Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:2.7.7.49 (reverse transcriptase)
31,746 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Abacavir (1592U89), a nucleoside reverse transcriptase inhibitor with in vitro activity against human immunodeficiency virus type-1 (HIV-1), has been evaluated for efficacy and safety in combination regimens with other nucleoside analogs, including zidovudine (ZDV) and lamivudine (3TC). To evaluate the potential pharmacokinetic interactions between these agents, 15 HIV-1-infected adults with a median CD4(+) cell count of 347 cells/mm3 (range, 238 to 570 cells/mm3) were enrolled in a randomized, seven-period crossover study. The pharmacokinetics and safety of single doses of abacavir (600 mg), ZDV (300 mg), and 3TC (150 mg) were evaluated when each drug was given alone or when any two or three drugs were given concurrently. The concentrations of all drugs in plasma and the concentrations of ZDV and its 5'-glucuronide metabolite, GZDV, in urine were measured for up to 24 h postdosing, and pharmacokinetic parameter values were calculated by noncompartmental methods. The maximum drug concentration (Cmax), the area under the concentration-time curve from time zero to infinity (AUC0-infinity), time to Cmax (Tmax), and apparent elimination half-life (t1/2) of abacavir in plasma were unaffected by coadministration with ZDV and/or 3TC. Coadministration of abacavir with ZDV (with or without 3TC) decreased the mean Cmax of ZDV by approximately 20% (from 1.5 to 1.2 microg/ml), delayed the median Tmax for ZDV by 0.5 h, increased the mean AUC0-infinity for GZDV by up to 40% (from 11.8 to 16.5 microg. h/ml), and delayed the median Tmax for GZDV by approximately 0.5 h. Coadministration of abacavir with 3TC (with or without ZDV) decreased the mean AUC0-infinity for 3TC by approximately 15% (from 5.1 to 4.3 microg. h/ml), decreased the mean Cmax by approximately 35% (from 1.4 to 0.9 microg/ml), and delayed the median Tmax by approximately 1 h. While these changes were statistically significant, they are similar to the effect of food intake (for ZDV) or affect an inactive metabolite (for GZDV) or are relatively minor (for 3TC) and are therefore not considered to be clinically significant. No significant differences were found in the urinary recoveries of ZDV or GZDV when ZDV was coadministered with abacavir. There was no pharmacokinetic interaction between ZDV and 3TC. Mild to moderate headache, nausea, lymphadenopathy, hematuria, musculoskeletal chest pain, neck stiffness, and fever were the most common adverse events reported by those who received abacavir. Coadministration of ZDV or 3TC with abacavir did not alter this adverse event profile. The three-drug regimen was primarily associated with gastrointestinal events. In conclusion, no clinically significant pharmacokinetic interactions occurred between abacavir, ZDV, and 3TC in HIV-1-infected adults. Coadministration of abacavir with ZDV or 3TC produced mild changes in the absorption and possibly the urinary excretion characteristics of ZDV-GZDV and 3TC that were not considered to be clinically significant. Coadministration of abacavir with ZDV and/or 3TC was generally well tolerated and did not produce unexpected adverse events.
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PMID:Single-dose pharmacokinetics and safety of abacavir (1592U89), zidovudine, and lamivudine administered alone and in combination in adults with human immunodeficiency virus infection. 1039 Feb 27

Several new nucleoside analogues have been developed which can inhibit hepatitis B replication by at least two logs. Lamivudine is the most widely studied of these new agents. Extensive phase II and III studies in patients with chronic hepatitis B have been completed. The sustained HBeAg seroconversion rate in patients who have received 100 mg lamivudine increases from 17% after a year of treatment to 27% after 2 years of treatment. Histological improvement has been noted in 38%-52% of lamivudine-treated patients, exceeding the improvement seen in placebo recipients. Similar histological improvement has been noted in anti-HBe-positive, DNA- positive patients. Lamivudine can prevent recurrence of hepatitis B after liver transplantation. It is likely that in the absence of immune clearance to accelerate elimination of infected hepatocytes, inhibitors of virus replication such as lamivudine will need to be administered for a long period to reduce the burden of infected hepatocytes in the liver, and to prevent relapse. The drug is generally well tolerated with few direct adverse events. Genotypic mutations have been observed in 23% (range 13-32%). In a study in Asian patients treated for two years the incidence of these mutants increased to 38% (as detected by PCR). Loss of susceptibility to lamivudine has been found to be due to reverse transcriptase amino acid substitutions. Lamivudine is likely to be reserved for patients with replicative hepatitis B infection with active chronic hepatitis, and/or active cirrhosis. Copyright 1998 John Wiley & Sons, Ltd.
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PMID:Lamivudine treatment of chronic hepatitis B. 1039 3

