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F R Seiler

Publications and source records attributed to F R Seiler.

At least 55 records · Page 3Linked to original sources
PubMed1989

Granulocyte-macrophage colony-stimulating factor binding sites and oxidative metabolism in human granulocytes.

We investigated the interaction between GM-CSF and its receptor on human granulocytes and on several human tumor cell lines. Specific high-affinity binding for GM-CSF was characterized by Scatchard plot analysis. The specific radioactivity of the 125I-labeled derivative of rH. GM-CSF was determined by self-displacement analysis and calculated to be 30 microCi/micrograms. The maximum concentration of binding sites (B max) in granulocytes was 40 fmol/mg protein (2,200 molecules GM-CSF bound/cell) and the dissociation constant (KD) was 0.42 nM. No binding sites for GM-CSF were found in two lung cancer cell lines, SCLC-16HV and NCI-N417 or in the urinary bladder carcinoma cell line 5637, whereas the promyelocytic leukemia cell line HL60 was positive for GM-CSF binding. Time course experiments showed maximum binding of GM-CSF in granulocytes after an incubation period of 60 min and a decrease in binding after an incubation period of 2 h. In parallel, we found a maximum biological signal when granulocytes were preincubated for 90 min with GM-CSF, and a decrease after an incubation time of 120 min. Preincubation of the cells with rH. GM-CSF induced an enhancement of the production of activated oxygen species by the cells in response to PMA.

Binding Sites↗
PubMed1988

C4-binding protein prevents spontaneous cleavage of C3 in sera of patients with hereditary angioedema.

We have studied the effects of polyclonal monospecific Fab' preparations against C1r, C1s, C1INH, C4, C4bp, and fragment Bb of factor B on complement activation in NHS and HAES. Furthermore, we have investigated complement activation in these sera after addition of purified C1s and purified C4bp. Blocking C1INH induced a spontaneous activation of the classical pathway in NHS and to a lesser extent in HAES. Addition of p-C1s resulted in a strong C3 conversion in NHS, but not in HAES. However, after the blocking of C4bp in HAES, addition of p-C1s produced a total C3 consumption. The ration of the protein concentration of C4bp to hemolytically active C4 was eight times higher in HAES than in NHS. This increased ratio may account for the resistance of HAES to the C1s induced C3 cleavage in our in vitro system and the stability of C3 in HAE despite C4 and C2 consumption in vivo.

Angioedema↗
PubMed1988

Rationals for the development of the human hematopoietic colony stimulating factors as therapeutical useful drugs.

Proliferation and differentiation of stem cells within the bone marrow into different cell types circulating in the peripheral blood is regulated by a series of hierarchically acting growth mediators--the colony-stimulating factors. In recent years, the genes of some of these physiological glycoproteins were cloned by means of molecular biology, and thus highly purified recombinant protein can now be produced in large scale. This has provided access to profound investigation of the role of these factors and the mechanisms for their action with regard to their potential therapeutical use. Part of their properties as revealed by the various in vitro test systems and by corresponding efficacy models using laboratory animals could also be found in recently performed phase I clinical studies, i.e. for granulocyte-macrophage colony-stimulating factor (GM-CSF) and granulocyte colony-stimulating factor (G-CSF). Perspectives for a potential use of these regulators as single therapy and in possible additive or synergistic combinations are currently intensively studied.

Colony-Stimulating Factors↗
PubMed1988

E. coli derived human granulocyte-macrophage colony-stimulating factor (rh GM-CSF) available for clinical trials.

Recombinant human GM-CSF has been expressed as a fusion protein in E. coli in the form of inclusion bodies. Using denaturing agents, acid cleavage and sulfitolysis, the biologically inactive GM-CSF protein could be highly purified and additionally renaturated under suitable reoxidizing conditions. The thorough repair of the two disulfide bridges could be confirmed by sequencing fragments obtained by tryptic digestion. Refolding of the molecule has been studied by CD spectrometry and identity by Western blotting and SDS-PAGE analysis. As could be demonstrated, full biological activity (colony-forming assay with fresh human bone marrow cells) was restored during renaturation of the GM-CSF protein. Further proof of biological equivalence of the E. coli-derived protein with a yeast-derived biologically active rh GM-CSF has been published elsewhere.

