Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:2.7.7.7 (DNA polymerase)
 17,007 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The human ISG20/HEM45 gene was identified independently on the basis of its increased level of expression in response to either interferon or estrogen hormone. Notably, the encoded protein is homologous with members of the 3' to 5' exonuclease superfamily that includes RNases T and D, and the proofreading domain of Escherichia coli DNA polymerase I. We provide here direct biochemical evidence that Isg20 acts as a 3' to 5' exonuclease in vitro. This protein displays a pH optimum of approximately 7.0, prefers Mn2+ as a metal cofactor, and degrades RNA at a rate that is approximately 35-fold higher than its rate for single-stranded DNA. Along with RNase L, Isg20 is the second known RNase regulated by interferon. Previous data showed that Isg20 is located in promyelocytic leukemia (PML) nuclear bodies, known sites of hormone-dependent RNA polymerase II transcription and oncogenic DNA viral transcription and replication. The combined data suggest a potential role for Isg20 in degrading viral RNAs as part of the interferon-regulated antiviral response and/or cellular mRNAs as a regulatory component of interferon and estrogen signaling.
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PMID:The human interferon- and estrogen-regulated ISG20/HEM45 gene product degrades single-stranded RNA and DNA in vitro. 1140 64

The term 'genotyping' describes the genetic characterization of a genome. The genotype analysis is performed to identify mutations that differentiate one individual or strain from another. The mutations may confer resistance to specific antiviral drugs or they may simply allow classification of a strain as to 'type' and 'subtype'. There are four human viruses for which genotype information is clinically useful. Hepatitis B virus (HBV) infections are being treated with antiretroviral drugs and resistance after prolonged treatment is common. Since HBV cannot be cultured, the only method of detecting resistance-conferring mutations in the genome is a genotypic analysis. Hepatitis C virus (HCV) infection can be cured by treatment with the combination of interferon and ribavirin but certain strains of virus are more resistant to treatment than others. The current recommendations are that all HCV type 1 infections be treated for 12 months whereas other types may be successfully treated in 6 months. Since interferon treatment may have significant side effects, the determination of HCV genotype is an important aspect of this therapeutic regimen. Treatment of cytomegalovirus (CMV) disease with nucleoside analogues occasionally results in resistant virus with mutations in the phosphotransferase gene (UL97) and/or the DNA polymerase gene (UL54) that can be tested with phenotypic or genotypic assays. Since CMV grows very slowly, it may be more clinically useful to perform a rapid genotypic assay although only the UL97 gene can be efficiently genotyped. Finally, the virus for which genotyping has become the standard of care, human immunodeficiency virus type 1 (HIV-1) can now be genotyped routinely by many clinical virology labs experienced with molecular amplification methods and automated DNA sequencing technology. All currently-available antiretroviral drugs are directed against either the protease or reverse transcriptase genes of HIV-1 and the mutations within these genes that confer resistance have been well described. Sequence-based genotyping methods are not necessarily the best approach for routine genotyping of these four viruses, but sequencing is the gold standard from which other methods are developed and against which they are compared.
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PMID:Clinically relevant sequence-based genotyping of HBV, HCV, CMV, and HIV. 1141 49

Interferon-alpha is the most widely used antiviral drug in chronic hepatitis B and C. Tolerability is usually good and serious adverse effects are rare. Most of the adverse effects are mild or transient and do not necessitate drug withdrawal. More than 90% of patients who are given interferon-alpha achieve 6 months to 1 year of treatment without serious adverse effects. The serious adverse effects usually occur in predisposed patients with pre-existing organ dysfunction. Nevertheless, careful selection of patients for therapy and observation during therapy are recommended. Nucleoside analogues are promising drugs in the treatment of chronic hepatitis B through inhibition of viral DNA polymerase. Lamivudine has been licensed for use in this indication. Its tolerability is excellent even when used for periods of 1 year or more. The main concern is the relatively high incidence of viral resistance resulting in breakthrough during or relapse after therapy. In the treatment of chronic hepatitis C, ribavirin, in combination with interferon-alpha is currently the reference therapy. The main adverse effect is haemolytic anaemia, which necessitates careful monitoring and adjustment of dosage in many cases. Recently, large trials showed the better efficacy of pegylated interferons as compared with standard interferon. The combination of pegylated interferon with ribavirin is under evaluation.
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PMID:Tolerability of treatments for viral hepatitis. 1141 64