A single amino acid substitution from methionine-184 to valine (M184V) of HIV-1 reverse transcriptase (RT) evokes the 1000-fold 3TC (Lamivudine) resistance by the HIV-1 virus observed in the clinic. The M184V mutant HIV-1 RT was studied to assess its catalytic efficiency during single nucleotide incorporation using a transient kinetic approach. The maximum rate of polymerization (k(pol)), binding affinity (K(d)), and incorporation efficiency (k(pol)/K(d)) were determined for incorporating dCTP and 3TC-TP by wild-type and 3TC-resistant HIV-1 RT. The 3TC-resistant HIV-1 RT showed a similar efficiency of incorporation compared with the wild-type enzyme during DNA-dependent DNA polymerization; however, the incorporation efficiency is reduced 3.5-fold during RNA-dependent polymerization. A dramatic 146- and 117-fold decrease in incorporation efficiency was observed for 3TC-MP incorporation by M184V RT for DNA- and RNA-dependent DNA polymerization, respectively, as compared with wild-type HIV-1 RT. While the k(pol) was slower and the K(d) was weaker for 3TC-TP incorporation by the M184V RT, the decrease in the efficiency of incorporation is primarily due to a substantially reduced binding affinity for the 3TC-TP to the enzyme.DNA (or RNA) complex poised for DNA elongation. The fidelity of M184V RT was also examined to evaluate mispair formation since this mutant has been suggested to exhibit a higher level of fidelity. The results of our studies indicate that there is a maximum 2.4-fold increase in fidelity for M184V RT as compared with wild-type HIV-1 RT. Both the wild-type and 3TC-resistant mutant RT showed higher fidelity using an RNA template as contrasted with the corresponding DNA template. This mechanistic information provides insight into our understanding of the molecular mechanism of 3TC-drug resistance and supports suggestions that increased RT fidelity and decreased fitness of the M184V HIV-1 virus may be factors contributing to the strong antiviral effect of AZT-3TC combination therapy.
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PMID:Mechanistic studies examining the efficiency and fidelity of DNA synthesis by the 3TC-resistant mutant (184V) of HIV-1 reverse transcriptase. 1041 20

To analyze the emergence and role of the lamivudine (3TC)-selected HIV-1 reverse transcriptase (RT) M184V mutation under triple therapy, we performed a retrospective study of 40 nucleoside RT inhibitor-pretreated and 16 drug-naive patients who were switched to combined treatment with zidovudine (ZDV) plus 3TC plus a protease inhibitor (PI). Plasma viral load and pol genotype were analyzed at baseline and after 24 and 48 weeks of combination therapy. Emergence of the M184V RT mutation at week 48 was detected in 3 of 16 (18.7%) initially drug-naive subjects as opposed to 21 of 40 (52.5%) ZDV-pretreated patients. Multivariate logistic analysis detected HIV-1 RNA load at week 24 as the best predictor of subsequent selection of the M184V mutant (p = .0121). Among ZDV-resistant study subjects at week 24 (n = 17), those with mutant RT M184V codon had a more favorable HIV-1 RNA slope than those with wild-type RT 184M codon (p = .0551). This trend was observed, although in a less evident manner, even in pretreated ZDV-sensitive patients. These findings suggest that development of the 3TC-resistance M184V mutation under triple therapy with 3TC, ZDV, and a PI may have unexpected beneficial effects in vivo in addition to those associated with resensitization of ZDV-resistant virus to ZDV.
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PMID:Development and significance of the HIV-1 reverse transcriptase M184V mutation during combination therapy with lamivudine, zidovudine, and protease inhibitors. 1042 Dec 43

