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)

Treatment of HIV-1-infected cells with the HIV-1-specific inhibitors hydroxyethoxymethylphenylthiothymine (HEPT), tetrahydroimidazobenzodiazepinones (TIBO), nevirapine, pyridinone, bis(heteroaryl)piperazines (BHAP), and tert-butyldimethylsilylspiroaminooxathioledioxide (TSAO) at a concentration of 0.1 microgram/ml resulted in a rapid breakthrough of resistant virus within three to four subcultivations. At drug concentrations of 0.5 to 1 microgram/ml, emergence of resistant virus was delayed. The drug-resistant HIV-1 strains that originated under these conditions were genetically and phenotypically characterized and showed differential sensitivities against the different classes of HIV-1-specific inhibitors depending on the amino acid substitutions in their reverse transcriptase. Novel amino acid substitutions were found in the reverse transcriptase of BHAP- and pyridinone-resistant mutant HIV-1 strains that had not been reported so far. At 2.5 to 10 micrograms/ml, that is at a concentration 100- to 250-fold higher than the 50% effective concentration (EC50), HEPT, TIBO, nevirapine, pyridinone, and BHAP prevented virus breakthrough after 15 subcultivations. In contrast, 3'-azido-2',3'-dideoxythymidine (AZT), even when administered at a 1000-fold higher concentration (i.e., 1.3 micrograms/ml) than its EC50 failed to prevent virus breakthrough after the second subcultivation. HIV-1-infected cell cultures could apparently be cleared from virus by the HIV-1-specific inhibitors when used at the "knocking-out" concentrations (2.5-10 micrograms/ml), as evidenced by (i) the lack of viral cytopathicity, (ii) the lack of virus-specific envelope glycoprotein expression, (iii) the lack of viral p24 antigen production, and (iv) the apparent absence of proviral DNA in the cells. Moreover, uninfected CEM cell cultures to which HIV-1-infected CEM cells (including syncytia) had been added were protected from destruction by high-concentration treatment with the HIV-1-specific inhibitors, but not with AZT and DDI (2',3'-dideoxyinosine).
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PMID:Knocking-out concentrations of HIV-1-specific inhibitors completely suppress HIV-1 infection and prevent the emergence of drug-resistant virus. 769 May 1

We have infected ten-day-old primary cultures of human monocyte-derived macrophages (MDM) with HIV-1 by cocultivation with chronically infected monocytic cell lines. This work has involved the U-937 monocytoid cell line, chronically infected with the HIV-IIIB strain of HIV-1 (U-937HIV IIIB) as well as a number of cell clones, termed UHC, which were derived from U-937HIV IIIB by limiting dilution. Cell-free virus, derived from each of U-937HIV IIIB cells and the UHC1 clone were noninfectious for MDM, as determined by failure to express viral p24 antigen (Ag). In contrast, viral p24 Ag production was detected in MDM that had been cocultivated with U-937HIV IIB, and with each of three UHC clones that produced infectious virus. Infection, in each case, was confirmed by polymerase chain reaction detection via the amplification of proviral DNA. In contrast, cocultivation with the UHC15.7 clone, which fails to cleave viral gp160 to its gp120 and gp41 products or the UHC8 clone, which lacks functional reverse transcriptase, did not lead to infection of MDM. Pretreatment of MDM for 2 hr with 1 microM AZT completely prevented infection by culture fluids containing HIVada, a macrophage-tropic virus, but did not affect infection mediated by cocultivation. These results suggest that cell-to-cell transmission of HIV-1, among monocyte-derived macrophages, can be mediated by proviral DNA. Moreover, gp120 at the surface of infected cells may play an important role in this process, since cell-to-cell HIV transmission could not be demonstrated with the UHC clone that is defective in cleavage of the viral envelope glycoprotein gp160.
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PMID:Infection of human monocyte-derived macrophages by human immunodeficiency virus mediated by cell-to-cell transmission. 769 62

We examined the ability of human immunodeficiency virus (HIV) type 1 (HIV-1) to infect in vitro, primary brain-derived human microvascular endothelial cells (HMEC) that constitute the blood-brain barrier (BBB). Immunofluorescence (IFA) and antigen capture assays failed to demonstrate p24 antigen from HIV inoculated endothelial cells and supernatants did not contain detectable levels of reverse transcriptase (RT). HIV could be rescued by cocultivation of infected HMEC with a susceptible T-lymphocyte line (CEM-SS), which were then shown to form syncytia and produce RT activity and p24 Ag (IFA, antigen captive assay). Polymerase chain reaction (PCR) was successfully used to amplify HIV-specific gag and env gene sequences from HMEC. CD4 expression was not identified on these cells by IFA. These results suggest that HIV infection of BBB endothelium occurs, but that viral replication is minimal. Infection of the BBB by HIV may give the virus a foothold in the CNS and suggests that the brain might be infected directly and may not be limited to just the passage of infected mononuclear cells.
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PMID:HIV-1 infection of human brain-derived microvascular endothelial cells in vitro. 769 39