Development of a novel group of antiviral agents, acyclic nucleoside phosphonates, has provided a new perspective for treating human immunodeficiency virus (HIV) infection. One of the compounds, 9-(R)-[2-(phosphonomethoxy)propyl]adenine (PMPA) (tenofovir), has been shown to confer complete protection against AIDS in a simian model of the infection. The aim of our study was to investigate whether the antiviral efficacy of PMPA, which depends mainly on inhibition of virus-induced DNA polymerase or of reverse transcriptase, could be contributed by immunomodulatory potential of this drug. We screened for its ability to activate production of cytokines and chemokines that are known to interfere with the replication and/or the entry of HIV in cells. Using the in vitro test system of mouse macrophages and lymphocytes, it has been found that PMPA stimulates macrophage secretion of interleukin-1beta (IL-1beta), IL-10, and tumor necrosis factor alpha. Production of the chemokines RANTES and macrophage inflammatory protein 1alpha was activated in both macrophages and lymphocytes, and also in human cell line U937. Other cytokines--i.e., IL-2, IL-12, IL-13, and gamma interferon-remained uninfluenced by PMPA. The cytokines were stimulated in a dose-dependent fashion, with rapid onset, and peak concentrations were achieved within 5 to 24 h. The findings contribute to a more complex understanding of mechanisms of antiviral effectiveness of PMPA and support the view that this drug could become a promising candidate for therapeutic exploitation in anti-HIV preventive medicine.
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PMID:Activation by 9-(R)-[2-(phosphonomethoxy)propyl]adenine of chemokine (RANTES, macrophage inflammatory protein 1alpha) and cytokine (tumor necrosis factor alpha, interleukin-10 [IL-10], IL-1beta) production. 1170 12

Lamivudine (3TC) is a nucleoside analogue which inhibits replication of HIV and HBV and which is used in the treatment of chronic hepatitis B-infected patients with safety and efficacy. The activity of lamivudine was evaluated by the measurement of DNA-HBV concentration in plasma using a very sensitive assay (1,000 copies/mL) (Amplicor VHB Monitor. Roche). Ten patients chronically infected with hepatitis B (group A) and 24 patients with HIV-1 co-infection (group B) were enrolled. In 9 patients of group A, HBVDNA load was undetectable a median of 3.5 months after the beginning of treatment and remained negative for 2 years with hepatitis Be antigen disappearing and normal alanine aminotransferase concentration. In the last immunodeficient patient, the virus which had been resistant to three interferon treatments, was also resistant to lamivudine. In five patients of group B, HBV DNA load remained undetectable after 18 months with HBe antigen disappearing and baseline concentration of alanine aminotransferase. In the remaining 19 patients after a transient decrease of HBV DNA concentration for one year, HBV DNA load increased again without disappearing of HBe antigen and without decrease of alanine aminotransferase concentration showing lamivudine resistant hepatitis B virus. Mutations in the YMDD motif of the DNA polymerase gene were identified in 11 patients (3 with M550V/I mutation; 7 with M550V/I and L256M mutations; 1 with M550V/I, L526M and V519L mutations). In 6 of these patients, was found a M184V mutation in the VIH polymerase. No correlation could be observed between the mutations detected in the two viruses. Using a sensitive HBV-DNA assay, efficacy of lamivudine for a long time in HBV infected patients was proved. However, the prevalence of lamivudine resistance is related to duration of treatment and it may be necessary to use a multitherapy.
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PMID:[Evaluation of a quantitative HBV-DNA PCR assay in lamivudine treated hepatitis B-infected patients]. 1236 44

The emergence of drug-resistant virus in hepatitis B virus (HBV) patients treated with lamivudine is well documented. In this study, we determined the mutations occurring in the tyrosine-methionine-aspartate-aspartate (YMDD) amino acid motif of the HBV DNA polymerase gene, as well as upstream and downstream of this region, in patients with breakthrough virus during lamivudine therapy. Thirty-one Turkish patients (20 patients HBeAg positive, 11 patients HBeAg negative and anti-HBe positive) with chronic HBV infection who completed at least 104 weeks of lamivudine treatment were investigated. All patients received lamivudine, (150 mg/day), for 104 weeks, with or without 4 months of interferon (IFN) combination. HBV-specific sequences were amplified by polymerase chain reaction (PCR) from sera of patients with breakthrough virus, and the PCR products were directly analysed by sequencing. Breakthrough virus was detected in seven of the 31 patients (22.6%) between 9 and 18 months of therapy. Of the seven patients, six were HBeAg positive at baseline, and four had a double mutation consisting of rtM204V and rtL180M, while two had an rtM204I change. In one patient, two base substitutions at rt204 (ATG --> AGT; T to G and G to T) lead to a methionine to serine change (YMDD --> YSDD). This novel DNA pol mutation was detected at month 18 of lamivudine treatment. In addition, this new variant had the rtL180M mutation and a 12 base pair deletion in the pre-S1 region between nucleotides 43-54. The YSDD mutation was still present 6 months after lamivudine discontinuation. In vitro transfection studies also confirmed that the YSDD strain is resistant to lamivudine. In conclusion, the results indicate that, in addition to a Met --> Val and Met --> Ile change in YMDD, a Met --> Ser change at rt204 (YMDD --> YSDD) associated with the rtL180M change can also emerge during lamivudine treatment, which confers lamivudine resistance in vivo and in vitro, leading to virological breakthrough and ALT increases.
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PMID:YSDD: a novel mutation in HBV DNA polymerase confers clinical resistance to lamivudine. 1282 91