The racemic nucleoside analogue 2'-deoxy-3'-oxa-4'-thiocytidine (dOTC) is in clinical development for the treatment of human immunodeficiency virus (HIV) type 1 (HIV-1) infection. dOTC is structurally related to lamivudine (3TC), but the oxygen and sulfur in the furanosyl ring are transposed. Intracellular metabolism studies showed that dOTC is phosphorylated within cells via the deoxycytidine kinase pathway and that approximately 2 to 5% of dOTC is converted into the racemic triphosphate derivatives, which had measurable half-lives (2 to 3 hours) within cells. Both 5'-triphosphate (TP) derivatives of dOTC were more potent than 3TC-TP at inhibiting HIV-1 reverse transcriptase (RT) in vitro. The K(i) values for dOTC-TP obtained against human DNA polymerases alpha, beta, and gamma were 5,000-, 78-, and 571-fold greater, respectively, than those for HIV RT (28 nM), indicating a good selectivity for the viral enzyme. In culture experiments, dOTC is a potent inhibitor of primary isolates of HIV-1, which were obtained from antiretroviral drug-naive patients as well as from nucleoside therapy-experienced (3TC- and/or zidovudine [AZT]-treated) patients. The mean 50% inhibitory concentration of dOTC for drug-naive isolates was 1.76 microM, rising to only 2.53 and 2.5 microM for viruses resistant to 3TC and viruses resistant to 3TC and AZT, respectively. This minimal change in activity is in contrast to the more dramatic changes observed when 3TC or AZT was evaluated against these same viral isolates. In tissue culture studies, the 50% toxicity levels for dOTC, which were determined by using [(3)H]thymidine uptake as a measure of logarithmic-phase cell proliferation, was greater than 100 microM for all cell lines tested. In addition, after 14 days of continuous culture, at concentrations up to 10 microM, no measurable toxic effect on HepG2 cells or mitochondrial DNA replication within these cells was observed. When administered orally to rats, dOTC was well absorbed, with a bioavailability of approximately 77%, with a high proportion (approximately 16.5% of the levels in serum) found in the cerebrospinal fluid.
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PMID:Anti-human immunodeficiency virus type 1 activity, intracellular metabolism, and pharmacokinetic evaluation of 2'-deoxy-3'-oxa-4'-thiocytidine. 1042

2'-Fluoro-2'3'-dideoxyarabinosyladenine (F-ddA), a nucleoside reverse transcriptase inhibitor of human immunodeficiency virus (HIV) replication, is currently being evaluated in clinical trials. Future monotherapy for the treatment of HIV is unlikely owing to the rapid emergence of drug-resistant viruses, so F-ddA was evaluated in combination with a variety of mechanistically diverse inhibitors of HIV replication. Such in vitro studies provide insights into whether certain drug combinations yield synergistic antiviral activity or, more importantly, antagonistic antiviral activity or synergistic cytotoxicity. F-ddA exhibited synergistic antiviral interactions with representatives of each of the major classes of anti-HIV compounds, including other nucleoside reverse transcriptase inhibitors, non-nucleoside reverse transcriptase inhibitors and protease inhibitors. Greatest levels of synergistic interaction were detected when F-ddA was used in combination with the non-nucleoside compounds nevirapine and costatolide, the nucleoside analogues and costatolide, the nucleoside analogues AZT, ddC and 3TC and the protease inhibitors ritonavir and nelfinavir. No evidence of either combination toxicity or antagonistic antiviral activity was detected with any of the tested compounds.
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PMID:Anti-human immunodeficiency virus type 1 (HIV-1) activity of 2'-fluoro-2',3'-dideoxyarabinosyladenine (F-ddA) used in combination with other mechanistically diverse inhibitors of HIV-1 replication. 1043 10

Of all of the nucleoside inhibitors approved by the FDA for treatment of AIDS, (-)-beta-2',3'-dideoxy-3'-thiacytidine (3TC, lamivudine) is the only one with the unnatural (-)-beta-L configuration. The fluorinated derivative (-)-beta-2', 3'-dideoxy-5-fluoro-3'-thiacytidine [(-)-FTC] and its triphosphate form have also been reported to have excellent antiretroviral activity against HIV-1 reverse transcriptase (RT). Preliminary results of clinical trials suggest that (-)-FTC is 6- to 10-fold more potent than 3TC. However, the molecular mechanism for the observed enhanced clinical potency of (-)-FTC to inhibit viral replication is not understood. The present mechanistic studies used a transient kinetic approach and were designed to compare the incorporation of 3TC-TP and (-)-FTC-TP into DNA by HIV-1 RT and illuminate key features that may play a role in the differential potency. Here we show that (-)-FTC-TP is incorporated 10-fold more efficiently than 3TC-TP during HIV-1 RT-catalyzed RNA-dependent DNA synthesis. The enhanced incorporation efficiency of (-)-FTC-TP may be a key mechanistic feature that, in part, is responsible for the enhanced potency of (-)-FTC observed in ongoing clinical trials.
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PMID:Mechanistic studies show that (-)-FTC-TP is a better inhibitor of HIV-1 reverse transcriptase than 3TC-TP. 1046 41

