Broad-spectrum antiviral activity of Virazole: 1-beta-D-ribofuranosyl-1,2,4-triazole-3-carboxamide.

Virazole is a synthetic nucleoside active in tissue culture against at least 16 DNA and RNA viruses. Applied topically, it inhibits herpetic keratitis in rabbits and tail lesions induced by herpes, vaccinia, and vesicular stomatitis viruses in mice. Injected intraperitoneally into mice, it inhibits splenomegaly and hepatomegaly induced by Friend leukemia virus and respiratory infections caused by influenza A(O), A(2), and B viruses and parainfluenza 1 virus. infections is also effective.

Synthesis and biochemical evaluation of nucleosides of naphthoquinone heterocycles.

The synthesis, characterization, and biochemical evaluation of 1-beta-D-ribofuranosylnaphtho[2,3-d]imidazole-4,9-dione (3), 2-beta-D-ribofuranosylnaphtho[2,3-d]pyrazole-4,9-dione (6), and 2-beta-D-ribofuranosylnaphthol[2,3-d]triazole-4,9-dione (9) are reported. These quinone nucleosides and the corresponding quinone heterocycles were tested as inhibitors of purine nucleotide biosynthesis in Ehrlich ascites cells. The nucleosides 3 and 9 and naphtho[2,3-d]imidazole-4,9-dione were effective inhibitors of hypoxanthine phosphoribosyltransferase.

Synthesis of 1-(4-thio-beta-D-ribofuranosyl)-1,2,4-triazole-3-carboxamide.

The synthesis of 1-(4-thio-beta-D-ribofuranosyl)-1,2,4-triazole-3-carboxamide (3) is described. The acid-catalyzed fusion procedure with 3-cyano-1,2,4-triazole and 1,2,3,5-tetra-O-acetyl-4-thio-D-ribofuranose provided 3-cyano-1-(2,3,5-tri-O-acetyl-4-thio-beta-D-ribofuranosyl-1,2,4-triazole (2) which was converted with NH4OH to 3. In contrast to 1-beta-D-ribofuranosyl-1,2,4-triazole-3-carboxamide (1), the 4'-thio nucleoside 3 did not exhibt significant antiviral activity in vitro.

Synthesis and antiviral acticity of some phosphates of the broad-spectrum antiviral nucleoside, 1-beta-D-ribofuranosyl-1,2,4-triazole-3-carboxamide (ribavirin).

1-beta-D-Ribofuranosyl-1,2,4-triazole-3-carboxamide 5'-phosphate (2) was prepared and converted into the following derivatives: the 5'-phosphoramidate 3, the 5'-diphosphate 4, the 5'-triphosphate 5, and the cyclic 3',5'-phosphate 6. The cyclic 2',3'-phosphate 7 was prepared from the parent nucleoside, 1-beta-D-ribofuranosyl-1,2,4-triazole-3-carboxamide (1), and was opened to the 2'(3')-phosphate 8. These compounds were found to exhibit significant antiviral activity against several viruses in cell culture. Ribavirin 5'-phosphate (2) was shown to be effective when tested against lethal infections in mice caused by influenza A2, influenza B, and murine hepatitis viruses.

Synthesis and antiviral activity of certain thiazole C-nucleosides.

A general reaction of glycosyl cyanides with liquid hydrogen sulfide in the presence of 4-dimethylaminopyridine to provide the corresponding glycosylthiocarboxamides is described. These glycosylthiocarboxamides were utilized as the precursors for the synthesis of 2-D-ribofuranosylthiazole-4-carboxamide and 2-beta-D-ribofuranosylthiazole-5-carboxamide (23). The structural modification of 2-beta-D-ribofuranosylthiazole-4-carboxamide (12) into 2-(2,3,5-tri-O-acetyl-beta-D-ribofuranosyl)thiazole-4-carboxamide (15), 2-beta-D-ribofuranosylthiazole-4-thiocarboxamide (17), and 2-(5-deoxy-beta-D-ribofuranosyl)thiazole-4-carboxamide (19) is also described. These thiazole nucleosides were tested for in vitro activity against type 1 herpes virus, type 3 parainfluenza virus, and type 13 rhinovirus and an in vivo experiment was run against parainfluenza virus. They were also evaluated as potential inhibitors of purine nucleotide biosynthesis. It was shown that the compounds (12 and 15) which possessed the most significant antiviral activity were also active inhibitors (40-70%) of guanine nucleotide biosynthesis.

Synthesis and biological activity of certain 1,2,3-triazole carboxamide nucleosides related to bredinin and pyrazofurin.

