greatly prolonged survival in three of five monkeys treated with much lower doses of anti-Tac. Anti-Tac was not shown to be synergistic with cyclosporine in this model. Animals treated with anti-Tac developed high titers of antibodies against the murine monoclonal antibody after 6-8 days of treatment, associated with the disappearance of plasma anti-Tac staining of activated lymphocytes as measured by flow cytometry. The data confirm the utility of the IL-2 receptor as a target for immunosuppressive therapy, and suggest that investigations of dosage and of methods to reduce the immunogenicity of anti-IL-2 receptor agents may be beneficial.

Animals↗
PubMed1989

Interleukin-2-diphtheria toxin fusion protein prolongs murine islet cell engraftment.

Allograft rejection is dependent upon complex cell-mediated processes, the primary effectors of which are activated T cells. As the expression of the cell surface protein interleukin 2 receptor is primarily limited to the subset of stimulated T cells, therapeutic agents that target this molecule may provide highly selective immunosuppression. A newly constructed chimeric IL-2 diphtheria toxin fusion protein specifically binds to and poisons activated T cells bearing the high-affinity IL-2R. We describe the in vivo effects of IL-2 toxin in preventing rejection of a crude pancreatic islet preparation transplanted across major and minor histoincompatibility barriers. IL-2 toxin administered once daily as the sole immunosuppressive agent prolongs islet graft survival and decreases the severity of the early mononuclear cell infiltrate into the graft site. Long-term survival of transplanted islets (greater than 100 days) was achieved following a short course (10 days) of more-intensive IL-2 toxin treatment. Thus IL-2 toxin, a highly selective immunosuppressive agent, leads to prolonged islet cell engraftment while sparing the resting or memory subset of the entire T cell repertoire.

Animals↗
PubMed1989

Prolongation of cardiac allograft survival in murine recipients treated with a diphtheria toxin-related interleukin-2 fusion protein.

A diphtheria toxin-related IL-2 fusion gene has been constructed that encodes a 68KD recombinant toxin in which the diphtheria toxin receptor-binding domain has been replaced with amino acids 2-133 of IL-2. This chimeric IL-2 toxin is cytotoxic for cells expressing the high-affinity IL-2 receptor but not for cells lacking this receptor. The ability of this IL-2 toxin to prolong allograft survival was examined in a murine vascularized, heterotopic heart transplant model in the strain combination B10.BR into C57B1/10. When given at a dose of 1.0 micrograms/day for 10 days, the IL-2 toxin significantly prolonged allograft survival in all recipients. CRM-45, a fragment of diphtheria toxin missing the binding domain, was ineffective, confirming the specificity of the therapy. The results demonstrate that this IL-2 toxin, which targets activated T cells expressing the IL-2 receptor, will prolong allograft survival, offering a new option for immunosuppressive therapy.

Animals↗
PubMed1989

Similar effects of cyclosporine and verapamil on lymphokine, interleukin 2 receptor, and proto-oncogene expression.

Since calcium channel blocking agents and CsA exert an antiproliferative effect upon T cell mitogenesis, we have compared and characterized their immunosuppressive properties at the level of gene activation. Verapamil (greater than or equal to 30 microM), which blocks T cell mitogenesis and a rise in cytosolic calcium, was added to cultures of peripheral blood mononuclear cells stimulated with phytohemagglutinin (5 micrograms/ml) and phorbol myristate acetate (5 ng/ml). Northern blot analysis was performed using cDNA probes for the p55 interleukin 2 receptor (Tac; IL-2R), interleukin 2 and c-myc at 20 hr of culture. Accumulation of IL-2 encoding mRNA within the cytoplasm was completely abrogated by verapamil. However, IL-2R and c-myc encoding mRNA were clearly detectable in verapamil-treated cell cultures. Surface expression of the Tac protein in mitogen-activated T cells was also not blocked by verapamil as shown by FACS analysis. In companion experiments with CsA, verapamil only partially inhibits the intracellular processes leading to T cell activation. A calcium-independent pathway may exist for the expression of IL-2R and c-myc, while an increase of intracellular Ca2+ may provide the additional signal for IL-2 gene expression. Although the in vitro concentrations of verapamil used in these experiments are in excess of common clinically therapeutic levels, the results help clarify the mode of CsA action and may provide a new tool to dissect the early events of T cell activation.

Calcium↗