Prophylactic and therapeutic intervention of Punta Toro virus (Phlebovirus, Bunyaviridae) infection in hamsters with interferon alfacon-1.

Punta Toro virus (PTV) is a member of the Bunyaviridae family, genus Phlebovirus, related to the highly pathogenic Rift Valley fever virus (RVFV). It produces a disease in hamsters that models severe Rift Valley fever (RVF) in humans. The recent outbreak of RVF in Kenya stresses the need to identify prophylactic and therapeutic measures for preventing and treating severe forms of disease. To this end, interferon (IFN) alfacon-1 (consensus IFN-alpha) was evaluated in cell culture against RVFV and PTV, and in the hamster PTV infection model. Survival outcome following treatment initiated pre- and post-virus challenge and the suppression of viral burden and liver disease in infected hamsters was determined. Pre-treatment of cell cultures with IFN alfacon-1 induced marked antiviral activity against both viruses. Intraperitoneal treatment of hamsters initiated 4 h prior to infection with PTV was highly protective and greatly limited liver disease and systemic and liver viral burden. Complete protection from a highly lethal challenge dose was afforded by treatment initiated 36 h following viral inoculation. Although efficacy was much reduced, IFN alfacon-1 therapy was still beneficial when started as late as 3-5 days post-virus exposure. These studies suggest that IFN alfacon-1 may be an effective treatment for early intervention following infection with RVFV.

Is the anti-psychotic, 10-(3-(dimethylamino)propyl)phenothiazine (promazine), a potential drug with which to treat SARS infections? Lack of efficacy of promazine on SARS-CoV replication in a mouse model.

Phenothiazine and derivatives were tested for inhibition of SARS-CoV replication. Phenothiazine slightly inhibited SARS-CoV replication in a neutral red (NR) uptake assay. Adding a propylamino group to give promazine reduced virus yields (VYR assay) with an EC(90)=8.3+/-2.8 microM, but without selectivity. Various substitutions in the basic phenothiazine structure did not promote efficacy. Phenazine ethosulfate was the most potent compound by VYR assay (EC(90)=6.1+/-4.3 microM). All compounds were toxic (IC(50)=6.6-74.5 microM) except for phenoxathiin (IC(50)=858+/-208 microM) and 10-(alpha-diethylamino-propionyl) phenothiazine.HCl (IC(50)=195+/-71.2 microM). Consequently, none were selective inhibitors of SARS-CoV replication (SI values <1-3.3 microM). These data portended the poor efficacy of promazine in a SARS-CoV mouse lung replication model. Intraperitoneal treatment with promazine using a prophylactic (-4h)/therapeutic regimen of 1, 10, or 50mg/(kg day) did not reduce virus lung titers at day 3, yet prolonged virus replication to 14 days. Similar therapeutic promazine doses were not efficacious. Thus, promazine did not affect SARS-CoV replication in vitro or in vivo, nor were any other phenothiazines efficacious in reducing virus replication. Therefore, treating SARS infections with compounds like promazine is not warranted.

Viral regulation of aquaporin 4, connexin 43, microcephalin and nucleolin.

The current study investigated whether human influenza viral infection in midpregnancy leads to alterations in proteins involved in brain development. Human influenza viral infection was administered to E9 pregnant Balb/c mice. Brains of control and virally-exposed littermates were subjected to microarray analysis, SDS-PAGE and western blotting at three postnatal stages. Microarray analysis of virally-exposed mouse brains showed significant, two-fold change in expression of multiple genes in both neocortex and cerebellum when compared to sham-infected controls. Levels of mRNA and protein levels of four selected genes were examined in brains of exposed mice. Nucleolin mRNA was significantly decreased in day 0 and day 35 neocortex and significantly increased in day 35 cerebellum. Protein levels were significantly upregulated at days 35 and 56 in neocortex and at day 56 in cerebellum. Connexin 43 protein levels were significantly decreased at day 56 in neocortex. Aquaporin 4 mRNA was significantly decreased in day 0 neocortex. Aquaporin 4 protein levels decreased in neocortex significantly at day 35. Finally, microcephalin mRNA was significantly decreased in day 56 neocortex and protein levels were significantly decreased at 56 cerebellum. These data suggest that influenza viral infection in midpregnancy in mice leads to long-term changes in brain markers for enhanced ribosome genesis (nucleolin), increased production of immature neurons (microcephalin), and abnormal glial-neuronal communication and neuron migration (connexin 43 and aquaporin 4).