5-Hydroxy-1-(beta-D-ribofuranosyl)-1,2,3-triazole-4-carboxamide (II) has been prepared by direct glycosylation of the trimethylsilyl derivative of 5-hydroxy-1,2,3-triazole-4-carboxamide (IV). The use of trimethylsilyltrifluoromethane sulfonate as a catalyst and 1-O-acetyl-2,3,5-tri-O-benzoyl-beta-D-ribofuranose in acetonitrile gave a 95% yield of a 1:1 mixture of V and VI as blocked nucleosides which were readily separated on a silica gel column. The synthesis of II was also achieved by the cycloaddition of 2,3,5-tri-O-benzoyl-beta-D-ribofuranosyl azide with ethyl malonamate. II shows significant antiviral activity against herpes and measles virus in vitro.

Effect of 1-beta-D-ribofuranosyl-1,2,4-triazole-3-carboxamide (ribavirin) on Friend leukemia virus infections in mice.

Ribavirin, when administered intraperitoneally to mice infected with Friend leukemia virus, significantly inhibited development of the disease as determined by inhibition of virus-induced splenomegaly and viable virus titers in the spleen and plasma of the infected animals. The drug was effective whether administered ip in multiple daily treatments, treatments every three days, or single injection. Greatest efficacy was seen when therapy began early in the infection, presumably while the virus was in its eclipse phase.

Suppression by 1-beta-D-ribofuranosyl-1,2,4-triazole-3-carboxamide (virazole, ICN 1229) of influenza virus-induced infections in mice.

1-beta-d-Ribofuranosyl-1,2,4-triazole-3-carboxamide (Virazole) was found to possess significant antiviral activity against aerosol-induced or intranasally induced influenza A(0), A(2), and B virus infections in mice. Significant protection was achieved by both oral and intraperitoneal routes of administration. Depending upon the level of virus infection, antiviral activity was best observed at the daily dose of 75 mg/kg. The efficacy of the compound was evidenced by an increase in survivor number, prolongation of mean survival time, suppression of lung consolidation, or decrease in hemagglutinin titer in the infected lung samples. The therapeutic value of this synthetic triazole nucleoside was evident as noted by a significant increase in survivor number even if the treatment was started as late as 24 h after infection with an aerosol of influenza A(2) virus.

In vitro effect of 1-beta-D-ribofuranosyl-1,2,4-triazole-3-carboxamide (virazole, ICN 1229) on deoxyribonucleic acid and ribonucleic acid viruses.

Virazole (1-beta-d-ribofuranosyl-1,2,4-triazole-3-carboxamide) is a highly soluble new synthetic nucleoside having significant, reproducible activity against a broad spectrum of deoxyribonucleic acid and ribonucleic acid viruses in vitro. The drug inhibited viral cytopathogenic effects in monolayers of cells infected for 3 days with type 3 adeno, types 1 and 2 herpes, myxoma, cytomegalo, vaccinia, infectious bovine rhinotracheitis, types 1A, 2, 8, 13, and 56 rhino, types 1 and 3 parainfluenza, vesicular stomatitis, subacute sclerosing panencephalitis, Semliki Forest, Newcastle disease, and measles viruses. Hemagglutinin production by influenza A(2), influenza B, and type 1 parainfluenza viruses in chicken embryo cells was reduced by Virazole treatment. Recoverable intra- and extracellular virus titers were reduced by the drug in experiments with type 1 herpes, vaccinia, type 3 parainfluenza, and vesicular stomatitis viruses. Plaque formation by type 1 herpesvirus was also inhibited by exposure of the infected cells to Virazole. Pretreatment of cells with the compound, followed by its removal before addition of type 1 herpesvirus, severely lessened the antiviral activity; the compound was still moderately effective in reducing the viral effects on the cells when added as long as 22 hr after the virus. Parallel experiments, in which the antiviral activity of a number of known active drugs was compared, indicated Virazole to have at least a comparable degree of activity, and it was also active against a wider variety of viruses than any of these known active materials. The CCED(50) of Virazole to chicken embryo cells was approximately 1,000 mug/ml, although concentrations as low as 10 mug/ml caused slight (15%) inhibition in total cellular protein after 72 hr of incubation.

In vitro and in vivo effect of 1-beta-D-ribofuranosyl-1,2,4-triazole-3-carboxamide (ribavirin) on types 1 and 3 parainfulenza virus infections.

