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:2.4.2.8 (hypoxanthine-guanine phosphoribosyltransferase)
2,527 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Enzyme activities were studied in peripheral blood lymphocytes from patients infected with, or at risk for, infection with human immunodeficiency virus (HIV). No significant differences were observed in the HIV-infected and HIV-seronegative high-risk patients with regard to enzyme activities of hypoxanthine-guanine phosphoribosyltransferase (EC 2.4.2.8) and purine nucleoside phosphorylase (EC 2.4.2.1) in peripheral blood. Adenosine deaminase (EC 3.5.4.4) was significantly (P less than 0.02) depressed in asymptomatic HIV-seropositive patients and HIV-seronegative patients at high risk of HIV infection as compared with a healthy HIV-seronegative population. Adenosine kinase (AK, EC 2.7.1.20) was significantly increased in the asymptomatic seropositive (P less than 0.02) and also in the HIV-seronegative high-risk groups (P = 0.01) compared with the normal controls. AK activity was significantly lower in subjects with AIDS than in the asymptomatic (P less than 0.002) and high-risk groups (P less than 0.01). Taken together, these results indicate that adenosine deaminase and AK activities are influenced by the health of the patient, and that measurement of AK activity may prove useful in monitoring the clinical progress of patients with HIV infection.
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PMID:Depressed activities of purine enzymes in lymphocytes of patients infected with human immunodeficiency virus. 254 31

It has been postulated that HIV-infected patients undergo an active production of virus and CD4+ T cell destruction from the early stages of the disease, and that an extensive postthymic expansion of CD4+ T cells prevents a precipitous decline in CD4+ T cell number. Based on the rebound of the CD4+ T cell number observed in patients undergoing antiretroviral therapy with protease inhibitors, it has been calculated that, on average, 5% of T cells are replaced every day in HIV-infected patients. To obtain an independent estimate of the recycling rate of T cells in the patients, we measured the frequency of cells carrying a loss-of-function mutation at the hypoxanthine guanine phosphoribosyl transferase (hprt) locus. Assuming a recycling rate of 5%/d, an accumulation of 2.6 mutations/10(6)/yr over the physiological accumulation was predicted. Indeed, we observed an elevated frequency of HPRT mutants in the CD4+ T cells of most patients with < 300 CD4+ T cells/mm3 of blood and in the CD8+ T cells of most patients with < 200 CD4+ T cells/mm3, consistent with an elevated and protracted increased division rate in both subsets. However, in earlier stages of the disease the mutant frequency in both CD4+ and CD8+ T cells was lower than in healthy controls. The cytokine production profile of most HPRT mutant CD4+ T cell clones from both healthy and HIV-infected patients was typical of T helper cells type 2 (high IL-4 and IL-10, low IFN-gamma), whereas the cytokine production pattern of wild-type clones was heterogeneous. The cytokine profile of CD8+ clones was indistinguishable between HPRT mutants and wild type. Our data provide evidence of increased CD4+ and CD8+ T cell recycling in the HIV-infected patients.
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PMID:Frequency and cytokine profile of HPRT mutant T cells in HIV-infected and healthy donors: implications for T cell proliferation in HIV disease. 904 68

Recombinant vaccinia viruses based on the highly attenuated Modified Vaccinia Ankara (MVA) strain expressing HIV-1 antigen genes were constructed by a novel procedure involving the transient use of two marker genes. The selectable markers used, the Escherichia coli guanine phosphoribosyltransferase (gpt) and the beta-galactosidase (lacZ) genes, are not retained within the final recombinant virus. The transient marker stabilisation (TMS) procedure allows the generation of marker-free recombinant viruses in a series of simple plaque purification steps. HIV-1 gag pol genes were inserted into two loci of vaccinia MVA demonstrating the efficiency of the procedure.
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PMID:Transient marker stabilisation: a general procedure to construct marker-free recombinant vaccinia virus. 957 48