The role of cerebellar genes in pathology of autism and schizophrenia.

Schizophrenia and autism are neurodevelopmental diseases that have genetic as well as environmental etiologies. Both disorders have been associated with prenatal viral infection. Brain imaging and postmortem studies have found alterations in the structure of the cerebellum as well as changes in gene expression. Our laboratory has developed an animal model using prenatal infection of mice with human influenza virus that has demonstrated changes in behavior, pharmacology, structure, and gene expression in the brains of exposed offspring. In the current communication we describe altered expression of cerebellar genes associated with development of brain disorder in a mouse model for schizophrenia and autism and correlate these changes with those involved in the pathology of these two disorders.

Differential pathogenesis of cowpox virus intranasal infections in mice induced by low and high inoculum volumes and effects of cidofovir treatment.

The causes of death from intranasal cowpox virus infections in mice remain unclear. Hypotheses include severe pneumonitis, hepatitis and/or hyperproduction of cytokines and chemokines. This work explores these hypotheses by studying the influence of low- and high-volume virus inocula on viral pathogenesis. BALB/c mice were infected intranasally with a syncytium-forming variant of cowpox virus in 5 microL or 50 microL volumes containing the same infectious virus challenge dose. The 50 microL infection produced a more rapidly lethal disease associated with severe pneumonitis, high lung and nasal virus titres and increased cytokine and chemokine levels in the lungs and nasal tissue, whilst liver infection was minimal. The 5 microL inoculum infection was also lethal, but the infection was primarily confined to the upper respiratory tract and included elevated nasal cytokine and chemokine levels. Levels of the pro-inflammatory cytokine interleukin-6 were particularly high in both infections. Treatment of the infections with cidofovir (100mg/kg/day for 2 days starting 24h after virus exposure) led to survival and suppression of tissue virus titres. Treatment reduced pneumonitis in the 50 microL infection and lessened cytokine hyperproduction in both infections. We conclude that a 5 microL volume inoculum of cowpox virus causes a lethal upper respiratory tract infection, whilst the 50 microL inoculum targets both upper and lower respiratory tracts, with excessive release of systemic pro-inflammatory factors. Cidofovir effectively treated both infections and slowed viral replication sufficiently to subdue the exaggerated release of pro-inflammatory mediators.

Potent in vitro activity of the albumin fusion type 1 interferons (albumin-interferon-alpha and albumin-interferon-beta) against RNA viral agents of bioterrorism and the severe acute respiratory syndrome (SARS) virus.

BACKGROUND: The type 1 interferons (INF-alpha and INF-beta) are potent antiviral agents. Albumin-INF-alpha and albumin-INF-beta are novel recombinant proteins consisting of IFN-alpha or IFN-beta genetically fused to human albumin. METHODS: The in vitro antiviral activity of albumin-IFN-alpha was evaluated against representative bioterrorism viral agents and the severe acute respiratory syndrome virus. Antiviral activity was assessed using inhibition of cytopathic effect and neutral red staining. RESULTS: EC(50) values for albumin-IFN-alpha ranged from <0.1 ng/ml for Punta Toro virus to 65 ng/ml for Venezuelan equine encephalitis virus in the neutral red assay. Albumin-IFN-beta showed 75- and 360-fold greater in vitro activity than albumin-IFN-alpha against Ebola virus and severe acute respiratory syndrome, respectively. CONCLUSION: Further evaluation of these long-acting albumin-IFN fusion proteins as prophylactic or therapeutic agents against these viral agents of bioterrorism in relevant primate models is warranted.