Beside the threat of infection via HIV-containing blood, the ophthalmologist is especially interested in the possibility of HIV infection via the tears of HIV-positive persons. In a first step, we tried to isolate HIV-1 from the peripheral blood lymphocytes (PBL) of 50 HIV-1-antibody-positive persons in different stages of disease and to detect reverse transcriptase (RT) and p24 antigen (p24-Ag) in the supernatant. Simultaneously we carried out the same tests on tears of these patients. In 10 persons tears were collected using Schirmer strips, in 40 persons by means of microcapillaries. In a second step 10 sample pairs (PBL and tears) were tested with the polymerase chain reaction to detect proviral sequences of HIV-1 (gag, pol, env). In the first step it was not possible to isolate HIV-1 from tears, nor was it possible to detect RT or p24-Ag from the supernatant. In contrast, this was successful in 32 of the 50 examined cases for the PBL. In the second step, it was possible to detect gag, pol and env in all 10 PBL samples, while gag and pol could be detected only in one tear sample and env not at all. Our results show that the tears of HIV-positive persons contain extremely low quantities of tissue-infectious units of HIV. In addition, proviral sequences seem to occur in much lower frequency in tears than in PBL. Infection with HIV via tears therefore appears very unlikely. These findings make it possible to assign tears a place in a semiquantitative ranking of different body fluids by HIV-1 concentration.
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PMID:[HIV-1 and tears. Results of virus isolation and polymerase chain reaction (PCR)]. 781 1

Forty-five subjects with symptomatic human immunodeficiency virus type 1 (HIV-1) infection, CD4+ lymphocyte counts of > or = 150 x 10(6)/L, and Karnofsky scores > or = 60 were enrolled in a multicenter, randomized, controlled trial that compared zidovudine monotherapy and combination therapy for 48 weeks with zidovudine and interferon-alpha (IFN-alpha). Zidovudine with IFN-alpha (n = 25) had a favorable effect on CD4+ cell counts compared with zidovudine alone (n = 20). At all time points analyzed, the mean change from baseline was higher, reaching significance at week 24 (+10% versus -21%; P = .029). At week 48 the difference was -12% versus -45% (P = .07). Anti-CD3 monoclonal antibody-induced T cell reactivity improved temporarily in both groups. Serum HIV p24 antigen levels decreased maximally during the first 12 weeks of treatment. At weeks 0 and 48, polymerase chain reaction analysis for mutations at codons 67 and 215 of the HIV-1 reverse transcriptase gene conferring zidovudine resistance was conducted in 10 subjects receiving zidovudine and in 8 subjects receiving combination therapy. At week 48, 1 of 8 and 4 of 6 samples from the groups receiving zidovudine only or combination therapy, respectively, contained wild type virus at codon 215. Grade 3 or 4 toxicity was uncommon. Drug-related malaise and anorexia were observed more frequently in patients receiving both zidovudine and IFN-alpha.
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PMID:Zidovudine and interferon-alpha combination therapy versus zidovudine monotherapy in subjects with symptomatic human immunodeficiency virus type 1 infection. 791 Aug 38

Mycosis fungoides (MF) is a rare form of cutaneous T cell lymphoma suspected of having a viral etiology. As in adult T cell leukemia, the virus involved may be human T lymphotropic virus type 1 (HTLV-1). We cultured the peripheral blood mononuclear cells (PBMC) of 29 patients with MF HTLV-1 seronegative by enzyme-linked immunosorbent assay and Western blot. The presence of reverse transcriptase (RT) and p24 antigen was investigated in the concentrate supernatant of the culture. The DNA of all studied patients was submitted to polymerase chain reaction and Southern blot analysis using primers and probes recognizing the tax region of HTLV-1/2 and the pol region of HTLV-1. 10 of 29 patients were found positive to HTLV-1, whereas they were always negative to RT and p24. The same results were confirmed in double blind after 6 mo. Our findings suggest HTLV-1 may be involved in the etiology of MF, at least in certain cases.
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PMID:Persistence of human T cell lymphotropic virus type 1 (HTLV-1) sequences in peripheral blood mononuclear cells from patients with mycosis fungoides. 796 73