Perinatal treatment with 3'-azido-3'-deoxythymidine (AZT) has been found to reduce the rate of maternal-infant transmission of HIV; however, AZT is genotoxic in mammalian cells in vitro and induces tumors in the offspring of mice treated in utero. The purpose of the present study was to investigate the relationships between incorporation of AZT into DNA, and the frequency and spectrum of mutations at the HPRT locus of the human lymphoblastoid cell line, TK6, following in vitro exposures to AZT. Cells were cultured in medium containing 0 or 300 microM AZT for 1, 3, or 6 day(s) (n = 5/group). The effects of exposure duration on incorporation of AZT into DNA and HPRT mutant frequency were determined using an AZT radioimmunoassay and a cell cloning assay, respectively. AZT accumulated in DNA in a supralinear manner, approaching a plateau at 6 days of treatment (101.9 +/- 14.7 molecules AZT/10(6) nucleotides). After 3 days of AZT exposure, HPRT mutant frequency was significantly increased (1.8-fold, p = 0.016) compared to background (mutant frequency = 3.78 x 10(-6)). Multiplex PCR amplification of genomic DNA was used to determine the frequency of exon deletions in HPRT mutant clones from untreated cells versus AZT-treated cells. Molecular analyses of AZT-induced mutations revealed a significant difference in the frequency of total gene deletions (44/120 vs. 18/114 in controls, p = 0.004 by the Mann-Whitney U-statistic). In fact, the Chi-square test of homogeneity demonstrate that the differences between the control and AZT-treatment groups is attributed mainly to this increase in total gene deletion mutations (p = 0.00001). These data indicate that the primary mechanism of AZT mutagenicity in human TK6 cells is through the production of large deletions which occur as a result of AZT incorporation into DNA and subsequent chain termination. The data imply that perinatal chemoprophylaxis with AZT may put children of HIV-infected women at potential risk for genetic damage.
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PMID:Genotoxicity of 3'-azido-3'-deoxythymidine in the human lymphoblastoid cell line, TK6: relationships between DNA incorporation, mutant frequency, and spectrum of deletion mutations in HPRT. 1052 9

Drug combinations that include nucleoside reverse transcriptase inhibitors (NRTIs) are remarkably effective in preventing maternal-viral transmission of HIV during pregnancy. However, there may be potential long-term risks for children exposed in utero. Examination of the genotoxic and mutagenic effects of two NRTIs, zidovudine [AZT (3'-azido-3'-deoxythymidine)] and didanosine [ddI (2',3'-dideoxyinosine)], in cultured human lymphoblastoid cells revealed multiplicative synergistic enhancement of AZT-DNA incorporation and mutant frequency induction in response to the combined drug exposure, as compared with single-drug exposures. Dose-related increases in DNA incorporation of AZT (as measured by a competitive RIA) and mutagenicity at the HPRT and TK loci (as assessed by cell-cloning assays) were observed in cells exposed in culture to AZT, or equimolar combinations of AZT + ddI, at exposure concentrations ranging from 3 to 30 times the maximum plasma levels found in humans. Because mutagenesis is strongly associated with tumor induction in experimental models, children exposed transplacentally to combinations of NRTIs may be at risk for cancer development later in life.
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PMID:Zidovudine-didanosine coexposure potentiates DNA incorporation of zidovudine and mutagenesis in human cells. 1105 53