Correlation between breakdown of the blood-brain barrier and disease outcome of viral encephalitis in mice.

Changes in the permeability of the blood-brain barrier (BBB) were evaluated in two mouse models of viral encephalitis. The ability of sodium fluorescein (NaFl) to cross the BBB from the serum into the central nervous system was assayed in animals inoculated with virulent strains of either Banzi or Semliki Forest viruses. To test the hypothesis that increases in BBB permeability were associated with poor disease outcome subsequent experiments measured BBB permeability in conjunction with treatment with the interferon inducer Ampligen (poly I:poly C(12)U). A single intraperitoneal injection of Ampligen (1 mg/kg) administered either 24 h or 4-6 h before, but not 24 h after, virus inoculation with Banzi virus provided significant improvements in survival, viral brain titers, weight change and BBB permeability. In comparison, a similar treatment with Ampligen administered either 24 h or 4-6 h before inoculation with Semliki Forest virus was able to significantly improve weight change, and BBB permeability, but only animals receiving Ampligen 4-6 h pre-virus showed a significantly improved mortality. In general, it was found that evaluation of BBB permeability was a more sensitive indicator of disease outcome and the antiviral efficacy Ampligen than either weight change or brain viral titers.

Efficacy of N-methanocarbathymidine in treating mice infected intranasally with the IHD and WR strains of vaccinia virus.

N-Methanocarbathymidine [(N)-MCT] is a newly identified inhibitor of orthopoxvirus replication in cell culture and in mice. Limited published animal studies indicated the compound is effective by intraperitoneal (i.p.) route at 10-100 mg/(kg day). More extensive studies using different treatment regimens in intranasally infected mice were conducted in order to further explore the potential of this compound compared to cidofovir in treating vaccinia virus infections. (N)-MCT was given twice a day for 7 days, whereas cidofovir was administered once a day for 2 days, each starting 24h after virus exposure for most experiments. (N)-MCT was not toxic up to 1000 mg/(kg day) by the i.p. treatment route. Oral and i.p. treatment regimens with (N)-MCT were directly compared during a vaccinia virus (IHD strain) infection, indicating that the nucleoside has good oral bioavailability in mice. Treatments by i.p. route with (N)-MCT (100 mg/(kg day)) reduced lung, nasal, and brain virus titers during an IHD virus infection, but not nearly to the same extent as i.p. cidofovir (100 mg/(kg day)). Treatment with both compounds decreased liver, spleen, and kidney virus titers, as well as reduced lung consolidation scores and lung weights. Onset of treatment could be delayed by 2 days with (N)-MCT and by 3 days with cidofovir, providing significant survival benefit during the IHD virus infection. Against a vaccinia virus (WR strain) infection in mice, i.p. (N)-MCT treatment prevented death at 500 mg/(kg day), which was comparable in activity to i.p. cidofovir (100 mg/(kg day)). Significant reductions in tissue virus titers occurred with both treatment regimens. (N)-MCT could be further pursued for its potential to treat orthopoxvirus infections in humans.

Prophylactic treatment with recombinant Eimeria protein, alone or in combination with an agonist cocktail, protects mice from Banzi virus infection.

