NMR investigations of the role of the sugar moiety in glycosylated recombinant human granulocyte-colony-stimulating factor.

Human granulocyte-colony-stimulating factor (G-CSF) is a hematopoietic growth factor that plays a major role in the stimulation of the proliferation and maturation of granulocyte neutrophil cells. With the recent increased understanding of its biological properties in vivo together with available preparations of recombinant human G-CSF, this growth factor has become an essential agent for clinical applications. The presence of an O-linked carbohydrate chain at position 133 greatly improves the physical stability of the protein.

Comparison of the potency of glycosylated and nonglycosylated recombinant human granulocyte colony-stimulating factors in neutropenic and nonneutropenic CD rats.

Recent studies in human bone-marrow culture and healthy human volunteers suggest that lenograstim [glycosylated, recombinant human granulocyte colony-stimulating factor (rHuG-CSF) produced in Chinese hamster ovary (CHO) cells] has greater in vivo potency than filgrastim [nonglycosylated, methionine-extended recombinant human granulocyte colony-stimulating factor (rmetHuG-CSF) produced in Escherichia coli]. To confirm and extend these results we investigated the in vivo potency of both products in normal rats and neutropenic CD rats as an animal model of chemotherapy-induced neutropenia. In normal rats, groups of eight normal male CD rats received four subcutaneous doses of 10, 30, or 100 micrograms/kg filgrastim or lenograstim on days 1-4 of the study, whereas a control group received the vehicle.

Metastatic phenotype of murine tumor cells expressing different cooperating oncogenes.

Four murine cellular tumor models expressing various combinations of oncogenes (SV40 large T and v-Ha-ras, SV40 large T and v-src, SV40 large T and neu, adenovirus EIA and v-Ha-ras) induce sarcoma when they are inoculated s.c. into the DBA/2 syngenic mice.

Mode of action of the antitumor compound girodazole (RP 49532A, NSC 627434).

Girodazole (RP 49532A) or 3-amino-1-[4-(2 amino-1H-imidazolyl]-propanol, 2HCl is an experimental antitumor compound which inhibits protein synthesis in cell cultures and in cell free systems. The compound has been evaluated for its capacity to inhibit specific assays of initiation, elongation and termination of protein synthesis. Girodazole inhibited the release of nascent peptides from polyribosomes in rabbit reticulocyte lysates indicating that the major effect of the compound is on the protein synthesis termination step.

Antitumor activity and mechanism of action of the marine compound girodazole.

Girodazole, a new marine compound has been isolated from the sponge Pseudaxinyssa cantharella. Girodazole is active in vivo on several murine grafted tumors including leukemias (P388, L1210, i.p.

Synthesis of new (+-)-3,5-dihydroxypentyl nucleoside analogues from 1-amino-5-(benzyloxy)pentan-3-ol and their antiviral evaluation.

The synthesis and antiviral evaluation of a series of (+-)-3,5- dihydroxypentyl nucleoside analogues related to acyclic nucleoside antiviral agents are reported. All purine and pyrimidine nucleoside analogues described in this paper have been obtained from 1-amino-5-(benzyloxy)pentan-3-ol. A synthesis of this amine is reported from 1-(benzyloxy)but-3-ene after epoxidation and regiospecific diethylaluminum chloride catalyzed opening of the epoxide by trimethylsilyl cyanide.

Inhibition of simian virus 40 DNA replication in CV-1 cells by an oligodeoxynucleotide covalently linked to an intercalating agent.

An octathymidylate covalently linked via its 3'-end to an acridine derivative inhibited the cytopathic effect of Simian Virus SV40 on CV-1 cells in culture. Control experiments revealed that this effect was virus-specific and did not arise as a result of oligonucleotide degradation by nucleases. A photoactive probe was covalently attached to the 5'-end of the oligonucleotide-acridine conjugate.

Protective activity of tetracycline analogs against the cytopathic effect of the human immunodeficiency viruses in CEM cells.

Tetracycline analogs were evaluated for anti-HIV activity in CEM cells; minocycline and doxycycline were the most active of these in inhibiting the virus-induced cytopathic effect between 7 and 14 days post-infection. The active concentrations (0.3-1.

Selective inhibition of tyrosine protein kinase by a synthetic multisubstrate analog.

