The effect of interferon-gamma treatment of rat tumour cells on their susceptibility to natural killer cell, macrophage and cytotoxic T-cell killing.

The effect of exposing tumour cells to interferon-gamma (IFN-gamma) has been studied investigating changes in MHC antigen expression and susceptibility to a variety of cellular effector mechanisms. Treatment of rat tumour cell lines with rat recombinant IFN-gamma increased their expression of class I MHC molecules, as monitored by quantitative immunofluorescence. Reduced sensitivity to lysis by NK cells was observed, although cold-target competition assays indicated NK cells were bound equally well by both interferon-treated and control cells.

Reverse transformation of Harvey murine sarcoma virus-transformed NIH/3T3 cells by site-selective cyclic AMP analogs.

Eighteen site-selective cAMP analogs modified at either the C-8 position or the C-6 position were tested for their growth regulatory effects on the Harvey murine sarcoma virus-transformed NIH/3T3 clone 13-3B-4 cells grown in a serum-free defined medium. All 18 analogs, when tested individually, exhibited an appreciable growth inhibitory effect at micromolar concentrations. The most potent growth inhibitory analogs contained a thio moiety at the C-8 position.

Site-selective cAMP analogs at micromolar concentrations induce growth arrest and differentiation of acute promyelocytic, chronic myelocytic, and acute lymphocytic human leukemia cell lines.

Cyclic AMP (cAMP)-dependent protein kinase may play a role in the functional and morphological differentiation of leukemic cells. In this study, we showed that the cAMP analogs, potent activators of protein kinase recently shown to be selective for either site 1 or site 2 cAMP binding sites of protein kinase, demonstrate potent growth inhibition of acute promyelocytic, chronic myelocytic, and acute lymphocytic leukemic cell lines with no sign of toxicity. The growth inhibition accompanied monocytic differentiation in HL-60 cells and a loss of nuclear terminal deoxynucleotidyl transferase activity in Molt-4 leukemic cells.

Site-selective cAMP analogs induce nuclear translocation of the RII cAMP receptor protein in Ha-MuSV-transformed NIH/3T3 cells.

Site-selective cAMP analogs, depending on the position of their substituents on the adenine ring, selectively bind to either site 1 or site 2 of the known cAMP binding sites of protein kinase. Treatment of Harvey murine sarcoma virus-transformed NIH/3T3 cells with such site-selective analogs results in growth inhibition and phenotypic reversion, and the combination of a C-8 thio or halogen analog (site 1 selective) with an N6 analog (site 2 selective) produces a synergistic effect. We report here that the growth inhibitory effect of the analogs correlates with the nuclear translocation of the RII cAMP receptor protein, the regulatory subunit of protein kinase type II.

The effects of immobilization on the biochemical, physiological and morphological features of Anabaena azollae.

Anabaena azollae, a presumptive isolate from Azolla filiculoides, was immobilized in polyurethane foam, hydrophilic polyvinyl foam and alginate. When viewed by low-temperature scanning electron microscopy a thick mucilage layer covered the surface of both cells and matrix; this closely resembles the mode of attachment of the symbiont Anabaena in the Azolla leaf cavity. The heterocyst frequency of the immobilized A.

Site-selective cyclic AMP analogs provide a new approach in the control of cancer cell growth.

Site-selective cyclic AMP analogs bind to site 1 or site 2 of the known cAMP-binding sites depending on the position of substituents on the purine ring, either at C-2 and C-8 (site 1) or at C-6 (site 2). The growth inhibitory effect of such site-selective cAMP analogs used in this investigation with 15 human cancer cell lines surpassed that of analogs previously tested. The most potent analogs were 8-chloro, N6-benzyl and N6-phenyl-8-p-chlorophenylthio-cAMP.

Production of Hyoscyamine by 'Hairy Root' Cultures of Datura stramonium.

'Hairy root' cultures of DATURA STRAMONIUM were established following infection of aseptic leaves with AGROBACTERIUM RHIZOGENES. Transformation was confirmed by Southern blotting using [ (32)P]-labelled fragments of the T-DNA as probes. The transformed cultures grew in the absence of added phytohormones and cell mass increased 55-fold during 28 days incubation.

