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T N Oeltmann

Publications and source records attributed to T N Oeltmann.

31 records · Page 2Linked to original sources
PubMed1985

Synthesis and in vitro activity of a hormone-diphtheria toxin fragment A hybrid.

The synthesis and in vitro biological activity of a hybrid protein composed of intact human chorionic gonadotropin and fragment A of diphtheria toxin in a disulfide conjugate is reported. This hybrid retained greater than 90% of the binding ability of uncoupled hCG and was shown to be specifically toxic to a mouse Leydig cell tumor which binds hCG while being non-toxic towards cells which lack receptors for hCG.

Animals↗
PubMed1983

Insulin-ricin B hybrid molecules mediate an insulin-associated effect on cells which do not bind insulin.

A hybrid molecule was constructed by covalently linking, by a disulfide bridge, the hormone insulin to the binding subunit B of the plant toxin ricin (specificity: Gal, GalNAc). Monolayer-cultured MDCK cells, which lack detectable levels of specific plasma membrane 125I-insulin binding but which readily bind 125I-insulin-ricin B, were used in these studies. Binding of insulin-ricin B to these cells could be displaced by lactose and ricin B, but not by insulin. The biological activity of the hybrid, as measured by [14C]glucose incorporation into glycogen, was stimulated in a dose-dependent manner by the hybrid (10(-11)-10(-8) M), whereas glycogen production was not stimulated by insulin alone. The stimulated glycogen labeling in response to the hybrid was also inhibited by lactose and ricin B. When ricin B alone was tested over the same range of concentrations, stimulation of glycogen synthesis was not observed, nor was there any evidence for stimulation when insulin and ricin B were added simultaneously. These data suggested that alternate cell surface receptors (i.e. ricin B receptors) may substitute for specific receptors (i.e. insulin receptors) to convey intracellular metabolic control signals in this cell line.

Animals↗
PubMed1983

Insulin-ricin B hybrid molecules: receptor binding and biological activity in a minimal-deviation hepatoma cell line.

Hybrid molecules were produced by covalently coupling the hormone insulin to the binding chain B of the plant toxin ricin. Binding of the insulin-ricin B hybrid to minimal-deviation hepatoma cells occurred primarily through ricin-specified cell-surface carbohydrates (galactose, N-acetylgalactosamine) since 125I-insulin-ricin B binding to cells could be 90% displaced by 50 mM lactose. [14C]Glucose incorporation into glycogen was maximally stimulated approximately 80% by insulin, whereas maximum stimulation by insulin-ricin B hybrid was greater than 100%. Ricin B chain alone was non-stimulating at concentrations tested (10(-9)-10(-7) M). Furthermore, the stimulation of [14C]glycogen labeling mediated by the hybrid was markedly inhibited by 1 mM lactose, while this sugar had no effect on the stimulation mediated by native insulin. Additionally, a preparation of ricin B shown to actively displace up to 80% of the binding of 125I-hybrid to cells also inhibited hybrid-mediated [14C]glycogen production. These results indicate that insulin-ricin B hybrid molecules possess toxin-specified binding abilities while evoking the insulin-associated cellular response of stimulated incorporation of [14C]glucose into glycogen. Such results thus suggest the possibility that alternate cell-surface receptors may play a role in conveying insulin's intracellular metabolic-control signals.

Animals↗
PubMed1983

Guinea pig line 10 hepatocarcinoma model: characterization of monoclonal antibody and in vivo effect of unconjugated antibody and antibody conjugated to diphtheria toxin A chain.

Monoclonal antibodies were raised against the guinea pig line 10 (L10) hepatocarcinoma, and an IgG1-producing hybridoma (D3) was selected for further study. D3 is a true monoclonal antibody as demonstrated by two-dimensional gel electrophoresis. Radioimmunoassays on live cells revealed no cross-reactivity with normal tissues or with the line 1 hepatocarcinoma which was used as a control. Membrane immunofluorescence assays demonstrated similar specificity. Immunoperoxidase staining of cryostat sections of tumor and normal tissues of both adult animals and fetuses showed that the D3 monoclonal antibody reacted primarily with the L10 tumor, but some cross-reactivity with smooth muscle, placenta, fetal skeletal muscle, and fetal liver was also demonstrated. Radioimmunoprecipitation of detergent extracts of iodinated L10 cells showed that the antigen is present on the cell surface as a dimer of Mr 290,000 (unit size, Mr 148,000). Therapy studies with unconjugated D3 antibody demonstrated a minor dose-dependent effect on tumor growth. D3 antibody conjugated to the A chain of diphtheria toxin (10(-7) M) was cytotoxic to 100% of L10 cells in vitro. Animals treated with a single 1-mg i.v. injection of this immunoconjugate on Day 7 following the intradermal injection of 10(5) tumor cells demonstrated a highly significant inhibition of tumor growth compared to control animals and those treated with unconjugated antibody.

Animals↗
PubMed1981

Mannose 6-, fructose 1-, and fructose 6-phosphates inhibit human natural cell-mediated cytotoxicity.