An important component of triple-drug anti-AIDS therapy is 2', 3'-dideoxy-3'-thiacytidine (3TC, lamivudine). Single mutations at residue 184 of the reverse transcriptase (RT) in HIV cause high-level resistance to 3TC and contribute to the failure of anti-AIDS combination therapy. We have determined crystal structures of the 3TC-resistant mutant HIV-1 RT (M184I) in both the presence and absence of a DNA/DNA template-primer. In the absence of a DNA substrate, the wild-type and mutant structures are very similar. However, comparison of crystal structures of M184I mutant and wild-type HIV-1 RT with and without DNA reveals repositioning of the template-primer in the M184I/DNA binary complex and other smaller changes in residues in the dNTP-binding site. On the basis of these structural results, we developed a model that explains the ability of the 3TC-resistant mutant M184I to incorporate dNTPs but not the nucleotide analog 3TCTP. In this model, steric hindrance is expected for NRTIs with beta- or L- ring configurations, as with the enantiomer of 3TC that is used in therapy. Steric conflict between the oxathiolane ring of 3TCTP and the side chain of beta-branched amino acids (Val, Ile, Thr) at position 184 perturbs inhibitor binding, leading to a reduction in incorporation of the analog. The model can also explain the 3TC resistance of analogous hepatitis B polymerase mutants. Repositioning of the template-primer as observed in the binary complex (M184I/DNA) may also occur in the catalytic ternary complex (M184I/DNA/3TCTP) and contribute to 3TC resistance by interfering with the formation of a catalytically competent closed complex.
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PMID:Lamivudine (3TC) resistance in HIV-1 reverse transcriptase involves steric hindrance with beta-branched amino acids. 1046 56

Lamivudine is effective in suppressing replication of hepatitis B virus (HBV). However, the emergence of HBV variants resistant to lamivudine is a concern. Lamivudine resistance has been attributed mainly to a substitution of isoleucine or valine for methionine at residue 550 (M550I or M550V) in the catalytic site of the virus polymerase. A substitution of methionine for leucine at residue 526 (L526M) has also been identified. To examine such virus genotypic mutations in Japanese patients, we studied five patients with chronic hepatitis B, who showed HBV breakthrough while on a 1-year lamivudine treatment. The entire nucleotide and deduced amino acid sequences of the proposed reverse transcriptase domain of the polymerase gene were determined on HBV DNA amplified by polymerase chain reaction from patient sera collected at the start and at the end of therapy. The HBV sequences from all five patients were of genotype C. In four patients, a substitution of valine or isoleucine for leucine at residue 426, which has not been reported previously, emerged in combination with M550I. One also harbored L526M. In the remaining patient, an alteration of leucine to methionine at residue 428 co-occurred with M550V. Longitudinal study of the mutations showed that the two or three mutations in each patient emerged almost simultaneously 4 weeks before or at the time of breakthrough and were replaced by wild-type virus after completing the therapy. Our results indicate that occurrence of HBV polymerase mutations at residue 426 in combination with M550I is frequent in Japanese or genotype C virus-in- fected patients who develop resistance to lamivudine.
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PMID:Novel patterns of amino acid mutations in the hepatitis B virus polymerase in association with resistance to lamivudine therapy in japanese patients with chronic hepatitis B. 1050 55

Hepatitis B virus (HBV), although a DNA virus, replicates using reverse transcriptase encoded by the HBV polymerase (pol) gene. The biochemical dissection of HBV pol has been hampered by failure to liberate enzymatically active protein from nucleocapsids. Here, we have employed a yeast-based genetic approach to express the HBV reverse transcriptase. In this strategy, the reverse transcriptase of yeast retrotransposon Ty1 element is replaced with the HBV pol gene to produce the hybrid Ty1/HBV element. Additionally, the indicator gene his3AI is combined in an antisense orientation to the transcripts of the hybrid Ty1/HBVRT element. The splicing of his3AI, cDNA synthesis of the Ty1/HBVRT RNA and subsequent integration relies on the reverse transcriptase activity. The production of histidine prototrophs results from the successful reverse transcription of Ty1/HBVRThis3AI transcripts followed by either homologous recombination or integrase-mediated insertion and subsequent expression of HIS3 gene. Using this approach we successfully detected the reverse transcriptase activity of HBV in yeast strains defective in endogenous Ty1 expression. Consistent with the unique priming activity associated with HBV pol, the minus strand DNA synthesis was protein-primed. Deletion of HBV reverse transcriptase (RT) or RNase H domains resulted in a dramatic drop in histidine prototrophs. The addition of HBV encoded HBx protein in virus-like particles during in vitro RT reaction stimulated the RT reaction by severalfold. Furthermore, in the presence of 3TC, a known inhibitor of HBV reverse transcriptase, yeast His(+) growth of His protrophs was not observed. Thus, this approach, which is based on genetic selection in yeast, is safe, economic, and a reliable strategy with a potential for large scale screening of cofactors and inhibitors of HBV polymerase functions.
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PMID:Expression of hepatitis B virus polymerase in Ty1-his3AI retroelement of Saccharomyces cerevisiae. 1053 36


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