Circular Dichroism↗
PubMed1988

Monoclonal antibodies directed to leukocyte differentiation antigens for therapeutic use.

During the past years monoclonal antibodies directed to human leukocyte differentiation antigens have resulted not only in fundamental new knowledge of function and differentiation of leukocyte-populations, but have also turned out to be excellent and new tools for the treatment of lymphoid malignancies. Most widely and successfully monoclonal antibodies directed to T-cell differentiation antigens have been used for immunosuppression in organ transplantation. In the present paper, experimental and clinical data are analyzed which may give information about the criteria for clinical effectivity of distinct monoclonal antibodies. The conclusions clearly show, that not only specificity of an antibody, e.g. recognition of a distinct antigen, but also the recognized epitope, affinity and isotype of an antibody play a crucial role for the biological effectivity of a monoclonal antibody. Clinical success in antibody-therapy will therefore be dependent on the possibilities to alter antibody molecules according to the special clinical situation.

Antibodies, Monoclonal↗
PubMed1988

Investigations with monoclonal antibody drug (anthracycline) conjugates.

The development of monoclonal antibody-drug (anthracycline) conjugates (immunocytostatics) that has emerged during the last decade is briefly reviewed. Stress is layed on the various procedures that have been employed for the chemical attachment of daunorubicin (daunomycin) and doxorubicin (adriamycin) to spacers, high-molecular weight carrier molecules, and immunoglobulins. Anthracycline conjugates that have proved effective in mice with respect to the treatment of cancer are especially evaluated. Also a new method developed for the conjugation of rhodosaminyl anthracyclinones via a hydrolysable spacer, developed in our laboratories, is briefly communicated. Finally, import aspects for further developments of monoclonal antibody-drug conjugates are discussed.

Antibiotics, Antineoplastic↗
PubMed1988

Preparation and use of synthetic blood group specific immunoadsorbents.

The carbohydrate structures determining the ABO blood group system have been almost completely characterized. Recent advances in the stereoselective synthesis of such complex structures have made it possible to prepare pure oligosaccharides in large quantities and to study their possible uses for diagnostics and pharmaceutical processing techniques. In the experiments described here A and B blood group specific determinants were synthesized and bound to solid phases by means of suitable spacers in order to study their use as immunoadsorbents. The objective was to adsorb blood group specific antibodies from positive sera and from immunoglobulin preparations. It was shown that the anti-A and anti-B antibodies bound to the immunoadsorbents with high affinity could be removed effectively. The effects were achieved both using affinity chromatography and "batch processing".

ABO Blood-Group System↗
PubMed1988

Human recombinant derived IL-3 and GM-CSF in hematopoiesis of normal cynomolgus monkeys.

Recent progress in molecular cloning has provided access to several major human colony-stimulating factors - GM-CSF, IL-3, G-CSF and M-CSF. Now they are available highly purified from different expression systems (e.g. yeast, E. coli, CHO cells). These molecules, acting multifunctionally in the hematopoietic system, are responsible for proliferation and differentiation of bone marrow-derived progenitor cells in vitro. First clinical studies performed with colony-stimulating factors have shown that neutropenia caused by drug-induced immunosuppression in patients with refractory cancer or autologous bone marrow transplantation was reversed using rh GM-CSF or rh G-CSF. We have investigated the effect of the consecutive administration of rh IL-3 and GM-CSF on hematopoiesis in normal cynomolgus monkeys. Whereas administration of rh IL-3 alone did not result in an increase of WBC counts, the combination therapy of rh IL-3 followed by rh GM-CSF exhibited significant synergistic effects and raised WBC numbers. Furthermore, application of both factors resulted in a platelet rise not seen when one of the factors was used alone. The response was dose dependent and implicated that the therapeutical potential of rh GM-CSF could be expanded if used in combination with rh IL-3.

