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Background: The antipoxviral activities and phosphorylation of N-methanocarbathymidine ([ N]-MCT) and four 5-halo-2'-deoxyuridines, namely 5-fluoro- (FdU), 5-chloro- (CldU), 5-bromo- (BrdU), and 5-iodo- (IdU) derivatives, were explored. Methods: Antiviral activities and nucleoside metabolism were determined in C127I mouse, LLC-MK2 monkey, and A549 human cells infected with thymidine-kinase-containing and -deficient (TK+and TK−) vaccinia (WR strain) viruses. Results: The antiviral potencies of CldU, BrdU and IdU were increased 16–26-fold in LLC-MK2 cells infected with TK+compared with TK−virus infections, but enhancement of activity was much less in the other cell lines. ( N)-MCT was nearly equally active against TK+and TK−viruses in the three cell lines. Antiviral activity of FdU was associated with cytotoxicity. Uninfected and infected cells metabolized compounds to mono-, di- and triphosphates. The thymidine, BrdU and IdU triphosphate levels were higher in C127I and LLC-MK2 cells infected with TK+than with TK−virus. ( N)-MCT monophosphate levels were much higher in TK+virus-infected cells, but without corresponding increases in ( N)-MCT triphosphate. Furthermore, TK+virus infections did not appreciably alter ( N)-MCT triphosphate levels in other mouse (L929), monkey (MA-104 and Vero) and human cell lines (A549). Antiviral potency of the compounds was greater in C127I than in LLC-MK2 cells, yet lower intracellular triphosphate levels were found in C127I cells. Conclusion: We conclude that viral TK plays an important role in increasing the antiviral potencies of these compounds in some cell lines, but minimally in others. These findings may have implications in treating infected animals with compounds that are dependent upon poxvirus TK for their activation, because viral TK activity may vary greatly due to cell type.

We designed and synthesized novel 2,3-disubstituted quinazolin-4(3 H)-ones by microwave technique and characterized them by spectral analysis. Synthesized compounds were screened for cytotoxicity and for antiviral activity against influenza A (H1N1, H3N2 and H5N1), severe acute respiratory syndrome corona, dengue, yellow fever, Venezuelan equine encephalitis (VEE), Rift Valley fever, and Tacaribe viruses in cell culture. A neutral red uptake assay was used to determine 50% virus-inhibitory concentrations (EC50) of test compounds and their 50% cytotoxicity concentration (CC50) in uninfected Madin-Darby canine kidney, Vero, and Vero 76 cells; selectivity indices (ratio of CC50 to EC50) were derived from the data. The compound 4-(6,8-dibromo-4-oxo-2-phenyl quinazolin-3(4 H)-yl)-N-(4,5-dimethyloxazol-2yl) benzenesulphonamide 15 inhibited the replication of avian influenza (H5N1) virus (EC50=8.4 µg/ml, CC50>100 µg/ml, SI>11.9) as did 4-(6-bromo-4oxo-2phenylquinazolin-3(4 H)-yl) benzene]sulphonamide 5 (EC50=3 µg/ml, CC50=32 µg/ml, SI=11). Compound 5 was also moderately active against VEE and Tacaribe viruses. The methodology described in this report is applicable for rapid synthesis of many compounds with potential antiviral properties.

Avian influenza H5N1 infections can cause severe, lethal human infections. Whether influenza A virus treatments effectively ameliorate avian influenza H5N1 human infections is uncertain. The research objective was to evaluate the efficacy of novel zinc and other metallo-ion formulations in two influenza A mouse models. Mice infected with influenza A/Duck/MN/1525/81 (H5N1) virus were treated orally 48 h before virus exposure and then twice daily for 13 days with ZnAL42. The optimal dosing regimen for ZnAL42 was achieved at 17.28 mg/kg 48 h prior to virus exposure, twice daily for 7 days. The survival rate was 80% compared with 10% in the untreated control group and a 100% survival rate with ribavirin (75 mg/kg/day, twice a day for 5 days, beginning 4 h before virus exposure). ZnAL42 treatment significantly lessened the decline in arterial oxygen saturation (SaO2; P2. The data suggest that the prophylactic use of ZnAL42 is effective against avian influenza H5N1 or H1N1 virus infection in mice and should be further explored as an option for treating human influenza virus infections.