Combinations of antiretroviral drugs that include nucleoside reverse transcriptase inhibitors (NRTIs) are superior to single-agent regimens in treating or preventing HIV infection, but the potential long-term health hazards of these treatments in humans are uncertain. In earlier studies, our group found that coexposure of TK6 human lymphoblastoid cells to 3'-azido-2',3'-dideoxythymidine (AZT) and 2',3'-dideoxyinosine (ddI), the first two NRTIs approved by the FDA as antiretroviral drugs, produced multiplicative synergistic enhancement of DNA incorporation of AZT and mutagenic responses in both the HPRT and TK reporter genes, as compared with single-drug exposures (Meng Q et al. [2000a]: Proc Natl Acad Sci USA 97:12667-12671). The purpose of the current study was to characterize the mutational specificity of equimolar mixtures of 100 microM or 300 microM AZT + ddI at the HPRT and TK loci of exposed cells vs. unexposed control cells, and to compare the resulting mutational spectra data to those previously found in cells exposed to AZT alone (Sussman H et al. [1999]: Mutat Res 429:249-259; Meng Q et al. [2000b]: Toxicol Sci 54:322-329). Molecular analyses of HPRT mutant clones were performed by reverse transcription-mediated production of cDNA, PCR amplification, and cDNA sequencing to define small DNA alterations, followed by multiplex PCR amplification of genomic DNA to define the fractions of deletion events. TK mutants with complete gene deletions were distinguished by Southern blot analysis. The observed HPRT mutational categories included point mutations, microinsertions/microdeletions, splicing-error mutations, and macrodeletions including partial and complete gene deletions. The only significant difference or shift in the mutational spectra for NRTI-treated cells vs. control cells was the increase in the frequency of complete TK gene deletions following exposures (for 3 days) to 300 microM AZT-ddI (P = 0.034, chi-square test of homogeneity); however, statistical analyses comparing the observed mutant fraction values (measured mutant frequency x percent of a class of mutation) between control and NRTI-treated cells for each class of mutation showed that the occurrences of complete gene deletions of both HPRT and TK were significantly elevated over background values (0.34 x 10(-6) in HPRT and 6.0 x 10(-6) in TK) at exposure levels of 100 microM AZT-ddI (i.e., 1.94 x 10(-6) in HPRT and 18.6 x 10(-6) in TK) and 300 microM AZT-ddI (i.e., 5.6 x 10(-6) in HPRT and 34.6 x 10(-6) in TK) (P < 0.05, Mann-Whitney U-statistic). These treatment-related increases in complete gene deletions were consistent with the spectra data for AZT alone (ibid.) and with the known mode of action of AZT and ddI as DNA chain terminators. In addition, cotreatments of ddI with AZT led to substantial absolute increases in the mutant fraction of other classes of mutations, unlike cells exposed solely to AZT [e.g., the frequency of point mutations among HPRT mutants was significantly increased by 130 and 323% over the background value (4.25 x 10(-6)) in cells exposed to 100 and 300 microM AZT-ddI, respectively]. These results indicate that, at the same time that AZT-ddI potentiates therapeutic or prophylactic efficacy, the use of a second NRTI with AZT may confer a greater cancer risk, characterized by a spectrum of mutations that deviates from that produced solely by AZT.
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PMID:Molecular analysis of mutations at the HPRT and TK loci of human lymphoblastoid cells after combined treatments with 3'-azido-3'-deoxythymidine and 2',3'-dideoxyinosinedagger. 1211 80

The current worldwide spread of the human immunodeficiency virus-1 (HIV-1) to the heterosexual population has resulted in approximately 800,000 children born yearly to HIV-1-infected mothers. In the absence of anti-retroviral intervention, about 25% of the approximately 7,000 children born yearly to HIV-1-infected women in the United States are HIV-1 infected. Administration of zidovudine (AZT) prophylaxis during pregnancy reduces the rate of infant HIV-1 infection to approximately 7%, and further reductions are achieved with the addition of lamivudine (3TC) in the clinical formulation Combivir. Whereas clinically this is a remarkable achievement, AZT and 3TC are DNA replication chain terminators known to induce various types of genotoxicity. Studies in rodents have demonstrated AZT-DNA incorporation, HPRT mutagenesis, telomere shortening, and tumorigenicity in organs of fetal mice exposed transplacentally to AZT. In monkeys, both AZT and 3TC become incorporated into the DNA from multiple fetal organs taken at birth after administration of human-equivalent protocols to pregnant dams during gestation, and telomere shortening has been found in monkey fetuses exposed to both drugs. In human infants, AZT-DNA and 3TC-DNA incorporation as well as HPRT and GPA mutagenesis have been documented in cord blood from infants exposed in utero to Combivir. In infants of mice, monkeys, and humans, levels of AZT-DNA incorporation were remarkably similar, and in newborn mice and humans, mutation frequencies were also very similar. Given the risk-benefit ratio, these highly successful drugs will continue to be used for prevention of vertical viral transmission, however evidence of genotoxicity in mouse and monkey models and in the infants themselves would suggest that exposed children should be followed well past adolescence for early detection of potential cancer hazard.
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PMID:Perinatal genotoxicity and carcinogenicity of anti-retroviral nucleoside analog drugs. 1531 87

We report here the use of TEV protease cleavable fusion proteins to produce glycosylated bioactive peptides and proteins. Bacterial expression was utilized to produce two fusion proteins, GPRT-C37-H6 and His-tagged interleukin-2 (amino acids 6-133), which when cleaved by the tobacco etch virus NIa protease (TEV protease) to generate HIV entry inhibitor peptide C37-H6 and a truncated version of the cytokine interleukin-2, both containing N-terminal cysteines. The N-terminal cysteine containing C37-H6 and truncated interleukin-2 were then joined to a synthetic glycopeptide thioester utilizing native chemical ligation under nondenaturing and denaturing conditions, respectively. The ligations of the glycopeptide to the C37-H6 peptide and the truncated interleukin-2 protein both proceeded in high yield, though the size, and physical properties of the two polypeptides differ greatly.
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PMID:A new strategy for glycoprotein synthesis: ligation of synthetic glycopeptides with truncated proteins expressed in E. coli as TEV protease cleavable fusion protein. 1565 56

