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:2.7.1.21 (thymidine kinase)
7,561 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Naive CD4+ T cells initially transcribe both IL-4 and IFN-gamma when stimulated with either the mitogen Con A or Ag in the presence of IL-4 or IL-12 and, therefore, appear uncommitted as to the pathway of differentiation they will follow. However, when stimulated either with Con A or with Ag in the presence of IL-4, CD4+ T cells become primed to follow the Th2 differentiation pathway, and we show now that by 48 h of culture in this environment these cells extinguish IFN-gamma gene transcription. Likewise, priming in the presence of IL-12 leads to the development of Th1 cells, which switch off the expression of the IL-4 gene. To clarify the Th1 differentiation pathway, we performed ablation studies using IL-4 thymidine kinase transgenic mice. When the antiviral drug ganciclovir was added 1 day after primary stimulation in the presence of IL-12, IFN-gamma- and IL-4-producing cells were ablated. In contrast, when ganciclovir was added 2 days after primary stimulation, IL-4-producing cells, but not IFN-gamma-producing cells, were ablated. Thus, our studies show that by 48 h after activation, Th1 or Th2 cells have already become polarized to the differentiation pathway that they will follow. As the differentiation toward Th1 and Th2 effector cells proceeds, substantial amounts of IFN-gamma and IL-4 mRNA accumulate, while the mRNAs of the corresponding lineage (i.e., IFN-gamma in the case of Th2 cells, and IL-4 in the case of Th1 cells) diminish to undetectable levels. IL-4R is up-regulated during T cell differentiation by a mechanism mediated mainly by IL-4. The fact that IL-12 priming does not suppress IL-4-dependent IL-4R up-regulation shows that both IL-4 mRNA and cytokine are produced by IL-12-primed naive CD4+ T cells during differentiation into Th1 cells. Naive CD4+ T cells, therefore, begin as uncommitted cells which express both Th1 and Th2 cytokines that rapidly extinguish the expression of the inappropriate cytokine as the commitment toward the effector lineages is made.
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PMID:Polarization of IL-4- and IFN-gamma-producing CD4+ T cells following activation of naive CD4+ T cells. 901 46

Direct delivery of the herpes simplex virus thymidine kinase (HSVtk) gene, in combination with the prodrug ganciclovir (GC), has been used for the treatment of localised, inoperable tumours. Several groups have shown that when rodent tumours are ablated in vivo with suicide genes, anti-tumour immunity can also be generated. Hence, this approach may also be useful in treating disseminated disease. Here we have studied the mechanisms associated with this anti-tumour immunity. In B16 HSVtk+ tumours being killed in vivo with GC treatment, we observed the induction of a pronounced intratumoural infiltrate of macrophages, CD4+ and CD8+ T cells. In addition, using reverse transcriptase polymerase chain reaction, expression of interleukin (IL)-2, IL-12, interferon-gamma (IFN-gamma), tumor necrosis factor-alpha (TNF-alpha) and granulocyte/macrophage colony-stimulating factor (GM-CSF) but not IL-4, IL-6 or IL-10, was observed, a profile of cytokine expression which resembles that of a Th1 immune response. To complement these findings, we also investigated the mechanisms by which expression of HSVtk leads to cell death. Our data show that B16/HSVtk+ cells die predominantly by necrosis, rather than apoptosis, on exposure to GC, a process which may be associated with the generation of anti-tumour inflammatory responses. From these data we propose a model for the induction of anti-tumour immunity using suicide genes and discuss the development of improved vectors for gene therapy to augment these effects in vivo.
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PMID:Generation of an anti-tumour immune response in a non-immunogenic tumour: HSVtk killing in vivo stimulates a mononuclear cell infiltrate and a Th1-like profile of intratumoural cytokine expression. 913 53

The potential of gene therapy strategies for malignant gliomas that are based on retroviral-mediated transfer of a "suicide gene" such as Herpes Simplex Virus-thymidine kinase HSV-tk and subsequent treatment by a prodrug (ganciclovir, for example), has been emphasized by the promising results obtained by several groups. However, further experimental data as well as preliminary clinical results indicate that the low efficiency of retroviral-mediated gene transfer in vivo as well as difficulties for the diffusion of the prodrug inside the tumour mass can limit the efficacy of this form of gene therapy. To achieve a more effective limitation of tumour growth other approaches may be combined with the "suicide gene" strategy and the enhancement of the immunological response to the tumour by cytokine gene transfer is prominent among these approaches. The authors' experiments in nude mice confirm the antineoplastic role of IL-4 and encourage testing the effects of the simultaneous transfer of IL-4 and HSV-tk genes in immunocompetent animals.
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PMID:Gene transfer of suicide genes for the treatment of malignant gliomas: efficacy, limitations, and perspectives for a combined immunotherapy. 923 23