A recombinant Eimeria protozoan protein antigen (rEA) has been shown to have antitumor and antiviral activity. The purpose of this study was to determine the effect of rEA treatment alone or in combination with an agonist cocktail consisting of granulocyte macrophage colony stimulating factor (GM-CSF), interferon gamma (IFN-gamma), interleukin 4 (IL-4), and anti CD-40 antibody, in the treatment of Banzi virus (BV) disease in BALB/c mice. Treatment with rEA resulted in a significant increase in survival, weight gain, and mean day to death in BV-infected mice and resulted in a significant decrease in brain virus titer. Treatment with rEA, in combination with a 4-agonist cocktail, improved disease parameters to a greater degree than rEA treatment alone. The effect of treatment with a reduced concentration of agonist cocktail or fewer components of the agonist cocktail, in combination with rEA, on disease outcome in BV-infected mice was also investigated. Treatment with rEA, alone or in combination with agonist cocktail, 24h after virus challenge did not improve disease. Treatment with rEA, alone or in combination with an agonist cocktail, is efficacious for the prophylaxis of BV infection in mice.

Comparison of the inhibitory effects of interferon alfacon-1 and ribavirin on yellow fever virus infection in a hamster model.

Antiviral compounds were evaluated for efficacy against yellow fever virus (YFV) in a hamster model of YFV-induced liver disease. Challenge with a 10(2) 50% cell culture infectious doses of YFV resulted in a 50-80% mortality rate in female hamsters. Virus was detected by quantitative real-time RT-PCR (QRT-PCR) in liver, kidney, spleen and serum with peak titers on 4-6 days post-viral challenge (dpi). Serum levels of alkaline phosphatase, alanine aminotransferase (ALT), bilirubin, blood urea nitrogen, potassium and creatinine were significantly elevated, while serum levels of albumin, amylase, glucose, calcium, globulin, phosphorus, sodium and total protein were significantly reduced. Packed cell volume and white blood cell count were significantly elevated during the course of the infection. Intraperitoneal treatment of hamsters with 0.5-5 microg/kg/day interferon (IFN) alfacon-1, 100mg/kg/day viramidine or 50 mg/kg/day ribavirin, initiated 4h prior to YFV challenge, resulted in significant improvement in survival and serum ALT levels. Treatment with IFN alfacon-1 or ribavirin starting 2dpi, also significantly improved survival and serum ALT levels in hamsters challenged with YFV. Pre- and post-virus exposure treatment with IFN alfacon-1 was efficacious in improving disease in YFV-infected hamsters.

Cell line dependency for antiviral activity and in vivo efficacy of N-methanocarbathymidine against orthopoxvirus infections in mice.

A novel carbocyclic thymidine analog, N-methanocarbathymidine [(N)-MCT], was evaluated for inhibition of orthopoxvirus infections. Efficacy in vitro was assessed by plaque reduction assays against wild-type and cidofovir-resistant strains of cowpox and vaccinia viruses in nine different cell lines. Minimal differences were seen in antiviral activity against wild-type and cidofovir-resistant viruses. (N)-MCT's efficacy was affected by the cell line used for assay, with 50% poxvirus-inhibitory concentrations in cells as follows: mouse=0.6-2.2 microM, rabbit=52-90 microM, monkey=87 to >1000 microM, and human=39-220 microM. Limited studies performed with carbocyclic thymidine indicated a similar cell line dependency for antiviral activity. (N)-MCT did not inhibit actively dividing uninfected cells at 1000 microM. The potency of (N)-MCT against an S-variant thymidine kinase-deficient vaccinia virus was similar to that seen against S-variant and wild-type viruses in mouse, monkey, and human cells, implicating a cellular enzyme in the phosphorylation of the compound. Mice were intranasally infected with cowpox and vaccinia viruses followed 24h later by intraperitoneal treatment with (N)-MCT (twice a day for 7 days) or cidofovir (once a day for 2 days). (N)-MCT treatment at 100 and 30 mg/kg/day resulted in 90 and 20% survival from cowpox virus infection, respectively, compared to 0% survival in the placebo group. Statistically significant reductions in lung virus titers on day 5 occurred in 10, 30, and 100mg/kg/day treated mice. These same doses were also active against a lethal vaccinia virus (WR strain) challenge, and protection was seen down to 10mg/kg/day against a lethal vaccinia virus (IHD strain) infection. Cidofovir (100mg/kg/day) protected animals from death in all three infections.