1-beta-D-ribofuranosyl-1,2,4-triazole-3-carboxamide (ribavirin) had significant in vitro activity against type 1 parainfluenza (Sendai) and type 3 parainogenic (HA-1) viruses. Activity was manifested as inhibition of both viral cytopathogenic effect and of recoverable virus or viral hemagglutinin titer. The minimum Sendai virus inhibitory concentration was determined to be approximately 3.2 mug/ml. Previous studies had determined the minimum concentration inhibiting HA-1 virus was approximately 1-10 mug/ml. The effect of time of addition of ribavirin to virus-infected cells was determined; maximal activity was seen when the drug added just prior to either virus or within 4-8 h after each virus, although anti-Senadi viral effects were still apparent when ribavirin was added as late as 24 h after the virus. Ribavirin had no effect on adsorption of HA-1 or Sendai virus to cells. Lethal Sendai virus infections of mice were significantly inhibited by multiple intraperitoneal ribavirin treatment, starting either 4 h before or up to 24 h after virus inoculation. Therapy starting 48, 72 or 96 h after virus exposure had a moderate degree of efficacy. Treatment using an aerosol chamber also was of moderate effectiveness, although the procedure was considered traumatic to the animals. A nonlethal, principally upper respiratory tract infection of hamsters induced by the HA-1 virus was inhibited by ribavirin therapy. Treatment administered intraperitoneally, per os or by aerosol chamber resulted in reduced 23-day antibody titers to the virus, presumably because of reduction of virus in the animal. In a separate experiment, intraperitoneal ribavirin therapy resulted in a 1 log10 or less reduction in virus titer in nasal washings from HA-1 virus-infected hamsters, whereas, when the drug was administered intranasally in a dry powder aerosol spray, nasal virus titers were reduced up to 2 log10 and a moderate virus-induced lung consolidation was completely inhibited.

Mechanism of action of 1- -D-ribofuranosyl-1,2,4-triazole-3-carboxamide (Virazole), a new broad-spectrum antiviral agent.

The antiviral activity of the synthetic nucleoside, Virazole (1-beta-D-ribofuranosyl-1,2,4-triazole-3-carboxamide), against measles virus in Vero cell cultures was substantially reversed by xanthosine, guanosine, and to a slightly lesser extent by inosine. Virazole 5'-phosphate was subsequently found to be a potent competitive inhibitor of inosine 5'-phosphate dehydrogenase (IMP:NAD(+) oxidoreductase, EC 1.2.1.14) isolated from Escherichia coli (K(m) = 1.8 x 10(-5) M) with a K(i) of 2.7 x 10(-7) M. Guanosine 5'-phosphate (GMP) was a competitive inhibitor of this enzyme with a K(i) of 7.7 x 10(-5) M. Virazole 5'-phosphate was similarly active against IMP dehydrogenase isolated from Ehrlich ascites tumor cells, with a K(i) of 2.5 x 10(-7) M. The K(m) for this enzyme was 1.8 x 10(-5) M, and the K(i) for GMP was 2.2 x 10(-4) M. These results suggest that the antiviral activity of Virazole might be due to the inhibition of GMP biosynthesis in the infected cell at the step involving the conversion of IMP to xanthosine 5'-phosphate. This inhibition would consequently result in inhibition of the synthesis of vital viral nucleic acid.

Effect of 1-beta-D-ribofuranosyl1-1,2,4-triazole-3-carboxamide (virazole, ICN 1229) on herpes and vaccinia keratitis and encephalitis in laboratory animals.

Topical application of 1-beta-d-ribofuranosyl-1,2,4-triazole-3-carboxamide (Virazole) significantly inhibited the development of herpetic keratitis in the eyes of rabbits, as determined by both infectivity and Draize scoring parameters. Significant inhibition of the infection was demonstrated with 10% concentrations of Virazole; a 1% solution had a moderate effect, whereas doses of 0.1 and 0.01% had little activity in this system. A 5% concentration of Virazole similarly inhibited vaccinia keratitis in rabbits. Encephalitis-induced mortality in hamsters initially infected intraocularly with herpesvirus was significantly prevented or inhibited by topical application of 5, 10, and 20% concentrations of Virazole. Surviving, treated hamsters had no signs of herpes keratitis. The 20% concentration was the approximate LD(50) in hamsters. Virazole administered subcutaneously or intraperitoneally to mice did not appreciably alter the course of herpes virus- or vaccinia virus-induced encephalitis in these animals, although in a herpesvirus experiment direct injection of the drug into the brains 3 hr prior to virus inoculation resulted in a significant survivor increase.