Hepatitis C Virus RNA-dependent RNA Polymerase (NS5B) as a Mediator of the Antiviral Activity of Ribavirin

Maag, David; Castro, Christian; Hong, Zhi; Cameron, Craig Ε.

doi:10.1074/jbc.c100349200

Cited by 267 publications

(213 citation statements)

References 15 publications

(19 reference statements)

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“…Recently, Cameron and colleagues (21,22) showed that ribavirin can be misincorporated into mRNA by viral RNA-dependent RNA polymerases because of its chemical similarity to guanosine, and this outcome leads to lethal mutagenesis of genomes of polio and HCV. Ribavirin triphosphate (RTP) binds the HCV polymerase with an observed dissociation constant (K d ) of 0.58 mM (23), which is consistent with micromolar concentrations necessary to achieve a therapeutic effect against HCV clinically (24). In addition to effects at millimolar concentrations, ribavirin is also active at micromolar concentrations.…”

mentioning

confidence: 80%

Ribavirin suppresses eIF4E-mediated oncogenic transformation by physical mimicry of the 7-methyl guanosine mRNA cap

Kentsis

Topisirović

Čuljković

et al. 2004

Proc. Natl. Acad. Sci. U.S.A.

271

357

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The eukaryotic translation initiation factor eIF4E is deregulated in many human cancers, and its overexpression in cells leads to malignant transformation. Oncogenic properties of eIF4E are directly linked to its ability to bind 7-methyl guanosine of the 5 mRNA. Here, we observe that the antiviral guanosine analogue ribavirin binds to eIF4E with micromolar affinity at the functional site used by 7-methyl guanosine mRNA cap, competes with eIF4E:mRNA binding, and, at low micromolar concentrations, selectively disrupts eIF4E subcellular organization and transport and translation of mRNAs posttranscriptionally regulated by eIF4E, thereby reducing levels of oncogenes such as cyclin D1. Ribavirin potently suppresses eIF4E-mediated oncogenic transformation of murine cells in vitro, of tumor growth of a mouse model of eIF4E-dependent human squamous cell carcinoma in vivo, and of colony formation of eIF4E-dependent acute myelogenous leukemia cells derived from human patients. These findings describe a specific, potent, and unforeseen mechanism of action of ribavirin. Quantum mechanical and NMR structural studies offer directions for the development of derivatives with improved cytostatic and antiviral properties. In all, ribavirin's association with eIF4E may provide a pharmacologic means for the interruption of posttranscriptional networks of oncogenes that maintain and enhance neoplasia and malignancy in human cancer.cancer ͉ drug ͉ virus ͉ oncogenic network W hereas the precise causes of neoplasia and malignancy are known only for a few human cancers, deregulated tumor suppressors and oncogenes that maintain and enhance the malignant phenotype are becoming relatively well described (1, 2). Among these molecules are tumor suppressors like p53, Rb, and APC and oncogenes such as myc, cyclin D1, and eIF4E. Their interactions constitute a network of self-reinforcing feedback loops, whereby inactivation of principal elements can lead to reversal and at times even sustained loss of neoplastic phenotype (3, 4). eIF4E is overexpressed in a wide variety of malignant cell lines and primary human tumors such as carcinomas of the breast (5), colon (6), and head and neck (7), non-Hodgkin's lymphomas (8), and chronic and acute M4͞M5 myelogenous leukemias (9). Consistently, even moderate overexpression of eIF4E in rodent cells leads to deregulated proliferation and malignant transformation (10). eIF4E is essential for growth and survival of eukaryotes by acting at a critical step of cap-dependent translation and recruiting transcripts to the ribosome as a result of a specific interaction with the 5Ј 7-methyl guanosine (m 7 G) mRNA cap (11). It is important to stress that, although the interaction of eIF4E with the 5Ј mRNA cap is required for initiation of translation of cap-dependent mRNAs, up-regulation of eIF4E does not increase translation of all cap-dependent transcripts, but only of a specific subset of eIF4E-sensitive transcripts, as described below.As much as 70% of eIF4E is present in the nuclei of mammalian cells, where it i...