In vitro human natural cell-mediated cytotoxicity (NCMC) to K-562, Molt-4, and F-265 cells is inhibited in a dose-dependent manner by mannose 6-phosphate, fructose 1-phosphate and fructose 6-phosphate. This inhibition is not observed with mannose, glucose, fucose, glucose 6-phosphate, mannose 1-phosphate, galactose 1-phosphate, or galactose 6-phosphate. Preincubation of the effector cells, obtained from fresh whole blood, with mannose-6-phosphate, fructose-1-phosphate, or fructose-6-phosphate did not inhibit cytotoxicity, which indicated that these hexose phosphates are not nonspecifically toxic towards the effector lymphocytes. Mannose-6-phosphate and the stereochemically similar fructose-1-phosphate are more potent inhibitors than fructose-6-phosphate in terms of concentration required and time of onset of effect. Inhibition of cytotoxicity by mannose-6-phosphate varied with target cell type: F-265 is protected at much lower concentrations of mannose-6-phosphate (less than 1 mM) than is either Molt-4 or K-562. The inhibition of NCMC is also observed with the inhibitors of lysosomal function, NH4Cl, and chloroquine. The presence of a functional mannose-6-phosphate receptor on target cells was demonstrated: (i) Gelonin, a seed protein that inactivates the eukaryotic ribosome but is nontoxic to intact cells, was covalently linked to monophosphopentamannose, and this conjugate ws toxic to both K-562 and F-265 target cells, the latter being by far the more sensitive; and (ii) chloroquine, NH4Cl, and mannose-6-phosphate all inhibited the toxicity of gelonin-monophosphopentamannose. These results suggest either that a cytolytic lymphokine contains a hexose phosphate residue and may be taken up by target cells through the lysosomal/mannose 6-phosphate pathway or that such a residue is involved in target cell-effector cell recognition.

Cytotoxicity, Immunologic↗
PubMed1979

A hybrid protein containing the toxic subunit of ricin and the cell-specific subunit of human chorionic gonadotropin. I. Synthesis and characterization.

In order to study the mechanisms involved in the interaction of cell-surface components and effector proteins, we synthesized a double-labeled dimeric hybrid molecule consisting of the toxic A subunit of the lectin ricin and the cell-specific beta subunit of human chorionic gonadotropin (hCGbeta). Utilizing methyl-5-bromovalerimidate, a disulfide cross-linked conjugate of hCGbeta and toxin subunit A was prepared in 30% yield relative to hCGbeta input. The dimeric protein, purified by gel filtration and ion exchange chromatography, was characterized by: (a) sodium dodecyl sulfate-containing polyacrylamide gel electrophoresis in the presence and absence of 2-mercaptoethanol, (b) analysis by gel filtration chromatography in the presence and absence of 2-mercaptoethanol, and (c) precipitation with antibodies specific for hCG and subunit A.

Animals↗
PubMed1979

A hybrid protein containing the toxic subunit of ricin and the cell-specific subunit of human chorionic gonadotropin. II. Biologic properties.

The biologic properties of a purified hybrid protein containing the toxic subunit A of ricin and the cell-specific subunit of human chorionic gonadotropin (hCGbeta) in a disulfide conjugate is reported. The biologic activity of the hybrid has been studied by assaying inhibition of protein synthesis in rat R2C Leydig cells and mouse L-cells. The results indicate that for the A subunit to exhibit its toxic effect it must first bind to cells through receptors which are specific for the beta subunit of the hybrid. Thus, only cells with beta-type receptors are sensitive to the toxic effects, which hCG is capable of blocking, while cells which are not target cells for hCG are not subject to the toxic effects. Thus it is concluded that the disulfide conjugate may act as a functional analog of ricin toxin but with altered cell specificity. Or, alternately, the hybrid may be regarded as an analog of hCG but with drastically altered physiologic effects.

Animals↗
PubMed1975

Isolation and characterization of a bacteriophage polysaccharide.

A high molecular weight heteropolysaccharide, composed of glucose, glucuronic acid, N-acetylglucosamine, and mannose in an approximate molar ratio of 1:2:2:5, respectively, was isolated from phage K-2 and from the soluble fraction of phage-infected Aerobacter aerogenes lysates. Treatment of pure phage with 8 M urea at 4 degrees quantitatively solubilizes the bound polysaccharide and capsular polysaccharide (Yurewicz, E.C., Ghalambor, M.A., Duckworth, D.H., and Heath, E.C. (1971) J. Biol. Chem. 246, 5607-5616) with the release of only traces of other phage constituents; on this basis, it was concluded that the polysaccharide, like the the glycanohydrolase, is externally localized in the phage structure. Phage polysaccharide and glycanohydrolase fractionate similarly on ion exchange resins and gel electrophoresis in sodium dodecyl sulfate, but each may be purified to homogeneity by the procedures employed. The biosynthesis of the polysaccharide was shown to be uniquely dependent upon phage K-2 infection by: (a) absence of the polysaccharide in cells, the culture filtrate, or sonicated extracts of uninfected cells; (b) kinetics of polysaccharide synthesis following phage infection; and (c) isotopic double-labeling experiments that demonstrated the synthesis of polysaccharide only after initiation of phage replication in infected cells.

Acetylglucosamine↗
PubMedDate not supplied

An antimelanoma-barley ribosome inactivating protein conjugate is cytotoxic to melanoma cells in vitro.

The ribosome inactivating protein (BRIP) from barley is a single polypeptide chain (Mr = 32,000 Dalton) and is nontoxic to intact cells. The BRIP has been purified to homogeneity by modifications of the methods of Roberts and Selitrennikoff and crosslinked to monoclonal antibodies by Succinimidyl-3 (2- Pyridyldithio) -Propionate (SPDP) and by the cystamine-EDAC methods. The resulting hybrids were purified from the free BRIP by gel filtration on a Sephadex G-75 column. The model suicide transport agents were assayed against melanoma cells; K-562 cells were used as control. The hybrids were found to be selectively toxic to melanoma cells in a dose dependent manner.

Antibodies, Monoclonal↗