Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:2.7.1.21 (thymidine kinase)
 7,561 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

We describe a new retroviral vector system pSXLC/pHa that utilizes a putative internal ribosome entry site (IRES) from encephalomyocarditis virus downstream from a multicloning site to co-express drug-selectable markers with a second non-selectable cDNA in a eukaryotic expression vector. The positive drug-selectable marker, MDR1, and the positive-negative marker, herpes simplex virus thymidine kinase (HSV-TK), were successfully introduced and expressed in the pSXLC/pHa system. The pSXLC-MDR and pSXLC-TK vectors contain the drug-selectable genes under translational control of the IRES and multiple cloning sites upstream for insertion of second cDNAs which can be co-expressed in this system. The inserts of these pSXLC plasmids were designed for easy transfer to the pHa retrovirus vector which has a strong promoter from Harvey murine sarcoma virus. The IRES-MDR-carrying retroviral vector, pHa-MCS-IRES-MDR, conferred resistance to vincristine and adriamycin. The IRES-TK-containing vector, pHa-MCS-IRES-TK conferred HAT-resistance in TK-deficient cells and the transfectants showed hypersensitivity to ganciclovir. These "flexible" vectors should be useful for co-expression of genes for selectable gene transfer and for positive-negative (suicide) selections in vitro and in vivo.
 ...
PMID:Efficient expression of drug-selectable genes in retroviral vectors under control of an internal ribosome entry site. 776 14

Five different gene transfer protocols have progressed into human clinical trials for the treatment of brain tumors. Two utilize the in vivo transfer of the Herpes Simplex-thymidine kinase (HS-tk) gene by either retroviral or adenoviral gene transfer. HS-tk confers a sensitivity to the anti-herpes drug ganciclovir (GCV). Insertion of HS-tk into tumors and subsequent treatment with GCV has successfully eliminated tumors in experimental animal models despite less than a 100% gene transfer efficiency. This phenomenon, the 'bystander effect', allows the destruction of neighboring tumor cells not transduced with HS-tk. Two other approaches use ex vivo gene transfer of either the IL-2 or antisense insulin-like growth factor type 1 (IGF-1) genes into autologous tumor cells. In animal models, tumor cells genetically altered with antisense IGF-1 or IL-2 genes induce a potent cell-mediated antitumor response. The fifth approach uses the genetic modification of hematopoietic stem cells instead of tumor cells. In this approach, the multiple drug resistance (MDR-1) gene is transferred into stem cells to protect them from the toxic effects of certain chemotherapy drugs. This may allow the administration of higher doses without increasing bone marrow toxicity. Together, these clinical trials will provide critical information needed to develop improved gene transfer technologies for humans and to attain clinical benefit for cancer patients.
 ...
PMID:Gene therapy for malignant neoplasms of the CNS. 897 99

Ha-MDR-IRES-TK is a bicistronic vector that coexpresses the MDR1 gene and the herpes simplex virus thymidine kinase (HSV-TK) gene. In the present study we examined the effect of ganciclovir on MDR1-positive tumors that have been transduced with Ha-MDR-IRES-TK. To establish a human tumor xenograft model of MDR1-transduced recurrent tumors, human KB-3-1 carcinoma cells were transduced with HaMDR or Ha-MDR-IRES-TK, and one each of representative clones, termed KB/MDR and KB/MDR-TK, respectively, were isolated. KB/MDR and KB/MDR-TK showed similar levels of multidrug resistance in vitro. Vinblastine strongly inhibited the growth of the parental KB-3-1 tumors in nude mice but showed little or no effect against KB/MDR-TK tumors. Ganciclovir inhibited the in vivo growth of KB/MDR-TK tumors almost completely under conditions that did not affect the growth of KB-3-1 tumors. Coadministration of vinblastine and ganciclovir inhibited the in vivo growth of KB/MDR-TK premixed with KB-3-1 at any ratio. Long-term, high-level expression of human P-glycoprotein was observed in peripheral blood cells of mice transplanted with Ha-MDR-IRES-TK-transduced bone marrow cells. Ganciclovir eliminated the P-glycoprotein-positive normal blood cells. However, no systemic toxicity was observed. These results clearly demonstrate that it is possible to use ganciclovir to treat MDR1-positive tumors that have been unintentionally transduced with Ha-MDR-IRES-TK. This safety-modified vector should be useful for introducing the MDR1 gene into bone marrow cells to protect normal cells from the toxic effects of cancer chemotherapy.
 ...
PMID:Coexpression of a multidrug resistance gene (MDR1) and herpes simplex virus thymidine kinase gene in a bicistronic retroviral vector Ha-MDR-IRES-TK allows selective killing of MDR1-transduced human tumors transplanted in nude mice. 901 51

A new retroviral vector, pSXLC/pHa, was constructed to coexpress drug-selectable markers with a second gene of interest as a part of a bicistronic mRNA in a retroviral vector using an internal ribosome entry site (IRES) from encephalomyocarditis virus. This system was used to develop a new retroviral vector pHa-MDR-IRES-TK which expresses a single mRNA from which translation of the MDR1 gene is cap dependent and translation of the herpes simplex virus thymidine kinase gene is IRES dependent. The pHa-MDR-IRES-TK transfectants showed high levels of P-glycoprotein expression and multidrug resistance. More than 95% of the vincristine-resistant cells transfected or transduced with pHa-MDR-IRES-TK showed hypersensitivity to ganciclovir, which selects against cells expressing herpes simplex virus thymidine kinase. An amphotropic retrovirus titer of 7.8 x 10(4)/ml was obtained with this vector. This safety-modified vector should be useful for introducing the MDR1 gene into bone marrow cells to protect normal cells from the toxic effects of cancer chemotherapy because this vector allows the elimination of cancer cells that have been unintentionally transduced with the MDR1 vector.
 ...
PMID:Coexpression of a multidrug-resistance gene (MDR1) and herpes simplex virus thymidine kinase gene as part of a bicistronic messenger RNA in a retrovirus vector allows selective killing of MDR1-transduced cells 981 3

It has been proposed that the declining efficiency of antiretroviral agents in human immunodeficiency virus (HIV) infection may also depend on cellular factors at their site of action. Two in particular have been proposed: (i) the defective intracellular metabolism of NRTI in target cells and the altered uptake; and (ii) efflux of nucleoside reverse transcriptase inhibitors (NRTI) and protease inhibitors (PI) by cellular transporter molecules. Several studies have shown that: changes in the activities of various purine and pyrimidine biosynthetic enzymes may occur in lymphocytes of HIV-infected patients; HIV-infected patients on prolonged treatment with nucleoside analogues, e.g. zidovudine, show significantly decreased activity of thymidine kinase (TK) compared with untreated HIV-infected people; and NRTI and PI are substrates for the multidrug membrane transporters. With regard to the latter issue, it is known that the ATP-binding cassette transporter proteins such as the P-glycoprotein (MDR), and the newly discovered family of multidrug resistance-associated proteins (MRP1-6), promote the active extracellular efflux of a wide variety of therapeutics drugs and overexpression of some of them lowers intracellular concentration of PI. In the very near future such mechanisms, also called 'cellular drug resistance', might be taken into account, together with other immunological, virological and behavioural factors, to explain the 'drug failure' and/or the variability of response in HIV patients undergoing antiretroviral treatment.
 ...
PMID:Cellular issues relating to the resistance of HIV to antiretroviral agents. 1500 May 83