Quantitation of MHC antigen expression on colorectal tumours and its association with tumour progression.

A flow cytometric technique has been established for accurately quantitating the cell surface density of MHC antigens and the percentage of cells expressing MHC antigens in 38 colorectal tumours. Thirty-four percent of tumours were partially or completely negative for HLA-ABC antigen expression. Although the quantity of HLA-ABC antigens varied widely, there was no correlation between the density of HLA-ABC antigens, or the percentage of cells expressing these antigens and clinicopathological stage.

Novel pyrazolo[3,4-d]pyrimidine nucleoside analog with broad-spectrum antiviral activity.

A novel nucleoside analog, 4(5H)-oxo-1-beta-D- ribofuranosylpyrazolo[3,4-d]pyrimidine-3-thiocarboxamide (N10169), was evaluated in cell culture and in animals for antiviral activity against DNA and RNA viruses. The compound was highly active against strains of adeno-, vaccinia, influenza B, paramyxo-, picorna-, and reoviruses, with 50% inhibition of virus-induced cytopathology at 1 to 10 microM. Lesser or no antiviral effects were observed against herpes simplex, cytomegalo-, corona-, influenza A, vesicular stomatitis, and visna viruses.

Quinoline Alkaloid Production by Transformed Cultures of Cinchona ledgeriana.

A culture of CINCHONA LEDGERIANA, transformed with AGROBACTERLUM TUMEFACIENS A6 capable of growing and producing quinoline alkaloids in medium free of exogenous phytohormones has been obtained. Unlike the untransformed culture of this species, addition of ZR in combination with either IAA or IBA to medium did not affect alkaloid production. Growing the transformed culture in the dark, however, produced a marked enhancement of alkaloid accumulation, up to 50 times that of cultures grown in the light.

Synthesis of 5'-O-beta-D-glucopyranosyl and 5'-O-beta-D-galactopyranosyl derivatives of ribavirin.

The synthesis of 5'-O-beta-D-glucopyranosyl and 5'-O-beta-D-galactopyranosyl derivatives (13 and 15, respectively) of the antiviral agent ribavirin are described. Direct glycosylation of 2',3'-O-isopropylideneribavirin with either tetra-O-acetyl-alpha-D-glucopyranosyl bromide (4) or tetra-O-acetyl-alpha-D-galactopyranosyl bromide (8) under Koenigs-Knorr conditions (i.e.

Synthesis of certain nucleoside methylenediphosphonate sugars as potential inhibitors of glycosyltransferases.

The synthesis of alpha-D-glucopyranosyl 1-(methylenediphosphonate) (11), alpha-D-galactopyranosyl 1-(methylenediphosphonate) (14), and alpha-D-mannopyranosyl 1-(methylenediphosphonate) (17) has been accomplished. [(Di-phenoxyphosphinyl)methyl]phosphonic acid (diphenyl-MDP) (5), synthesized by two different procedures, was fused with beta-D-glucopyranose pentaacetate followed by catalytic hydrogenation to give 2,3,4,6-tetra-O-acetyl-alpha-D-glucopyranosyl methylenediphosphonate (glucose-MDP) (10). The anomeric configuration of 10 was assigned on the basis of NMR spectral studies.

Synthesis of glycopyranosylphosphonate analogues of certain natural nucleoside diphosphate sugars as potential inhibitors of glycosyltransferases.

The synthesis of alpha-D-glucopyranosyl-, alpha-D-galactopyranosyl-, and alpha-D-mannopyranosylphosphonate is described. Condensation of tris(trimethylsilyl) phosphite with 2,3,4,6-tetrakis-O-(phenylmethyl)-1-O-acetyl-alpha-D-glucopyranose generated 2,3,4,6-tetrakis-O-(phenylmethyl)-alpha-D-glucopyranosylphosphonic acetic anhydride (13). The benzyl blocking groups were removed by catalytic hydrogenation, and the anhydride bond was cleaved by alkaline hydrolysis to obtain alpha-D-glucopyranosylphosphonate (15).