Measles virus (MV) infections have been almost eradicated in some industrialized nations. However, MV continues to cause severe disease and mortality in the world and is responsible for clusters of exogenous-borne disease in essentially disease-free countries. Because of the ebb and flow of immunization campaigns, especially in the poverty-stricken and war-torn Third World, and the ominous potential for severe disease and mortality, it is vital that research for discovery of therapeutic countermeasures should continue. To that end, a number of compounds have been evaluated for efficacy in vitro and in animal models, and several therapeutic modalities have been tested in the clinic. The only current therapies used in the clinic include ribavirin administered orally or intravenously, alone or in combination with immune serum globulin; these therapies have demonstrated variable efficacy. Therefore, drug discovery efforts have been launched to supplement the existing treatments for MV infections. Antisense molecules, adenosine and guanosine nucleosides, including ring-expanded ‘fat’ nucleoside analogues, brassinosteroids, coumarins, peptide inhibitors, modulators of cholesterol synthesis and a variety of natural products have been screened for efficacy and toxicity both in vitro and in animals. However, none of these agents has gone into human clinical trials and most will not merit further development due to toxicity concerns and/or low potency. Thus, further research is needed to develop more potent and less toxic drugs that could be used for treating MV infections to supplement the existing MV vaccine campaigns.

We evaluated two types of compounds for efficacy in inhibiting SARSCoV replication in vitro: calpain inhibitors (a class of cellular cysteine proteinases) and a number of nucleoside analogues. Cytopathic effect reduction assays visually determined with spectrophotometric verification by neutral red (NR) uptake assay were used to evaluate cytotoxicity and antiviral potency of the compounds. Significantly inhibitory compounds were then evaluated in virus yield reduction assays. Two calpain inhibitors, Val-Leu-CHO (calpain inhibitor VI) and Z-Val-Phe-Ala-CHO (calpain inhibitor III), were the most potent inhibitors of SARSCoV. By virus yield reduction assay, calpain inhibitor VI had a 90% effective concentration (EC90) of 3 μM and calpain inhibitor III had an EC90 of 15 μM. β-D-N4-hydroxycytidine was the most selective nucleoside analogue inhibitor with an EC90 of 6 μM by virus yield reduction assay. These compounds or analogues warrant further evaluation as potential therapies for treating SARS or could be used as lead compounds for discovery of more potent SARSCoV inhibitors.

A series of acyclic phosphonomethylether nucleosides were synthesized and then evaluated for inhibitory activity against respiratory viruses of clinical significance using CPE inhibition, neutral red uptake and virus yield reduction assays. Of the 20 compounds synthesized, none significantly inhibited influenza A or B viruses or respiratory syncytial virus strains A2, Long or 18537; the selective indices (SI) were less than 10. A new compound, GS-2128 (2R, 5R-9-[2,5-dihydro-5-(phosphonomethoxy)-2-furanyl]adenine; D4API), selectively inhibited adenovirus 5 (SI>10) as did GS-0577 (9-(3-hydroxy-2-phosphonylmethoxypropyl)-adenine; HPMPA) and GS-0504 [(S)-1-[3-hydroxy-2-(phosphonylmethoxypropyl)]-cytosine; HPMPC]. The 50% effective concentrations (EC50) ranged from 8–100 μg mL−1 and 50% cell inhibitory concentrations (CC50) from 40–1000 μg mL−1. All three compounds were also found to be active against laboratory strains and clinical isolates of adenovirus types 1, 2, 8 and 41 with EC50 values ranging from 0.2 to 10 μg mL−1. Two compounds, GS-438 (9-(2-phosphonylmethoxyethyl)guanine, PMEG) and GS-2542 (9-[3-phosphonomethoxy)methoxymethyl]guanine) inhibited parainfluenza virus 3 strain C243, with SI of 52 and >333, respectively. PMEG also inhibited measles virus strains CC, Halonen and Chicago with EC50 values ranging from 0.03–9 μg mL−1. These data suggest that these compounds should be considered for possible development as therapeutic agents for respiratory virus infections.

A series of germanium- or silicon-centred heteropolytungstates (polyoxometalates) with the Barrel, Keggin or double Keggin structure were evaluated in vitro for their effects against influenza A (IV-A) and B (IV-B) viruses. Their 50% effective concentrations (EC50) against recent isolates of IV-A (H1N1) and IV-B ranged from 0.1 to 7.8 μM against IV-A (H3N2), the EC50 concentrations were often 10-fold higher. Recent clinical isolates of IV-A were generally more susceptible to these antiviral effects than older, laboratory-adapted strains. These experiments used inhibition of viral CPE in MDCK cells as determined microscopically and by Neutral Red (NR) uptake. Virus yield reduction studies indicated the 90% effective concentrations (EC90) ranged from 0.2 to 32 μM against these viruses. Cytotoxic or cell inhibitory concentrations (CC50), determined by NR uptake and total cell count, ranged from 38 to 189 μM, indicating high selective indices for some of these compounds. Altering time of addition of an active compound relative to infecting cells with IV-A (HINl) showed greatest efficacy when given early in viral replication. Five of the most active polyoxometalates were evaluated against IV-B infections in mice using intraperitoneal treatment beginning 4 h prior to virus exposure. Two of the compounds, one with the Barrel structure and the other with a double Keggin structure, were particularly inhibitory, preventing deaths, reducing arterial oxygen decline and lowering lung consolidation. Lung virus titres were reduced by a maximum of 0.7 log10. Therapy initiated 8 h post-virus exposure was not effective against this in vivo infection.