n-(n-Nonyl)-deoxygalactonojirimycin (n,n-DGJ), an alkylated imino sugar, reduces the amount of HBV DNA produced within the stably transfected HBV-producing HepG2.2.15 line in culture and is under consideration for development as a human therapeutic. n,n-DGJ does not appear to inhibit HBV DNA polymerase activity or envelop antigen production (A. Mehta, S. Carrouee, B. Conyers, R. Jordan, T. Butters, R. A. Dwek, and T. M. Block, Hepatology 33:1488-1495, 2001), and the mechanism of antiviral action is unknown. In this study, the step in the virus life cycle affected by n,n-DGJ was explored. Using Northern analysis and immunoprecipitation with anti-HBc antibody, we found that, under conditions in which cell viability was not affected but viral DNA production was substantially reduced, neither the amount of HBV transcription products nor the core polypeptide was detectably reduced. However, the pregenomic RNA, endogenous polymerase activity, and core polypeptide sedimenting in sucrose gradients with a density consistent with that of assembled nucleocapsids were significantly less in the HepG2.2.15 cells incubated with n,n-DGJ. These data suggest that n,n-DGJ either prevents the maturation of HBV nucleocapsids or destabilizes the formed nucleocapsids. Although the cellular and viral mediators of this inhibition are not known, depletion of nucleocapsid has been attributed to some other compounds as well as interferon's mechanism of anti-HBV action. The similarities and differences between this alkylated imino sugar and these other mediators are discussed.
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PMID:The alkylated imino sugar, n-(n-Nonyl)-deoxygalactonojirimycin, reduces the amount of hepatitis B virus nucleocapsid in tissue culture. 1458 30

Breakthroughs during lamivudine therapy were assessed according to hepatitis flares and mutational polymorphism of hepatitis B virus (HBV) infecting patients. Of 42 patients with chronic hepatitis B and positive for hepatitis B e antigen in serum, 13 (30%) harbored HBV mutants with lamivudine resistance after a mean duration of 29 months on lamivudine. The virological breakthrough occurred 14.5 months after the start of lamivudine treatment, and all the patients with it developed breakthrough hepatitis 3 months later. The clinical course of breakthrough hepatitis was self-limited except in one patient who had already developed cirrhosis at the baseline. One year after breakthrough hepatitis, serum ALT, albumin, prothrombin time and platelet counts were maintained well on conventional treatments without resorting to interferon. Major HBV mutants during breakthrough hepatitis were those with M552I in the YMDD motif of viral DNA polymerase/reverse transcriptase in 7 patients (54%), M552I/L528M in 4 patients (31%) and M552V/L528M in 2 patients (15%). There were no patients in whom mutations at nucleotide 529 occurred including the 2 who later developed hepatocellular carcinoma. There was no clear relationship between distinct mutational patterns and clinical courses. Further studies are needed for making out the effects of lamivudine-resistant mutants on clinical outcomes, taking into considerations genotypes of HBV.
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PMID:Subclones of drug-resistant hepatitis B virus mutants and the outcome of breakthrough hepatitis in patients treated with lamivudine. 1468 51