Synthetic compounds were designed in an attempt to mimic the possible transition state of tyrosine protein kinases. One representative compound (RP 53801) inhibited the enzyme purified from RSV-transformed cells. A serine/threonine kinase (kinase C) was 45 fold less sensitive.

Selective inhibition of the cytopathic effect of type A influenza viruses by oligodeoxynucleotides covalently linked to an intercalating agent.

Oligodeoxynucleotides covalently linked to an acridine derivative were targeted to part of the 3'-terminal sequence which is common to the eight RNAs of type A influenza viruses. The cytopathic effect of the virus on MDCK cells in culture was strongly decreased by a heptanucleotide covalently attached to the acridine ring. Control experiments using other oligonucleotide sequences showed that the effect was specific for the complementary sequence of the 3'-terminal region of the viral RNAs.

Inactivation of herpes simplex virus by protein components of bovine neutrophil granules.

From an acid extract of granules of bovine neutrophils we isolated fractions of cationic proteins, exhibiting significant anti-herpesvirus activity at concentrations which were devoid of cytotoxicity and of activity against a picornavirus (rhinovirus). The mechanism of action seems to involve a direct neutralization of the virions. Two antiviral peptides with an approximate MW 7500 were purified to homogeneity by reversed phase high-performance liquid chromatography.

Antirhinovirus compound 44,081 R.P. inhibits virus uncoating.

44,081 R.P., or 2-[(1,5,10,10a-tetrahydro-3H-thiazolo[3,4-b]isoquinolin- 3-ylidine)amino]-4-thiazole acetic acid, is a compound which selectively inhibits rhinovirus in cell cultures.

(+/-)-2-Amino-3,4-dihydro-7-[2,3-dihydroxy-4-(hydroxymethyl)-1- cyclopentyl]-7H-pyrrolo[2,3-d]pyrimidin-4-ones: new carbocyclic analogues of 7-deazaguanosine with antiviral activity.

5-Allyl-2-amino-4,6-dihydroxypyrimidine (3) was chlorinated and ozonized to yield (2-amino-4,6-dichloro-pyrimidin-5-yl)acetaldehyde (5). Acetalization of 5 with ethanol afforded a new pyrimidine intermediate 6 which can lead to 2-amino-3,4-dihydro-7-alkyl-7H-pyrrolo[2,3-d]pyrimidin-4-ones and therefore to carbocyclic analogues of 7-deazaguanosine. The 7-substituent was a cyclopentyl analogue of the arabinofuranosyl moiety in 10a, lyxofuranosyl moiety in 10b, and ribofuranosyl moiety in 10c.

[Use of permeabilized cells in the study of inhibitory products of herpes virus replication].

Cells made permeable by exposure to lysolecithin following infection by HSV-1 synthesize DNA (in greater amounts than non-infected cells) in the presence of the four deoxyribonucleoside-triphosphates (dNTPs) : dATP, dCTP, dGTP, and dTTP. DNA synthesis also occurs if dTTP is replaced by dT or dTMP, indicating activity of enzymes such as thymidine kinase, thymidylate kinase, deoxyribonucleoside-diphosphate kinase and ADN polymerase. Examination of DNA synthesis in permeabilized cells enables detection of antiviral activity of agents incapable of penetrating into intact cells and therefore ineffective in cell cultures.

Studies on 44 081 R.P., a new antirhinovirus compound, in cell cultures and in volunteers.

A synthetic compound, 2-[(1,5,10,10a-tetrahydro-3H-thiazolo[3,4b]isoquinolin-3-ylidene) amino]-4-thiazoleacetic acid (S), 44 081 R.P., inhibits the multiplication of rhinoviruses in cell cultures.

Immunostimulating hexapeptide from human casein: amino acid sequence, synthesis and biological properties.

A hexapeptide obtained from human casein by enzymatic digestion has been purified, sequenced and synthesized; its structure is: Val-Glu-Pro-Ile-Pro-Tyr. In vitro this hexapeptide stimulates the phagocytosis of opsonized sheep red blood cells by murine peritoneal macrophages. Administered intravenously to adult mice, it enhances the resistance to infection with Klebsiella pneumoniae.