Effect of oral gavage treatment with ZnAL42 and other metallo-ion formulations on influenza A H5N1 and H1N1 virus infections in mice.

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; P < 0.001). This regimen was also well tolerated by the mice. Manganese and selenium formulations were not inhibitory to virus replication when given therapeutically. Mice were also infected with influenza A/NWS/33 (H1N1) virus and were treated 48 h before virus exposure with three dosages of ZnAL42 (8.64, 1.46 or 0.24 mg/kg/day). Treatment was by oral gavage twice daily for 13 days. The highest dose of ZnAL42 was significantly inhibitory to the virus infection as seen by prevention of deaths and lessening of decline in SaO2. 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.

Synthesis of cyclopentenyl carbocyclic nucleosides as potential antiviral agents against orthopoxviruses and SARS.

A practical and convenient methodology for the synthesis of chiral cyclopentenol derivative (+)-12a has been developed as the key intermediate that was utilized for the synthesis of biologically active carbocyclic nucleosides. The selective protection of allylic hydroxyl group followed by the ring-closing metathesis (RCM) reaction with Grubbs catalysts provided (+)-12a on a 10 g scale with 52% overall yield from D-ribose (4). The key intermediate (+)-12a was utilized for the synthesis of unnatural five-membered ring heterocyclic carbocyclic nucleosides. The newly synthesized 1,2,3-triazole analogue (17c) exhibited potent antiviral activity (EC(50) 0.4 microM) against vaccinia virus and moderate activities (EC(50) 39 microM) against cowpox virus and severe acute respiratory syndrome coronavirus (SARSCoV) (EC(50) 47 microM). The 1,2,4-triazole analogue (17a) also exhibited moderate antiviral activity (EC(50) 21 microM) against SARSCoV.

Respiratory syncytial virus infections: recent prospects for control.

Respiratory syncytial virus (RSV) infections remain a significant public health problem throughout the world, although recently developed and clinically approved anti-RSV antibodies administered prophylactically to at-risk populations appear to have significantly affected the disease development. Much effort has been expended to develop effective anti-RSV therapies, using both in vitro assay systems and mouse, cotton rat, and primate models, with several products now in various stages of clinical study. Several products are also being considered for the treatment of clinical symptoms of RSV. In this review, updates on the status of the approved anti-RSV antibodies, ribavirin, and recent results of studies with potential new anti-RSV compounds are summarized and discussed.

Enhancement of the infectivity of SARS-CoV in BALB/c mice by IMP dehydrogenase inhibitors, including ribavirin.

Because of the conflicting data concerning the SARS-CoV inhibitory efficacy of ribavirin, an inosine monophosphate (IMP) dehydrogenase inhibitor, studies were done to evaluate the efficacy of ribavirin and other IMP dehydrogenase inhibitors (5-ethynyl-1-beta-D-ribofuranosylimidazole-4-carboxamide (EICAR), mizoribine, and mycophenolic acid) in preventing viral replication in the lungs of BALB/c mice, a replication model for severe acute respiratory syndrome (SARS) infections (Subbarao, K., McAuliffe, J., Vogel, L., Fahle, G., Fischer, S., Tatti, K., Packard, M., Shieh, W.J., Zaki, S., Murphy, B., 2004. Prior infection and passive transfer of neutralizing antibody prevent replication of severe acute respiratory syndrome coronavirus (SARS-CoV) in the respiratory tract of mice. J. Virol. 78, 3572-3577). Ribavirin given at 75 mg/kg 4 h prior to virus exposure and then given twice daily for 3 days beginning at day 0 was found to increase virus lung titers and extend the length of time that virus could be detected in the lungs of mice. Other IMP dehydrogenase inhibitors administered near maximum tolerated doses using the same dosing regimen as for ribavirin were found to slightly enhance virus replication in the lungs. In addition, ribavirin treatment seemed also to promote the production of pro-inflammatory cytokines 4 days after cessation of treatment, although after 3 days of treatment ribavirin inhibited pro-inflammatory cytokine production in infected mice, significantly reducing the levels of the cytokines IL-1alpha, interleukin-5 (IL-5), monocyte chemotactic protein-1 (MCP-1), and granulocyte-macrophage colony stimulating factor (GM-CSF). These findings suggest that ribavirin may actually contribute to the pathogenesis of SARS-CoV by prolonging and/or enhancing viral replication in the lungs. By not inhibiting viral replication in the lungs of infected mice, ribavirin treatment may have provided a continual source of stimulation for the inflammatory response thought to contribute to the pathogenesis of the infection. Our data do not support the use of ribavirin or other IMP dehydrogenase inhibitors for treating SARS infections in humans.