Animals↗
PubMed1988

Influence of recombinant human granulocyte-macrophage colony-stimulating factor on granulocyte functions.

Besides its effect on bone marrow progenitors, GM-CSF is able to interact with mature cells such as granulocytes which express GM-CSF receptors at their surface. Using an incubation period of 90 min, which corresponds to the maximum expression of GM-CSF receptors, we studied oxidative metabolism, random migration and chemotaxis as well as elastase release. Elastase was chosen as a marker for primary granules. We observed inhibition of chemotaxis and random migration, and stimulation of oxidative metabolism as well as elastase release in presence of rh GM-CSF. These different effects were dependent on the dosage of rh GM-CSF applied.

Chemotaxis, Leukocyte↗
PubMed1988

Comparison of relevant biological assays for the determination of biological active erythropoietin.

Human recombinant erythropoietin (rh EPO), a glycoprotein hormone which is an obligatory growth factor for the proliferation and differentiation of committed erythroid progenitor cells, has been purified to homogeneity and compared to human urinary erythropoietin (EPO). Erythropoietin levels will be determined and standardized by in vivo bioassays in which endogenous erythropoietin production has been reduced by hypertransfusion or hyperbaric atmosphere. The effect of EPO on the rate of red cell production or Fe59 incorporation is used as measure of its erythropoietin content. Both in vivo assay systems will be compared with respect to standardization for routine laboratory use. For in vitro characterization of EPO the classical assay for Fe59 incorporation in in vitro rat bone marrow cells will be compared to the proliferation and H3 Thd uptake by treatment with phenylhydrazin enriched erythroid progenitors from mouse spleens. (The direct physiological effect of EPO on stem cells will be shown by differences in BFUe progenitor levels). Rh EPO is jointly developed by Integrated Genetics (Boston) and Behringwerke AG (Marburg).

Animals↗
PubMed1988

Biological characterization of recombinant human erythropoietin.

Highly purified, recombinant, human erythropoietin (rh EPO) has been compared to natural urinary derived erythropoietin (nEPO). Both EPO preparations have been characterized biologically: The proliferation of murine spleen cells in vitro after pretreatment with phenylhydrazine and the 59Fe incorporation into the heme of polycythemic mice have been determined. Further, the effect of rh EPO on the erythropoiesis of normal mice was studied. Depending on the applied rh EPO dosage an increase in hematocrit was observed. Treatment with rh EPO in rats with anemia due to subtotal nephrectomy also showed a dose-dependent rise of hematocrit and hemoglobin, and therefore a reversal of the anemic status. The comparison of rh EPO and natural EPO showed full biological activity of the recombinant protein and its equivalence to the natural hormone.

Anemia↗
PubMed1988

Modulation of functions of granulocytes by recombinant human GM-CSF and possible complications of GM-CSF therapy.

Besides its effect on bone marrow progenitors, GM-CSF is able to modulate functions of mature cells such as neutrophils. It inhibits random migration and chemotaxis through action on both cells and chemotactic factors, and stimulates oxidative metabolism as well as elastase release. Furthermore, it strongly enhances the response of the cells to the usual stimulants such as f-Met-Leu-Phe and phorbol esters. The role of neutral proteinases and activated oxygen species in different diseases such as ARDS, emphysema, coagulation defects, arthritis, and inflammation, is recognized. The remarkable in vitro release of neutral proteinases and activated oxygen species from granulocytes after GM-CSF stimulation may be of importance in vivo. This should be considered in clinical application of GM-CSF, particularly with high-dose therapy.

Cell Movement↗
PubMed1988

On the nature of IgG dimers. I. Dimers in human polyclonal IgG preparations: kinetic studies.