In a cancer gene therapy model recombinant adenoviruses expressing the herpes simplex virus thymidine kinase (HSVtk) gene were injected into tumors in situ, either alone or in combination with adenoviruses (Avs) engineered to express IL-2, IL-6 or the costimulatory molecule B7-1. HSVtk phosphorylates the prodrug ganciclovir, thus converting it into an antimetabolite which kills not only HSVtk expressing cells, but also by the 'bystander effect', neighboring untransduced tumor cells. The tumors regressed in 80% of mice upon AvTK/ganciclovir treatment: combinations with AvIL-2, AvIL-6, or AvB7-1 did not improve these results. Cured mice were protected from further challenge with wild-type tumor but not from challenges with an unrelated syngeneic tumor cell line. Since cytotoxic T lymphocyte responses in this tumor model were weak, we analyzed cytokine secretion from spleen cells of treated animals. The best correlate of antitumor immunity in this model was enhanced secretion of GM-CSF, while secretion of IL-2, IL-6 and IFN gamma was also frequently increased but not as consistently. The enhanced IFN gamma secretion associated with unchanged IL-4 secretion suggests that AvTK treatment results in a predominantly Th1-mediated antitumor immune response.
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PMID:Characterization of the antitumor immune response generated by treatment of murine tumors with recombinant adenoviruses expressing HSVtk, IL-2, IL-6 or B7-1. 947 56

In order to investigate if immune responses to the fusion (F) protein of respiratory syncytial virus (RSV) could be influenced by cytokines, recombinant vaccinia viruses (rVV) carrying both the F gene of RSV and the gene for murine interleukin-2 (IL-2), IL-4, or gamma interferon (IFN-gamma) were constructed. In vitro characterization of rVV revealed that insertion of the cytokine gene into the VP37 locus of the vaccinia virus genome resulted in 100- to 1,000-fold higher expression than insertion of the same gene into the thymidine kinase (TK) locus. In comparison, only a two- to fivefold difference in the level of expression of the F protein was observed when the gene was inserted into either of these two loci. Mice vaccinated with rVV expressing the F protein and high levels of IL-2 or IFN-gamma cleared rVV more rapidly than mice inoculated with a control rVV and developed only low levels of RSV-specific serum antibody. In addition, these recombinants were much less effective at priming RSV-specific memory cytotoxic T lymphocytes (CTL) and IFN-gamma production by spleen cells than rVV expressing the F protein alone. In contrast, mice vaccinated with rVV expressing high levels of IL-4 showed signs of delayed rVV clearance. RSV-specific serum antibody responses were biased in favor of immunoglobulin G1 (IgG1) in these mice, as there was a significant reduction in IgG2a antibody responses compared with serum antibody responses in mice vaccinated with rVV expressing the F protein alone. However, vaccination with rVV expressing the F protein together with high levels of IL-4 did not alter the development of RSV-specific memory CTL or IFN-gamma production by RSV-restimulated splenocytes.
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PMID:Recombinant vaccinia virus coexpressing the F protein of respiratory syncytial virus (RSV) and interleukin-4 (IL-4) does not inhibit the development of RSV-specific memory cytotoxic T lymphocytes, whereas priming is diminished in the presence of high levels of IL-2 or gamma interferon. 955 97

To explore the potential for molecular immunotherapies in the treatment of malignant gliomas, we evaluated the efficacy of subcutaneous tumor cell vaccines in the treatment of intracranial 9L tumors, using 9L gliosarcoma cell lines stably transduced with the murine interleukin-4 cDNA (9L-IL4), the herpes simplex virus-thymidine kinase cDNA (9L-Tk) or both (9L-IL4-Tk). The expression of multiple genes from a single transcript was achieved by incorporating internal ribosomal entry site (IRES) cassettes in the retroviral constructs. Subcutaneous immunization of rats with nonirradiated 9L-IL4 cells or 9L-IL4-Tk cells followed by treatment with ganciclovir (GCV) completely protected the animals from a subsequent intracranial challenge with wild-type 9L cells. In contrast, only 50% of animals immunized with 9L-Tk cells and 0% of 9L-neo immunized animals rejected the same challenge with wild-type 9L. More importantly, treatment of established (day 3) intracranial 9L tumors with genetically engineered tumor cells resulted in long-term survival (> 100 days) for 25-43% of 9L-IL4-Tk immunized animals and for 27% of nonirradiated 9L-IL4 immunized animals. In striking contrast, no 9L-Tk, 9L-neo or irradiated 9L-IL4 immunized animals survived for more than 33 days. As a marker of a cellular immune response, splenocytes from nonirradiated 9L-IL4, 9L-Tk or 9L-IL4-Tk immunized animals produced interferon-gamma (IFN-gamma) in greater amounts than those from 9L-neo immunized or Hank's balanced salts solution (HBSS) treated animals when stimulated with wild-type 9L in vitro. Our findings support the use of tumor cell vaccines expressing the IL-4 and HSVtk genes for the treatment of malignant gliomas.
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PMID:Effective cytokine gene therapy against an intracranial glioma using a retrovirally transduced IL-4 plus HSVtk tumor vaccine. 1043 6