Identification of a monoclonal antibody-defined breast carcinoma antigen in body fluids.

The monoclonal antibody NCRC-11 defines antigens associated with secretory glandular epithelia as well as most epithelial malignancies. These components have been identified in, and isolated from, normal body fluids including urine and skim milk. The immunoadsorbent purified antigens from urine and milk were very similar to those purified from breast and ovarian carcinomas; by sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS PAGE) and immunoblotting, NCRC-11 antibody-binding antigens from all sources were of high apparent molecular weight (greater than 400 kD) with the major component(s) present as a single band or a doublet.

A spectrophotometric assay for strictosidine synthase.

A spectrophotometric assay for strictosidine synthase is described. Strictosidine is extracted with ethyl acetate and, where high substrate concentrations are used, the organic extract is washed with dilute ammonia to remove coextracted secologanin; after evaporation of the solvent, the residue is heated with 5 M H2SO4 for 45 min and the A348 value is measured. Strictosidine production is calculated from the response of similarly treated standards.

Synthesis, structure, and biological activity of certain 2-deoxy-beta-D-ribo-hexopyranosyl nucleosides and nucleotides.

2-Deoxy-beta-D-ribo-hexopyranosyl nucleosides with adenine (2), hypoxanthine (17), guanine (23), cytosine (13), and uracil (7) as the aglycon were synthesized by the Lewis-acid catalyzed condensation of an appropriate trimethylsilylated heterocyclic base and 2-deoxy-1,3,4,6-tetrakis-O-(4-nitrobenzoyl)-beta-D-ribo-hexopyranose+ ++ (5) to provide the desired beta anomers in good yield. When the synthesis of 7 via an SN2 displacement was attempted by reaction between silylated uracil and 2-deoxy-3,4,6-tris-O-(4-nitrobenzoyl)-alpha-D-ribo-hexopyranosyl bromide (8), the major product, 1-(2-deoxy-3,4,6-tris-O-(4-nitrobenzoyl)-alpha-D-ribo-hexopyranosyl)-2,4 - pyrimidinedione (9), had retained the alpha configuration at the anomeric carbon. The structures of both anomers of 1-(2-deoxy-D-ribo-hexopyranosyl)-2,4-pyrimidinedione were assigned by single-crystal X-ray methods.

Potential for use of nicotinic acid as a selective agent for isolation of high nicotine-producing lines of Nicotiana rustica hairy root cultures.

The addition of exogenous nicotinic acid, nicotinamide or nicotine was studied with reference to their effects on growth and alkaloid production by hairy root cultures of Nicotiana rustica. Nicotinic acid and nicotinamide were toxic (50% phytostatic dose being 2.4 and 9 mM respectively) while nicotine was not toxic below 10 mM.

Toxicity of quinoline alkaloids to cultured Cinchona ledgeriana cells.

The toxicity of Cinchona alkaloids to cell cultures of C. ledgeriana has been studied in relation to alkaloid uptake and possibilities for selecting high-yielding cell lines. The most toxic, quinine, was completely toxic at 5.

Tumour aneuploidy, prognostic parameters and survival in primary breast cancer.

Cellular DNA content of primary tumours from 280 patients with operable breast cancer was determined by flow cytometry using nuclei from paraffin sections stained with DAPI, and 199 of these patients were followed for 8-13 years after surgery. Tumours from 67 patients have also been analyzed for their DNA content using single cell suspensions from fresh tumour tissue stained with mithramycin and ethidium bromide, and the results compared with those obtained from paraffin blocks of the same tumours. Overall 60% of the tumours contained cells with abnormal DNA content (DNA-aneuploid populations).

Synergistic activity of purine metabolism inhibitors in cultured human tumor cells.