Methionine deprivation imposes a metabolic stress, termed methionine stress, that inhibits mitosis and induces cell cycle arrest and apoptosis. The methionine-dependent central nervous system tumor cell lines DAOY (medulloblastoma), SWB61 (anaplastic oligodendroglioma), SWB40 (anaplastic astrocytoma), and SWB39 (glioblastoma multiforme) were compared with methionine-stress resistant SWB77 (glioblastoma multiforme). The cDNA-oligoarray analysis and reverse transcription-PCR verification indicated common changes in gene expression in methionine-dependent cell lines to include up-regulation/induction of cyclin D1, mitotic arrest deficient (MAD)1, p21, growth arrest and DNA-damage-inducible (GADD)45 alpha, GADD45 gamma, GADD34, breast cancer (BRCA)1, 14-3-3sigma, B-cell CLL/lymphoma (BCL)1, transforming growth factor (TGF)-beta, TGF-beta-induced early response (TIEG), SMAD5, SMAD7, SMAD2, insulin-like growth factor binding protein (IGFBP7), IGF-R2, vascular endothelial growth factor (VEGF), TNF-related apoptosis-inducing ligand (TRAIL), TNF-alpha converting enzyme (TACE), TRAIL receptor (TRAIL-R)2, TNFR-related death receptor (DR)6, TRAF interacting protein (I-TRAF), IL-6, MDA7, IL-1B convertase (ICE)-gamma, delta and epsilon, IRF1, IRF5, IRF7, interferon (IFN)-gamma and receptor components, ISG15, p65-NF-kappaB, JUN-B, positive cofactor (PC)4, C/ERB-beta, inositol triphosphate receptor I, and methionine adenosyltransferase II. On the other hand, cyclins A1, A2, B1 and B2, cell division cycle (CDC)2 and its kinase, CDC25 A and B, budding uninhibited by benzimidazoles (BUB)1 and 3, MAD2, CDC28 protein kinase (CKS)1 and 2, neuroepithelial cell transforming gene (NET)1, activator of S-phase kinase (ASK), CDC14B phosphatase, BCL2, TGF-beta activated kinase (TAK)1, TAB1, c-FOS, DNA topoisomerase II, DNA polymerase alpha, dihydrofolate reductase, thymidine kinase, stathmin, and MAP4 were down-regulated. In the methionine stress-resistant SWB77, only 20% of the above genes were affected, and then only to a lesser extent. In addition, some of the changes observed in SWB77 were opposite to those seen in methionine-dependent tumors, including expression of p21, TRAIL-R2, and TIEG. Despite similarities, differences between methionine-dependent tumors were substantial, especially in regard to regulation of cytokine expression. Western blot analysis confirmed that methionine stress caused the following: (a) a marked increase of GADD45alpha and gamma in the wt-p53 cell lines SWB61 and 40; (b) an increase in GADD34 and p21 protein in all of the methionine-dependent lines; and (c) the induction of MDA7 and phospho-p38 in DAOY and SWB39, consistent with marked transcriptional activation of the former under methionine stress. It was additionally shown that methionine stress down-regulated the highly active phosphatidylinositol 3'-kinase pathway by reducing AKT phosphorylation, especially in DAOY and SWB77, and also reduced the levels of retinoblastoma (Rb) and pRb (P-ser780, P-ser795, and P-ser807/811), resulting in a shift in favor of unphosphorylated species in all of the methionine-dependent lines. Immunohistochemical analysis showed marked inhibition of nuclear translocation of nuclear factor kappaB under methionine stress in methionine-dependent lines. In this study we show for the first time that methionine stress mobilizes several defined cell cycle checkpoints and proapoptotic pathways while coordinately inhibiting prosurvival mechanisms in central nervous system tumors. It is clear that methionine stress-induced cytotoxicity is not restricted by the p53 mutational status.
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PMID:Modulation of gene expression in human central nervous system tumors under methionine deprivation-induced stress. 1549 78

ISG20 is an interferon-induced antiviral exoribonuclease that acts on single-stranded RNA and also has minor activity towards single-stranded DNA. It belongs to the DEDDh group of RNases of the DEDD exonuclease superfamily. We have solved the crystal structure of human ISG20 complexed with two Mn2+ ions and uridine 5'-monophosphate (UMP) at 1.9 A resolution. Its structure, including that of the active site, is very similar to those of the corresponding domains of two DEDDh-group DNases, the epsilon subunit of Escherichia coli DNA polymerase III and E. coli exonuclease I, strongly suggesting that its catalytic mechanism is identical to that of the two DNases. However, ISG20 also has distinctive residues, Met14 and Arg53, to accommodate hydrogen bonds with the 2'-OH group of the UMP ribose, and these residues may be responsible for the preference of ISG20 for RNA substrates.
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PMID:Crystal structure of human ISG20, an interferon-induced antiviral ribonuclease. 1552 70


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