In this study, we examined the impact of the predominantly Th2-type lymphokines interleukin 13 (IL-13) and interleukin 4 (IL-4) on acute infection of human bronchoalveolar macrophages with a macrophage-tropic isolate of human immunodeficiency virus type 1 (HIV-1). Addition of 0.01-10 ng of IL-4 or IL-13 per milliliters significantly blocked HIV-1 replication in infected cells, judging from levels of reverse transcriptase and p24 antigen in the supernatants of infected cells. Both IL-4 and IL-13 were almost as efficient as interferon-gamma (IFN-gamma) in preventing HIV-1 replication, when given in equivalent amounts. Moreover, neither IL-13 nor IL-4 interfered with the IFN-gamma-mediated enhancement of anti-HIV-1 activity in alveolar macrophages. Both IL-4 and IL-13 interfered with enhanced replication of HIV-1 in macrophages pulsed with the growth factor granulocyte-macrophage colony-stimulating factor (GM-CSF). Interleukin 13 also prevented HIV-1 release from peripheral blood mononuclear cells in a cocultivation experiment with feeder cells from a seronegative subject. These data suggest that Th2-derived lymphokines have significant anti-HIV-1 activity in cells of the macrophage lineage, although they may enhance the susceptibility of HIV-1-infected subjects to some opportunistic pathogens.
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PMID:Interleukin 13 and interleukin 4 protect bronchoalveolar macrophages from productive infection with human immunodeficiency virus type 1. 798 85

Treatment of monocytes with interferon-gamma 1 day before, or at the time of infection with human immunodeficiency virus type-1 (HIV-1) induced complete resistance in monocytes against HIV-1 infection. There was no evidence of viral RNA, proviral DNA, p24 antigen, or reverse transcriptase activity through 2 weeks after inoculation. Ultrastructural examination of these cells showed no detectable virus particles. When interferon-gamma was added to monocytes 1 to 3 days post-infection, virus integration occurred, but the viral expression was either ablated (1 day post-infection) or significantly inhibited (3 days post-infection). Treatment of monocytes with interferon-gamma before or after infection with HIV-1 produced significantly higher levels of tumor necrosis factor-alpha and interleukin-8 than untreated or uninfected monocytes. These results suggest that altered regulation of cytokines may mediate antiviral activity of interferon-gamma in monocytes.
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PMID:Interferon-gamma induces resistance in primary monocytes against human immunodeficiency virus type-1 infection. 800 12

Several groups have shown that peripheral CD8+ lymphocytes can be infected with human immunodeficiency virus type 1 (HIV-1), resulting in noncytopathic infection and persistent production of viral particles. We studied the ability of 3'-azido-3'-deoxythymidine (AZT) and 2',3'-dideoxyinosine (ddI) to inhibit the establishment of HIV-1 infection in CD8+ cells that were derived from cultures of peripheral blood lymphocytes exposed to both virus and drug. In situ infection of CD8+ cells was demonstrated by double flow cytometry analysis by using both anti-glycoprotein 120 (anti-gp120) and anti-CD8 monoclonal antibodies. At higher concentrations of drug (e.g., 0.4 microM AZT), the production of viral particles was inhibited for over 2 months, as assessed by p24 antigen levels in the culture medium. We also performed a time course experiment to determine whether HIV-1 infection of CD8+ cells would be affected by treatment of peripheral blood lymphocytes with AZT or ddI for different intervals following exposure to virus. Quantitative PCR revealed that 0.4 microM AZT, added as late as 24 h after infection, interfered with the formation of proviral DNA in CD8+ cells. Both HIV-1 load and the production of progeny virions by CD8+ cells, as monitored by reverse transcriptase activity in culture fluids, were inhibited by both AZT and ddI in a dose-dependent manner.
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PMID:Effect of 3'-azido-3'-deoxythymidine and 2',3'-dideoxyinosine on establishment of human immunodeficiency virus type 1 infection in cultured CD8+ lymphocytes. 806 81

We sought to validate an in vitro system which could predict the minimal effect dose of antiretroviral agents. Mixtures of uninfected CEM cells and CEM cells chronically infected with human immunodeficiency virus (HIV) type 1 MN were exposed to 2',3'-didehydro-3'-deoxythymidine (D4T) in vitro in a hollow-fiber model which simulates the plasma concentration-time profile of D4T in patients. Drug concentration was adjusted to simulate continuous intravenous infusion, or an intravenous bolus administered twice daily. The effect of the dosing regimen was measured with viral infectivity, p24 antigen, and reverse transcriptase or PCR for unintegrated HIV DNA. Dose deescalation studies on a twice-daily dosing schedule predicted a minimum effect dose of 0.5 mg/kg of body weight per day which correlated with the results of a clinical trial. Antiviral effect was demonstrated to be independent of schedule for every 12-h dosing versus continuous infusion. Finally, at or near the minimal effect dose, efficacy appeared to depend on the viral load. The ability of this in vitro pharmacodynamic model to assess the response of HIV-infected cells to different doses and schedules of antiviral agents may be useful in the design of optimal dosing regimens for clinical trials but requires validation with other types of antiretroviral agents.
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PMID:Effect of 2',3'-didehydro-3'-deoxythymidine in an in vitro hollow-fiber pharmacodynamic model system correlates with results of dose-ranging clinical studies. 809 42


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