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mentioning

confidence: 80%

Ribavirin suppresses eIF4E-mediated oncogenic transformation by physical mimicry of the 7-methyl guanosine mRNA cap

Kentsis

Topisirović

Čuljković

et al. 2004

Proc. Natl. Acad. Sci. U.S.A.

271

357

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“…It is probable that this situation will extend to other RdRPs, especially the enzyme from hepatitis C virus (47). Therefore, it is reasonable to target the elongation complex for antiviral drug development.…”

Section: Implications For Development Of Antiviralsmentioning

confidence: 99%

Poliovirus RNA-Dependent RNA Polymerase (3D^p^ol): Pre-Steady-State Kinetic Analysis of Ribonucleotide Incorporation in the Presence of Mg²⁺

2004

Self Cite

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We have solved the complete kinetic mechanism for correct nucleotide incorporation catalyzed by the RNA-dependent RNA polymerase from poliovirus, 3D pol . The phosphoryl-transfer step is flanked by two isomerization steps. The first conformational change may be related to reorientation of the triphosphate moiety of the bound nucleotide, and the second conformational change may be translocation of the enzyme into position for the next round of nucleotide incorporation. The observed rate constant for nucleotide incorporation by 3D pol (86 s -1 ) is dictated by the rate constants for both the first conformational change (300 s -1 ) and phosphoryl transfer (520 s -1 ). Changes in the stability of the "activated" ternary complex correlate best with changes in the observed rate constant for incorporation resulting from modification of the nucleotide. With the exception of UTP, the K d values for nucleotides are at least 10-fold lower than the cellular concentration of the corresponding nucleotide. Our data predict that transition mutations should occur at a frequency of 1/15000, transversion mutations should occur at a frequency of less than 1/150000, and incorporation of a 2′-deoxyribonucleotide with a correct base should occur at a frequency 1/7500. Together, these data support the conclusion that 3D pol is actually as faithful as an exonuclease-deficient, replicative DNA polymerase. We discuss the implications of this work on the development of RNA-dependent RNA polymerase inhibitors for use as antiviral agents.One positive aspect of the human immunodeficiency virus (HIV) 1 pandemic was the realization that, given the appropriate effort, antiviral agents targeting enzymes required for virus multiplication can be developed. The first HIV-encoded enzyme that emerged as a tractable target was the viral polymerase, reverse transcriptase (RT) (1-3). While most of the inhibitors of HIV RT currently in use are nucleoside analogues (1,4), many nonnucleoside analogue inhibitors of HIV RT have also been reported (1,4). In all cases, however, an understanding of the specificity and/or precise mechanism of action of these inhibitors has required evaluation of these compounds in the context of the complete kinetic mechanism for nucleotide incorporation catalyzed by this enzyme (1,4-7) and has benefited from structural information available for HIV RT in the absence and presence of substrates and/or inhibitors (8)(9)(10)(11)(12)(13)(14). Kinetic analysis of site-directed mutants of HIV RT has established some of the fundamental mechanisms employed by this enzyme to recognize its substrates, both nucleic acid and nucleotide, and to cooperate with other viral proteins during the process of converting the viral single-stranded RNA genome into double-stranded DNA (1,4,15,16). In addition, mechanistic studies of HIV RT have suggested alternative strategies to target regions of this enzyme distinct † This work was supported, in part, by a Howard Temin Award (CA75118) from the NCI, National Institutes of Health, and by a grant ...

