Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:1.5.1.3 (dihydrofolate reductase)
 5,819 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Altered mouse dihydrofolate reductase gene (DHFRR) was expressed in murine cells using Abelson murine leukemia provirus genome as a prototype vector. A cDNA clone of DHFRR was inserted into a plasmid structure containing retroviral transcriptional as well as packaging signals. The recombinant plasmid was transfected into psi-2 ecotropic cells and the transient virus was used to infect amphotropic PA-12 cells. Recombinant virus (ABL-DHFRR) was detected in the culture medium of transfected PA-12 cells and was free of helper virus. The ABL-DHFRR was capable of conferring methotrexate (MTX) resistance to a variety of cells in culture. The titer of ABL-DHFRR virus was at least tenfold higher than other DHFR retroviruses. The ABL-DHFRR virus titer was increased by selection at increasing concentrations of MTX. The presence of the DHFRR in the virus-infected cells was confirmed by assays which showed reduced inhibition of enzyme activity by MTX. A helper-virus-free, amphotropic, high-titer retrovirus containing the altered DHFR was obtained which may be of use as a dominant selectable marker in infecting hematopoietic progenitor cells.
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PMID:Development of an amphotropic, high-titer retrovirus vector expressing the dihydrofolate reductase gene and conferring methotrexate resistance. 303 Aug 94

The demonstration tha RNA can be cleavaged by cis-ribozyme(catalytic RNAs, RNA enzyme) has potentially important therapeutic implications. Ribozymes are effective for modulation of gene expression because of their simple structure, site-specific cleavage activity and catalytic potential. The targets of ribozyme-mediated gene modulation have ranged from cancer cells to foreign genes that cause infectious diseases. Additional target sites for ribozymes are in initial phases of development and design. Ribozymes have been targeted against myriad genes, including oncogenes (ras, BCR-ABL) and drug resistance genes(MDR-1, c-fos, DHFR). These ribozymes have cleaved the target RNAs in culture system(in vitro) and developed in vivo system. We reported that anti-fos ribozyme has altered the expression of c-fos and DNA repair genes in cisplatin-resistance cancer cells, and reversed the sensitivity to ciaplatin. Furthermore, we have developed high efficiency by the transfer system using an electroporation in vivo.
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PMID:[Circumventing multidrug resistance in human cancer by anti-ribozyme]. 915 62

Chronic myelogenous leukemia (CML) is a malignant disease of the human hematopoietic stem cell caused by the BCR/ABL gene rearrangement. The only curative therapy is allogeneic transplantation. Although autologous transplants may prolong survival, most patients relapse because of disease persisting in the host and in the graft. Continued administration of chemotherapy after transplant could reduce the incidence of relapse provided that the autograft can be protected by transfer of a drug-resistance gene. However, CML autografts will almost certainly contain malignant stem cells that will also be rendered drug-resistant. The presence of the BCR/ABL oncoprotein is necessary and sufficient for malignant transformation seen in CML. We thus hypothesized that transfer of a vector that combines a drug-resistance gene with anti-BCR/ABL antisense (AS) sequences may allow for posttransplant chemotherapy to decrease persistent disease while rendering inadvertently transduced CML stem and progenitor cells functionally normal. We constructed a retroviral vector, LasBD, that combines the methotrexate (MTX)-resistant tyrosine-22 dihydrofolate-reductase (tyr22-DHFR) gene and AS sequences directed at the b3a2 BCR/ABL breakpoint. b3a2 BCR/ABL containing 32D and MO7e cells were transduced with LasBD and selected in MTX for 14 days. Expression of the AS sequences reduced BCR/ABL mRNA and p210(BCR/ABL) protein levels by 6- to 10-fold in most cells. This subsequently led to the restoration of normal function of BCR/ABL cDNA+ cells: they grew significantly slower in the presence of interleukin-3 (IL-3); they underwent apoptotic cell death when cultured without IL-3; and they had restored expression and function of adhesion receptors. These effects were specific, because LasBD-containing AS sequences directed at the b3a2 BCR/ABL breakpoint did not affect p190(BCR/ABL)-containing cells. LasBD also rendered 20% to 30% of primary Ph- and Ph+ CD34(+) cells MTX-resistant and decreased BCR/ABL mRNA levels in MTX resistant Ph+ CD34(+) cells by 10-fold. Expression of the MTX-resistant DHFR gene and the AS sequences has been stable for at least 1 year in vitro and for more than 70 days in vivo. Finally, LasBD decreased tumorigenicity of 32DBCR/ABL cells in vivo by 3 to 4 logs. In conclusion, the tyr22-DHFR gene in the LasBD vector can protect normal hematopoietic cells from MTX-mediated toxicity, whereas the AS sequences in LasBD can suppress expression of the BCR/ABL gene and restore normal function of BCR/ABL cDNA-containing cells. The LasBD vector may therefore prove to be an extremely useful adjunct in autologous transplantation for CML.
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PMID:Gene therapy for chronic myelogenous leukemia (CML): a retroviral vector that renders hematopoietic progenitors methotrexate-resistant and CML progenitors functionally normal and nontumorigenic in vivo. 938 83

Active site mutations that disrupt drug binding are an important mechanism of drug resistance. Computational methods capable of predicting resistance a priori are poised to become extremely useful tools in the fields of drug discovery and treatment design. In this paper, we describe an approach to predicting drug resistance on the basis of Dead-End Elimination and MM-PBSA that requires no prior knowledge of resistance. Our method utilizes a two-pass search to identify mutations that impair drug binding while maintaining affinity for the native substrate. We use our method to probe resistance in four drug-target systems: isoniazid-enoyl-ACP reductase (tuberculosis), ritonavir-HIV protease (HIV), methotrexate-dihydrofolate reductase (breast cancer and leukemia), and gleevec-ABL kinase (leukemia). We validate our model using clinically known resistance mutations for all four test systems. In all cases, the model correctly predicts the majority of known resistance mutations.
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PMID:Efficient a priori identification of drug resistant mutations using Dead-End Elimination and MM-PBSA. 2265 99