[Comparative effects of 4 cephalosporins on the incorporation of tritiated diaminopimelic acid during bacterial growth].

When comparing antibiotic activities, it might be of interest to study parameters other than minimal inhibitory concentrations (MIC's). Specific activity of beta-lactams on bacterial cell wall makes it possible to determine radiolabelled diaminopimelic acid (DAP) incorporation in growing cultures. We have studied the effects of various concentrations of cefalotin , cefotaxime, latamoxef (moxalactam) and ceftiolene (42 980 RP) on DAP incorporation in 6 strains of E.

Synergistic activities of type I (alpha, beta) and type II (gamma) murine interferons.

Type I (alpha, beta) and type II (gamma) murine interferons are able to potentiate each other with respect to the inhibition of encephalomyocarditis (EMC) virus and of herpes simplex virus type 1 (HSV-1) multiplication in a murine cell line (DBT). Examination of two double-stranded RNA-dependent enzymes in DBT cells, the 2-5A synthetase and the 67,000 MW protein phosphokinase indicates that mixed interferon preparations act synergistically at least with respect to an increase in the activity of the former enzyme. The results obtained with gamma interferons of different origin and of different specific activity suggest that interferon itself, rather than the lymphokines present in the interferon preparations, is responsible for the synergistic effect.

Immunostimulating substances from human casein.

Delipidated human casein was digested with trypsin and the enzymatic digest was fractionated on Sephadex G-50. The peptidic fractions were assayed for immunomodulating activity in two in vitro models. Fractions corresponding to molecular weights in the range of 2,000 +/- 600 were found to possess stimulating activity in these models.

Comparative effects of an inflammatory reaction on the resistance of mice to bacterial and viral infections.

The induction in mice of a sterile subcutaneous granuloma exerted no influence upon the mortality following their infection with herpes type 1, murine hepatitis or encephalomyocarditis viruses. Attempts to reproduce the resistance -- which has been found to occur as a result of the granulomatous reaction, in the case of bacterial, fungal or protozoa infections and tumour invasions -- by varying the route and timing of the virus inoculation or the strain of mice have failed. We conclude that it is not merely through their inflammatory properties that some non-specific immunostimulating substances enhance resistance against viral infection.

[Immunostimulating and adjuvant activities of a low molecular weight lipopeptide].

From crude extracts of a Streptomyces strain exhibiting immunopotentiating effects, a tetrapeptide was isolated and its structure established as L Ala leads to D isoGlu leads to L, L Dap comes from Gly. This peptide was devoid of biological activity but its chemical coupling with lauric acid gave a substance endowed with adjuvant and immunostimulating properties. This substance and the corresponding synthetic lauroyltetrapeptide were as active in this respect as the muramyl-dipeptide, thus far considered as the minimal adjuvant-active structure of bacterial cell walls: the presence of a sugar moiety is therefore not a prerequisite for immunopotentiating activities.

Glycolipids stimulate DNA polymerase activity in a DNA-membrane fraction and in a partially purified polymerase system extracted from pneumococci.

We have assayed the ability of various lipids to affect DNA polymerases activity in a DNA-membrane complex extracted from Streptococcus pneumoniae by the Sarkosyl-M-band technique. In addition, to determine which DNA polymerases were affected by the lipids, we partially purified three DNA polymerase activities from cell lysates, the first such demonstration outside of Escherichia coli and Bacillus subtilis. Glycolipids are unique among polar lipids in stimulating the rate and extent of DNA polymerase activity in M-bands and in Sarkosyl lysates from which the M-band is derived.

Membrane fusion: studies with a calcium-sensitive dye, arsenazo III, in liposomes.

Fusion between vesicles, cells, or organelles may be defined as confluence of two membrane-bound compartments without access of their solutes to external milieu. To study fusion by this criterion, we have trapped the metallochromic calcium-sensitive dye, arsenazo III (AIII), partially calcium-saturated (AIII-Ca) in one population of liposomes (phoshatidylcholine 90:dicetylphosphate 10), and ethylene glycol-bis(beta-aminoethyl ether)-N,N'-tetraacetate (EGTA) in a second. In such mixtures, interaction of EGTA with AIII-Ca was measured by a large color shift from blue leads to red (decreased absorbance at 660 nm).