Inhibition of Dengue Virus by an Ester Prodrug of an Adenosine Analog

Chen, Yen-Liang; Yin, Zheng; Lakshminarayana, Suresh B.; Qing, Min; Schul, Wouter; Duraiswamy, Jeyaraj; Kondreddi, Ravinder Reddi; Goh, Anne; Xu, Hao; Yip, Andy M.; Liu, Boping; Weaver, M. Libby; Dartois, Véronique; Keller, Thomas H.; Shi, Pei Yong

doi:10.1128/aac.00397-10

Cited by 54 publications

(74 citation statements)

References 20 publications

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“…Other recent studies have documented in vitro inhibition of flavivirus replication by antimetabolites which were also unable to prolong survival in in vivo models of infection (34,35). A few antimetabolites have successfully prolonged survival but also exhibited severe toxicity (36,37). Similarly, we observed that MTX was toxic at high concentrations.…”

Section: Discussionsupporting

confidence: 57%

Flaviviruses Are Sensitive to Inhibition of Thymidine Synthesis Pathways

Fischer

Smith

Shum

et al. 2013

J Virol

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bDengue virus has emerged as a global health threat to over one-third of humankind. As a positive-strand RNA virus, dengue virus relies on the host cell metabolism for its translation, replication, and egress. Therefore, a better understanding of the host cell metabolic pathways required for dengue virus infection offers the opportunity to develop new approaches for therapeutic intervention. In a recently described screen of known drugs and bioactive molecules, we observed that methotrexate and floxuridine inhibited dengue virus infections at low micromolar concentrations. Here, we demonstrate that all serotypes of dengue virus, as well as West Nile virus, are highly sensitive to both methotrexate and floxuridine, whereas other RNA viruses (Sindbis virus and vesicular stomatitis virus) are not. Interestingly, flavivirus replication was restored by folinic acid, a thymidine precursor, in the presence of methotrexate and by thymidine in the presence of floxuridine, suggesting an unexpected role for thymidine in flavivirus replication. Since thymidine is not incorporated into RNA genomes, it is likely that increased thymidine production is indirectly involved in flavivirus replication. A possible mechanism is suggested by the finding that p53 inhibition restored dengue virus replication in the presence of floxuridine, consistent with thymidine-less stress triggering p53-mediated antiflavivirus effects in infected cells. Our data reveal thymidine synthesis pathways as new and unexpected therapeutic targets for antiflaviviral drug development. Dengue virus (DENV) poses a significant human health risk for approximately 40% of the world's population (http://www.who .int/mediacentre/factsheets/fs117/en/). DENV occurs as four serotypes, with increasing and widespread circulation of all serotypes considered a contributing factor for the rising incidence of dengue disease. A particularly severe complication is dengue hemorrhagic fever (DHF), which results in approximately 22,000 deaths annually and of which the incidence is currently on the rise (http://www .who.int/csr/disease/dengue/impact/en/index.html). However, even nonfatal, self-limiting DENV infection can result in a severe, painful disease that is also known as "break-bone fever." No vaccine or antiviral therapeutics are licensed to address DENV infections, and treatment is currently limited to supportive care.Many studies are ongoing with the hopes of identifying effective novel anti-DENV therapeutics and/or potential therapeutic targets. Several viral factors have been explored as therapeutic targets, including the viral protease, helicase, and RNA-dependent RNA polymerase (reviewed in reference 1). As an alternative to directly targeting viral proteins, it is conceivable to target host cell metabolic pathways that are required for viral replication. Flaviviruses, small RNA viruses that include DENV as well as West Nile virus (WNV), yellow fever virus, and Japanese encephalitis virus, are highly dependent on the host cell to provide support for viral entry, repl...

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Section: Discussionsupporting

confidence: 57%

Flaviviruses Are Sensitive to Inhibition of Thymidine Synthesis Pathways

Fischer

Smith

Shum

et al. 2013

J Virol

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“…Both nucleoside analogues and non-nucleoside inhibitors of DENV RdRp have recently been reported (25,(32)(33)(34). The crystal structures of WNV and DENV RdRps revealed two conserved cavities that could be used for development of allosteric inhibitors (18).…”