Human polyclonal IgG for therapeutic or prophylactic purposes is usually prepared from pooled plasmas taken from more than 1000 donors. After storage in solution under physiological conditions for a sufficiently long period, a considerable percentage of dimers (approx. 10-30% [w/w] at a protein concentration of approx. 160 mg/ml) represents the main component of aggregates in contrast to the essentially monomeric IgGs of monoclonal or single donor origin. Analysing the kinetics of monomer-dimer equilibration suggests assuming approx. 10(6) different antibody (ab) populations interacting independently and simultaneously with a specific partner of the reaction. Concerning the average apparent (functional) equilibrium constant of association, Kapp., a distinction could be made between two main populations characterized by values in the range if approx. 2.5-3.0 X 10(10) M-1 and 1.0 X 10(12) M-1, respectively. The results were obtained by computer simulation, taking an association rate constant, k+1, of 5 X 10(5) M-1 s-1 as a basis. Since the main part of the individual populations was found to interact Fc-independently via Fab-located binding sites, we suppose that the dimers are for the most part complexes of idiotypic (Ids) and anti-idiotypic abs (anti-Ids). Moreover, dimerization seems to be mainly a bivalent binding reaction, at least at the experimental concentrations. The results are in line with the concept of an idiotypic network regulation in man.

Animals↗
PubMed1987

Stimulation of oxidative metabolism of granulocytes by recombinant granulocyte-macrophage-colony-stimulating-factor and a conditioned medium of a urinary bladder carcinoma cell line.

Neutrophils (PMN) are the major host defence cells protecting the body against invasion by microorganisms. Products of oxidative metabolism mediate PMN microbicidal and tumoricidal activity, but the mechanisms by which these pathways become activated are not well understood. The colony stimulating factors (CSF) are known to stimulate proliferation and differentiation of committed bone marrow stem cells. These regulators may probably play an important role in non specific resistance to infections. We studied the oxidative metabolism of neutrophils after stimulation with recombinant GM-CSF (r.GM-CSF) and the concentrated conditioned medium of the UBC-5637 cell line (UBC-CM) showing CSF activity. It could be demonstrated that the r.GM-CSF, as well as the UBC-CM, induce an activation of the neutrophil respiratory burst without any cofactors such as f-MLP, PMA, or zymosan. In addition, we observed an increase of the response to those stimulants in the presence of either r.GM-CSF or UBC-CM. These effects were not endotoxin-induced, since stimulation persisted after addition of Polymyxin B, which is known to inhibit the action of endotoxins.

Cell Line↗
PubMed1987

Fc fragments of human IgG may influence allergic reactions.

The antiallergic effect of Fc fragments prepared from human polyclonal IgG was investigated in several experimental models. In an in vitro assay, isolated ileum of cynomolgus monkeys was sensitized with serum from atopic patients. In six of fifteen monkeys the subsequent addition of specific allergens reproducibly resulted in an ileum contraction measured in the Schultz-Dale apparatus. In all six positive monkeys pre-treatment of the isolated ileum with Fc (papain) before sensitization inhibited ileum contraction. Fc (plasmin) or Fc (pepsin) was less or not effective, aggregated IgG, monomeric IgG, sulfonated IgG, the Fc-free F(ab')2 moiety, or albumin had no significant or no reproducible inhibitory effect. In an in vivo assay passive cutaneous anaphylaxis was studied in 28 cynomolgus monkeys. Different dilutions in PBS of the particular specific allergens were subsequently administered to sensitized skin sites, simultaneously to i.v. injection of Evans blue. The degree of the local allergic (passive cutaneous anaphylactia) reaction was evaluated by measuring the diameter of the blue area at the dermal injection sites. About 50% of sera induced positive reaction in monkeys. Compared to the control (application of PBS), the degree of the positive reactions could be reduced by about 50% if Fc (papain) (optimal dose 25 mg/kg body weight) was injected i.v. either 2 h before or after local sensitization. No significant inhibitory effect could be found with the Fab moiety of IgG. Follow-up studies showed that the inhibitory effect of Fc lasted for about 10 days. The pharmacological basis of the observed antiallergic action of Fc is not yet understood.

Animals↗