Experiments were performed to investigate the impact of didanosine (ddI), lamivudine (3TC), and stavudine (d4T) on cell survival and mutagenicity in two reporter genes, hypoxanthine-guanine phosphoribosyltransferase (HPRT) and thymidine kinase (TK), using a cell cloning assay for assessing the effects of individual nucleoside analogs (NRTIs)/drug combinations in human TK6 B-lymphoblastoid cells. Three-day treatments with 0, 33, 100, or 300 microM ddI, 3TC, or ddI-3TC produced positive trends for increased HPRT and TK mutant frequencies. While dose-related trends were too small to reach significance after treatments with d4T or d4T-3TC, pairwise comparisons with control cells indicated that exposure to 100 microM d4T or d4T-3TC caused significant elevations in HPRT mutants. Measurements of mutagenicity in cells exposed to d4T (or d4T-3TC) were complicated by the cytotoxicity of this NRTI. Enhanced increases in mutagenic responses to combined NRTI treatments, compared with single drug treatments, occurred as additive to synergistic effects in the HPRT gene of cells exposed to 100 microM ddI-3TC or 100 microM d4T-3TC, and in the TK gene of cells exposed to 100 or 300 microM ddI-3TC. Comparisons of these data to mutagenicity studies of other NRTIs in the same system (Meng Q et al. [2000c]: Proc Natl Acad Sci USA 97:12667-126671; Torres SM et al. [2007]: Environ Mol Mutagen) indicate that the relative mutagenic potencies for all drugs tested to date are: AZT-ddI > ddI-3TC > AZT-3TC congruent with AZT-3TC-ABC (abacavir) > AZT >/=ddI > d4T-3TC > 3TC > d4T >/= ABC. These collective data suggest that all NRTIs with antiviral activity against HIV-1 may cause host cell DNA damage and mutations, and impose a cancer risk.
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PMID:Relative mutagenic potencies of several nucleoside analogs, alone or in drug pairs, at the HPRT and TK loci of human TK6 lymphoblastoid cells. 1735 29

The current incidence of human immunodeficiency virus (HIV-1)/AIDS affects around 7000 pregnant women in the United States. When given during pregnancy, the nucleoside analog 3'-azido-3'-deoxythymidine (AZT) significantly reduces maternal-fetal transmission. It has been previously shown that AZT is incorporated into DNA, where it causes mutations in the HPRT and TK genes. It also changes cell cycle gene expression, and induces S-phase arrest, micronuclei, chromosomal aberrations, sister chromatid exchanges, telomeric attrition, and other genotoxic effects in cultured cells. A predicted consequence of these events is genomic instability that together, with clastogenicity may contribute to the carcinogenic potency of AZT. Various aspects of genotoxicity are explored in this contribution seeking to understand the multiple effects of this antiretroviral agent in animal models and humans. This mini-review describes some of the experimental models used to elucidate the genotoxicity induced by antiretroviral therapy during human pregnancy. The use of diverse methods to detect biomarkers of exposure, such as an AZT-specific radioimmunoassay, micronuclei bearing intact chromosomes, and telomeric DNA attrition highlight the role of in vitro models to elucidate exposure and risk. The relevance of the in vitro models is followed by the introduction of the role of the nucleoside analogs in transplacental carcinogenesis along with the description of a transplacental perfusion model and a transplacental carcinogenesis rodent model. In a more direct clinical application the use of AZT-DNA incorporation as a biomarker of exposure, in experiments conducted in vivo in Erythrocebus patas monkeys and in humans, addresses the possibility of elucidation of potential cancer risk in those infants exposed in utero. Two relevant aspects of this contribution are the potential application of some of the models described in this mini-review, as diagnostic tools in antiretroviral-exposed populations, and the use of these models to understand the nature of the genotoxicities and minimize the undesirable side effects of the antiretroviral therapy.
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PMID:Relevance of experimental models for investigation of genotoxicity induced by antiretroviral therapy during human pregnancy. 1829 33


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