Section: Discussionmentioning

confidence: 99%

Functional Analysis of Two Cavities in Flavivirus NS5 Polymerase

Zou

Chen

Dong

et al. 2011

Journal of Biological Chemistry

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118

128

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Flavivirus NS5 protein encodes methyltransferase and RNAdependent RNA polymerase (RdRp) activities. Structural analysis of flavivirus RdRp domains uncovered two conserved cavities (A and B). Both cavities are located in the thumb subdomains and represent potential targets for development of allosteric inhibitors. In this study, we used dengue virus as a model to analyze the function of the two RdRp cavities. Amino acids from both cavities were subjected to mutagenesis analysis in the context of genome-length RNA and recombinant NS5 protein; residues critical for viral replication were subjected to revertant analysis. For cavity A, we found that only one (Lys-756) of the seven selected amino acids is critical for viral replication. Alanine substitution of Lys-756 did not affect the RdRp activity, suggesting that this residue functions through a nonenzymatic mechanism. For cavity B, all four selected amino acids (Leu-328, Lys-330, Trp-859, and Ile-863) are critical for viral replication. Biochemical and revertant analyses showed that three of the four mutated residues (Leu-328, Trp-859, and Ile-863) function at the step of initiation of RNA synthesis, whereas the fourth residue (Lys-330) functions by interacting with the viral NS3 helicase domain. Collectively, our results have provided direct evidence for the hypothesis that cavity B, but not cavity A, from dengue virus NS5 polymerase could be a target for rational drug design.The genus Flavivirus from the family Flaviviridae contains more than 70 viruses, many of which are important human pathogens causing major public health threats worldwide. Dengue virus (DENV) 2 is a mosquito-borne flavivirus responsible for 50 -100 million human infections and ϳ20,000 deaths each year (1). Besides DENV, West Nile virus (WNV), Japanese encephalitis virus, yellow fever virus, and tick-borne encephalitis virus also cause significant human diseases (1). No antiviral therapy is currently available for treatment of flavivirus infections. Therefore, development of antiviral therapy is urgently needed for flaviviruses.The flavivirus genome is a single strand, plus-sense RNA of about 11 kb in length. The genomic RNA contains a 5Ј-untranslated region (UTR), a single open reading frame, and a 3Ј-UTR. The single open reading frame encodes a long polyprotein that is processed by viral and host proteases into 10 mature viral proteins (2). Three structural proteins (capsid, pre-membrane, and envelope) are primarily involved in virus particle formation, and seven nonstructural proteins (NS1, NS2A, NS2B, NS3, NS4A, NS4B, and NS5) are mainly responsible for viral replication (2). Among these, NS3 and NS5 possess enzymatic activities and have been targeted for antiviral development. NS3 functions as a protease (with NS2B as a cofactor), helicase, 5Ј-RNA triphosphatase, and nucleoside triphosphatase (3-5). NS5 is the largest and the most conserved viral protein. The N-terminal part of NS5 is a methyltransferase that methylates the N-7 and 2Ј-O positions of the viral RNA cap structure (6 -9);...

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“…Nucleoside analogues have been widely used as inhibitors of numerous medically important viruses, including HIV, herpesvirus, and hepatitis B virus (10,11). Regarding flaviviruses, nucleoside analogues have been demonstrated to be efficacious in HCV replicon assays (12)(13)(14)(15)(16) and also against DENV (17)(18)(19)(20)(21), WNV (22), and YFV (23). Some of them were described as broad-spectrum inhibitors of various RNA viruses (24)(25)(26)(27).…”

mentioning

confidence: 99%

Nucleoside Inhibitors of Tick-Borne Encephalitis Virus

Eyer

Valdés

Gil

et al. 2015

Antimicrob Agents Chemother

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The anti-TBEV effect of 2=-CMA in cell culture diminished gradually after day 3 posttreatment. 7-Deaza-2=-CMA showed no detectable cellular toxicity (CC 50 > 50 M), and the antiviral effect in culture was stable for >6 days posttreatment. Computational molecular analyses revealed that compared to the other two compounds, 7-deaza-2=-CMA formed a large cluster near the active site of the TBEV polymerase. High antiviral activity and low cytotoxicity suggest that 7-deaza-2=-CMA is a promising candidate for further investigation as a potential therapeutic agent in treating TBEV infection.

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Inhibition of Dengue Virus by an Ester Prodrug of an Adenosine Analog

Cited by 54 publications

References 20 publications

Flaviviruses Are Sensitive to Inhibition of Thymidine Synthesis Pathways

Flaviviruses Are Sensitive to Inhibition of Thymidine Synthesis Pathways

Functional Analysis of Two Cavities in Flavivirus NS5 Polymerase

Nucleoside Inhibitors of Tick-Borne Encephalitis Virus

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