Protective immunity against acute phleboviral infection elicited through immunostimulatory cationic liposome-DNA complexes.

Cationic liposome-DNA complexes (CLDC) have been demonstrated to induce potent antitumor activities. The ability of these complexes to elicit protective immunity against viral infections has not been fully explored. Here we report findings on the use of CLDC as an antiviral agent in a mouse model of acute phleboviral (Punta Toro virus) disease. CLDC treatment of mice challenged with Punta Toro virus (PTV) resulted in dramatic increases in survival and reduced viral burden and other parameters indicative of protection against disease. CLDC were effective when administered by intraperitoneal and intravenous routes and elicited protective immunity when given within 1 day of virus challenge. Treatments administered 36 h or longer after challenge, however, were not effective in preventing mortality or disease. CLDC treatment induced release of a number of potential antiviral cytokines including IFN-gamma, IL-12, and IFN-alpha. Taken together, our findings indicate that non-specific immunotherapy with CLDC appears to be an effective treatment for blocking PTV-induced disease and suggests that further exploration in other viral disease models may be warranted.

Activities of oseltamivir and ribavirin used alone and in combination against infections in mice with recent isolates of influenza A (H1N1) and B viruses.

Mouse models have been widely used for evaluating potential influenza virus inhibitors. However, the viral strains traditionally used in these models are fairly old and do not represent currently circulating viruses in nature. We developed two new lethal infection models in mice using mouse-adapted influenza A/New Caledonia/20/99 (H1N1) and influenza B/Sichuan/379/99 viruses. Both virus infections were used to study oral treatment with oseltamivir and ribavirin, both alone and in combination. Oral treatments were given twice daily for 5 days starting 4 h before infection in initial studies. Against influenza A, oseltamivir was active at 10, 20, and 40 mg/kg/day, protected 80-100% of mice from death and reduced lung consolidation - ribavirin was similarly effective at 20, 40, and 80 mg/kg/day. When treatments were initiated after virus challenge, delaying treatment with oseltamivir even 1 day caused it to be ineffective. Ribavirin prevented mortality by 50-80% when treatments were delayed 1-4 days after infection. The combination of the two drugs (oseltamivir at 20 mg/kg/day and ribavirin at 40 mg/kg/day) was no better than ribavirin alone. In contrast to what we observed with influenza A virus infections, oseltamivir and ribavirin showed similar dose-related antiviral activities against influenza B virus infections. The compounds both significantly increased survival when treatments started up to 4 days after infection, but ribavirin was more active than oseltamivir (50-80% survival compared to 30-40% survival, respectively, when starting treatments on days 2-4 after infection). By varying the doses of each drug that were used in combination (oseltamivir at 1.25, 2.5 and 5 mg/kg/day; ribavirin at 5, 10 and 20 mg/kg/day) certain dosage combinations were superior to either compound used alone as assessed by decreased mortality, lung virus titre, lung score and lung weight parameters. These activities differed from published results with older, more established virus strains as oseltamivir was less effective and ribavirin was more active than previously reported.