The expression of antigens or other molecules from recombinant vaccinia viruses requires the insertion of coding sequence at specific sites in the viral genome. Here we investigate the influence of two different sites on the level of protein expressed during a viral infection. The level of immune response in mice to vaccinia virus-expressed murine interleukin 2 (IL-2) or IL-4 varied depending on whether the coding sequence was inserted into the vaccinia virus thymidine kinase (tk) gene or into the HindIII F fragment of the viral genome where herpes simplex virus (HSV) tk was used as a selectable marker. In each case the intensity of the response was greater when the relevant gene was expressed from the HindIII F insertion site. In order to quantify these differences a series of recombinant viruses expressing luciferase was constructed. Luciferase activity from coding sequence inserted into the HindIII F fragment was significantly higher than that from the tk gene insertion, provided HSV tk(+) constructs were compared. Insertion of a marker gene (HSV tk) into the HindIII F site with disruption of the F7L open reading frame led to a reduced level of luciferase expressed from the tk insert, despite more than 45 kb of intervening sequence. In mice, luciferase expression was higher from the HindIII F inserted gene than from the tk insert in both lungs and ovaries.
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PMID:Insertion sites for recombinant vaccinia virus construction: effects on expression of a foreign protein. 1064 42

Vaccination with cytokine-transduced tumor cells represents a potentially important approach to the treatment of central nervous system tumors. We have recently demonstrated the therapeutic efficacy of tumor cell vaccines expressing the murine interleukin 4 (IL-4) and the herpes simplex virus-thymidine kinase in a rat brain tumor model in which nonirradiated vaccine cells can be eliminated by the subsequent administration of ganciclovir. In this report, we demonstrate the construction and characterization of a retroviral vector that encodes human IL-4, neomycin phosphotransferase, and herpes simplex virus-thymidine kinase genes for use in human clinical trials. An MFG-based retroviral vector was used to generate the recombinant retrovirus, TFG-hIL4-Neo-Tk, in which a long terminal repeat-driven polycistronic transcript encodes three cDNAs that are linked and coexpressed using two intervening internal ribosome entry site fragments from the encephalomyocarditis virus. The amphotropic retroviral vector TFG-hIL4-Neo-Tk was then used to infect human primary glioma cultures and skin-derived fibroblasts. After infection and G418 selection, cells produced 89-131 ng/10(6) cells/48 hours of human IL-4, which was determined to be biologically active. Transduced glioma cells were highly sensitive to the cytotoxic effect of ganciclovir. These data demonstrate the suitability of the TFG-hIL4-Neo-Tk vector for therapeutic studies of cytokine-transduced autologous tumor vaccination in patients with malignant gliomas.
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PMID:Characterization and transduction of a retroviral vector encoding human interleukin-4 and herpes simplex virus-thymidine kinase for glioma tumor vaccine therapy. 1076 55

Nutritional deficiency of zinc is widespread throughout developing countries, and zinc-deficient persons have increased susceptibility to a variety of pathogens. Zinc deficiency in an experimental human model caused an imbalance between Th1 and Th2 functions. Production of interferon-gamma and interleukin (IL)-2 (products of Th1) were decreased, whereas production of IL-4, IL-6, and IL-10 (products of Th2) were not affected during zinc deficiency. Zinc deficiency decreased natural killer cell lytic activity and percentage of precursors of cytolytic T cells. In HuT-78, a Th0 cell line, zinc deficiency decreased gene expression of thymidine kinase, delayed cell cycle, and decreased cell growth. Gene expression of IL-2 and IL-2 receptors (both alpha and beta) and binding of NF-kappaB to DNA were decreased by zinc deficiency in HuT-78. Decreased production of IL-2 in zinc deficiency may be due to decreased activation of NF-kappaB and subsequent decreased gene expression of IL-2 and IL-2 receptors.
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PMID:Effects of zinc deficiency on Th1 and Th2 cytokine shifts. 1094 85

The class II transactivator (CIITA) is induced by gamma interferon (IFN-gamma) and activates major histocompatibility complex class II; however, this report shows it suppresses other genes. An N-terminal 36 amino acids of CIITA mediates suppression of the collagen alpha(2)(I) promoter via binding to CREB-binding protein (CBP). Reconstitution of cells with CBP reverts this suppression. IFN-gamma is known to inhibit collagen gene expression; to test if CIITA mediates this gene suppression, a mutant cell line defective in CIITA induction but not in the activation of STAT1/JAK/IRF-1 is studied. IFN-gamma suppression of the collagen promoter and the endogenous gene is observed in the wild-type control but not in the mutant line. Suppression is restored when CIITA is introduced. Other targets of CIITA-mediated promoter suppression include interleukin 4, thymidine kinase, and cyclin D1.
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PMID:A 36-amino-acid region of CIITA is an effective inhibitor of CBP: novel mechanism of gamma interferon-mediated suppression of collagen alpha(2)(I) and other promoters. 1156 90


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