Cytotoxic effects of 3-deazaguanosine (3-DGUO) result from the inhibition of DNA synthesis and incorporation of the drug into DNA. Synergistic antiproliferative effects of a combination of 3-deazaguanosine and 2-beta-D-ribofuranosylthiazole-4-carboxamide, a potent inhibitor of inosine monophosphate dehydrogenase, was observed in human tumor cells. Inosine reversed the antiproliferative effects of the 3-deazaguanosine but not 2-beta-D-ribofuranosylthiazole-4-carboxamide.

Biological activity of analogs of guanine and guanosine against American Trypanosoma and Leishmania spp.

The growth inhibitory effects of six guanine and guanosine analogs, 3-deazaguanine (compound 1); 3-deazaguanosine (compound 2); 6-aminoallopurinol (compound 3); 9-beta-xylofuranosyl guanine (compound 4); a ribosylated derivative of compound 3, 6-aminopyrazolo(3,4-d)pyrimidin-4-one (compound 5); and 5-aminoformycin B (compound 6), were tested against some pathogenic members of the family of American Trypanosomatidae. Compounds 1 and 2 were highly active against Trypanosoma cruzi, Trypanosoma rangeli, and American Leishmania spp. in in vitro culture forms.

Use of tiazofurin to enhance the metabolism and cytotoxic activities of analogues of guanine, guanosine, and deoxyguanosine.

An effective modulator of cellular guanine nucleotide pools, 2-beta-D-ribofuranosylthiazole-4-carboxamide (tiazofurin) was tested for its ability to affect utilization of certain guanine, guanosine, and deoxyguanosine analogues by Chinese hamster ovary cells and hypoxanthine guanine phosphoribosyltransferase (HGPRTase)-deficient variants. The nucleoside analogues investigated were chosen for their potential to be metabolized to the nucleotide level by pathways other than those requiring the action of HGPRTase. Exposure of tiazofurin-treated (500 microM for 3 h) cells to 3-deazaguanosine (200 microM for 3 h) resulted in enhanced 3-deazaGTP formation and an increase (5-10-fold) in the ratio 3-deazaGTP/GTP.

2'-Deoxy-3,7-dideazaguanosine and related compounds. Synthesis of 6-amino-1-(2-deoxy-beta-D-erythro-pentofuranosyl) and 1-beta-D-arabinofuranosyl-1H-pyrrolo[3,2-c]pyridin-4(5H)-one via direct glycosylation of a pyrrole precursor.

The synthesis of two new analogs of 2'-deoxyguanosine, 6-amino-1-(2-deoxy-beta-D-erythro-pentofuranosyl)-1H-pyrrolo[3,2-c] pyridin-4(5H)-one (8) and 6-amino-1-beta-D-arabinofuranosyl-1H-pyrrolo[3,2-c]-pyridin-4(5H)-one (13) has been accomplished by glycosylation of the sodium salt of ethyl 2-cyanomethyl-1H-pyrrole-3-carboxylate (4c) using 1-chloro-2-deoxy-3,5-di-O-p-toluoyl-alpha-D-erythro-pentofuranose( 5) and 1-chloro-2,3,5-tri-O-benzyl-alpha-D-arabinofuranose (9), respectively. The resulting blocked nucleosides, ethyl 2-cyanomethyl-1-(2-deoxy-3,5-di-O-p-toluoyl-beta-D-erythro- pentofuranosyl)-1H-pyrrole-3-carboxylate (6) and ethyl 2-cyanomethyl-1-(2,3,5-tri-O-benzyl-beta-D-arabinofuranosyl)- 1H-pyrrole-3-carboxylate, were ring closed with hydrazine to form 5-amino-6-hydrazino-1-(2-deoxy-beta-D-erythro-pentofuranosyl)-1H- pyrrolo[3,2-c]-pyridin-4(5H)-one (7) and 5,6-diamino-1-(2,3,5-tri-O-benzyl-beta-D-arabinofuranosyl)-1H- pyrrolo[3,2-c]pyridin-4(5H)-one (11), respectively. Treatment of 7 with Raney nickel provided the 2'-deoxyguanosine analog 8 while reaction of 11 with Raney nickel followed by palladium hydroxide/cyclohexene treatment gave the 2'-deoxyguanosine analog 13.