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“…[66][67][68][69] It is well absorbed from the proximal gut and has a large apparent volume of distribution because of its rapid entry into all cell types, which is mediated by small es-nucleoside transporters. It is not bound to plasma proteins and is eliminated slowly from intracellular and extracellular compartments.…”

Section: Ifn-␣mentioning

confidence: 99%

“…66,68,69 Polarization of a T H 1 response would be expected to enhance both cytokine-and CTL-mediated clearance of HCV-in- …”

Section: Ifn-␣mentioning

confidence: 99%

The clinical and immunologic impact of using interferon and ribavirin in the immunosuppressed host

Braun

Vierling

2003

Liver Transplantation

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C irrhosis caused by chronic hepatitis C virus (HCV) infection is the leading indication for cadaver and living donor liver transplantation. 1 Recurrent HCV infection, which occurs within days in virtually all allografts, is associated with a 10-to 20-fold increase in serum HCV RNA levels because of enhanced replication stimulated by immunosuppression. 2,3 A majority develop acute hepatitis C 1 to 4 months post-orthotopic liver transplantation (OLT) that is characterized by lymphocytic infiltration and hepatocyte apoptosis. Progression to chronic hepatitis C is associated with: (1) decline in HCV RNA levels observed in the acute phase because of effects of HCVspecific CD4 T helper subtype 1 (T H 1) and CD8 cell responses, (2) profibrotic effects of interferon-␥ (IFN-␥), and (3) proapoptotic effects of increased hepatocyte expression of Fas. 3 Progression of chronic hepatitis C is accelerated in immunosuppressed patients post OLT, resulting in a cumulative probability of cirrhosis in 30% by 5 years. 4 In addition, a minority develop severe, usually fatal, cholestatic hepatitis C within the first 6 months after OLT that is characterized by a direct cytopathic effect of excessive intrahepatic replication of HVC, intrahepatic anti-inflammatory T H 2 cytokine expression, and absence of an HCV-specific immune response. 3 Studies of the impact of different immunosuppressive regimens on recurrent hepatitis C also have been performed in an attempt to reduce hepatitis C progression while preventing acute cellular rejection (ACR). 5 Overall, HCV infection significantly reduces graft and patient survival compared with patients who undergo OLT for non-HCV-related indications. 6 Results of re-OLT are poor, 7 and this is an increasingly unrealistic option in view of the shortage of organ donors.To prevent morbidity and mortality associated with recurrent hepatitis C, many centers have used interferon alfa (IFN-␣) and/or ribavirin (RVN) as either antiviral prophylaxis or treatment of established hepatitis C or hepatitis B. Regimens have ranged from

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Hepatitis C Virus RNA-dependent RNA Polymerase (NS5B) as a Mediator of the Antiviral Activity of Ribavirin

Cited by 267 publications

References 15 publications

Ribavirin suppresses eIF4E-mediated oncogenic transformation by physical mimicry of the 7-methyl guanosine mRNA cap

Ribavirin suppresses eIF4E-mediated oncogenic transformation by physical mimicry of the 7-methyl guanosine mRNA cap

Poliovirus RNA-Dependent RNA Polymerase (3D^p^ol): Pre-Steady-State Kinetic Analysis of Ribonucleotide Incorporation in the Presence of Mg²⁺

The clinical and immunologic impact of using interferon and ribavirin in the immunosuppressed host

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Hepatitis C Virus RNA-dependent RNA Polymerase (NS5B) as a Mediator of the Antiviral Activity of Ribavirin

Cited by 267 publications

References 15 publications

Ribavirin suppresses eIF4E-mediated oncogenic transformation by physical mimicry of the 7-methyl guanosine mRNA cap

Ribavirin suppresses eIF4E-mediated oncogenic transformation by physical mimicry of the 7-methyl guanosine mRNA cap

Poliovirus RNA-Dependent RNA Polymerase (3Dpol): Pre-Steady-State Kinetic Analysis of Ribonucleotide Incorporation in the Presence of Mg2+

The clinical and immunologic impact of using interferon and ribavirin in the immunosuppressed host

Contact Info

Product

Resources

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Poliovirus RNA-Dependent RNA Polymerase (3D^p^ol): Pre-Steady-State Kinetic Analysis of Ribonucleotide Incorporation in the Presence of Mg²⁺