Anti-influenza virus activities of 4-[(1,2-dihydro-2-oxo-3H-indol-3-ylidene)amino]-N-(4,6-dimethyl-2-pyrimidin-2-yl)benzenesulphonamide and its derivatives.

4-[(1,2-Dihydro-2-oxo-3H-indol-3-ylidene)amino]-N-(4,6-dimethyl-2-pyrimidinyl)-benzenesulphonamide (SPIII-5H) and related compounds were tested for antiviral activity against influenza A (H1N1, H3N2, and H5N1) and B viruses in Madin Darby canine kidney (MDCK) cell culture. Among the compounds tested, SPIII-5H and four derivatives (5-chloro [SPill-5Cl], 5-bromo [SPIII-5Br], 5-methyl [SPIII-5Me] and N-acetyl [SPIII-NA]) showed similar antiviral potencies, with only the 5-fluoro (SPIII-5F) derivative being ineffective. Fifty percent effective concentration (EC50) values were determined in cytopathic effect (CPE) inhibition assays quantified by neutral red dye uptake. By this method, the active compounds were inhibitory to the H1N1 strain of influenza A at 2.7-5.2 microg/ml, to the H3N2 strain of influenza A at 13.8-26.0 microg/ml, to the H5N1 strain of influenza A at 3.1-6.3 microg/ml and to influenza B at 7.7-11.5 microg/ml. Confirmatory virus yield reduction studies against influenza A (H1N1) virus demonstrated antiviral activity (90% inhibition) at concentrations of 2-10 microg/ml. No cytotoxic effects were evident in actively growing uninfected cells or stationary monolayers at 100 microg/ml. Potencies of the compounds were similar to those of ribavirin, but much less than those of oseltamivir carboxylate against the various viruses. Time-of-addition studies indicated the compounds inhibited an early step in the virus replication cycle, probably virus adsorption/penetration, and no virucidal activity was evident. The basic molecule is amenable to diverse chemical modifications, which may improve water solubility and antiviral potency.

Antiviral activity of nucleoside analogues against SARS-coronavirus (SARS-coV).

The recent outbreak of severe acute respiratory syndrome (SARS), which is an acute respiratory illness, is caused by newly discovered SARS coronavirus (SARS-CoV). Herein we describe the antiviral activity of several classes of nucleoside analogues evaluated against SARS-CoV in Vero 76 cells, some of which exhibited moderate activity.

Combinatorial ribavirin and interferon alfacon-1 therapy of acute arenaviral disease in hamsters.

Several arenaviruses endemic to South America (Junin, Machupo, and Guanarito) and Africa (Lassa) are known to cause frequently fatal haemorrhagic fever. With the exception of ribavirin, which has demonstrated efficacy in cases of Lassa fever, there is no other effective therapeutic for the treatment of arenaviral haemorrhagic fever. We have recently reported that consensus interferon-a (IFN alfacon-1) can protect hamsters from lethal Pichinde virus (PCV) infection, which serves as a model for acute arenaviral disease in humans. Here we demonstrate highly effective therapy through the combined use of ribavirin with IFN alfacon-1 for the treatment of PCV infection in hamsters. Ribavirin was given orally, twice per day for 7 days, and IFN alfacon-1 was administered intraperitoneally once per day for 10 days. Treatments were initiated 1-5 days post-virus challenge using various dose combinations, many of which were less than optimal when the drugs were given independently. Combining suboptimal doses of ribavirin (5-10 mg/kg/day) with IFN alfacon-1 (5-10 microg/kg/day), we were able to demonstrate increased protection from mortality, reduced viral burden and liver disease, and greatly extended survival times as compared to treatments where drugs were administered alone. Our data indicate that combination therapy results in synergistic activity that may slow down the progression of the disease and decrease fatality rates associated with severe arenaviral infections in humans. Further, combination therapy reduces the effective dosage of ribavirin, which would serve to limit its toxicity.