Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:3.5.1.4 (deaminase)
 5,113 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

To study the possibility of oral gene therapy using live attenuated Salmonella, eukaryotic expression vectors EGFPN1, pLCDSN were introduced into a live attenuated AraA(-) auxotrophic mutant of Salmonella typhimurium (SL3261) and were administered orally to BALB/c and C57BL/6 mice. After six weeks, these mice were challenged with 4T(1) and Lewis cancer cells. Until the tumors reached to about 10 mm in diameter, 5-fluorocytosine was given through intraperitoneal injection. Flow cytometry, confocal microscopy and PCR methods were used to detect the integration and expression of the genes. The inhibition of the tumor and the survival time of the mice were also investigated. Results showed that cytosine deaminase gene integration could be detected in almost all kinds of mice tissue. And the GFP expression was much stronger in spleen and tumor than in other tissues. Cytosine deaminase/5-fluorocytosine system had significant antitumoractivities in vivo. The anti-tumor activities of cytosine deaminase/5-fluorocytosine at 500 mg/kg on 4T(1) and Lewis carcinoma in BALB/c and C57BL/6 mice were more potent than the efficiency of 5-fluorouracil 10 mg/kg(P 0.05). Therefor, this experiment demonstrates the potential value of live attenuated Salmonella as carrier for oral gene therapy.
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PMID:Treatment of Tumor in Mice by Oral Administration of Cytosine Deaminase Gene Carried in Live Attenuated Salmonella. 1205 Aug 17

Uracil in DNA results from deamination of cytosine, resulting in mutagenic U : G mispairs, and misincorporation of dUMP, which gives a less harmful U : A pair. At least four different human DNA glycosylases may remove uracil and thus generate an abasic site, which is itself cytotoxic and potentially mutagenic. These enzymes are UNG, SMUG1, TDG and MBD4. The base excision repair process is completed either by a short patch- or long patch pathway, which largely use different proteins. UNG2 is a major nuclear uracil-DNA glycosylase central in removal of misincorporated dUMP in replication foci, but recent evidence also indicates an important role in repair of U : G mispairs and possibly U in single-stranded DNA. SMUG1 has broader specificity than UNG2 and may serve as a relatively efficient backup for UNG in repair of U : G mismatches and single-stranded DNA. TDG and MBD4 may have specialized roles in the repair of U and T in mismatches in CpG contexts. Recently, a role for UNG2, together with activation induced deaminase (AID) which generates uracil, has been demonstrated in immunoglobulin diversification. Studies are now underway to examine whether mice deficient in Ung develop lymphoproliferative malignancies and have a different life span.
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PMID:Uracil in DNA--occurrence, consequences and repair. 1248 10

Post-transcriptional RNA editing generates novel gene products by changing the coding sequence of the transcript from that in the genome. Two classes of RNA editing exist in mammals, each of which involves an enzymatic deamination. These reactions have stringent sequence and structural requirements for their target RNAs, and each requires distinctive enzymatic machinery. Alterations in the expression or abundance of RNA-editing factors produce unanticipated alterations in the processing or expression of RNAs, in some cases outside their physiological targets. Recent findings suggest that unregulated expression of the cytidine-deaminase gene family might lead to deamination of deoxycytidine nucleotides in DNA. Aberrant or dysregulated RNA editing, or altered expression of editing factors, might contribute to genomic instability in cancer.
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PMID:Hydrolytic nucleoside and nucleotide deamination, and genetic instability: a possible link between RNA-editing enzymes and cancer? 1272 40

Functional antibody genes assembled by V(D)J joining are subsequently diversified by somatic hypermutation, gene conversion and class-switch recombination. Recent evidence indicates that all three processes are caused by the deamination of cytosine to uracil at sites within the immunoglobulin (Ig) loci, with the pattern of diversification depending on the pathway used for resolving the initiating dU-dG lesion. Whereas DNA deamination targeted to the endogenous Ig locus triggers a program of somatic gene diversification that underpins adaptive immunity, deamination targeted to foreign DNA might have arisen initially as a form of innate immunity. Furthermore, the observation that members of the DNA deaminase family can target inappropriate genes suggests they might also contribute to mutations during genome evolution, as well as in cancer.
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PMID:Immunity through DNA deamination. 1282 2

Soft tissue and bone sarcomas of the extremities can be difficult to eradicate, and standard treatment may require limb amputation. New therapies to decrease tumor size could improve the effectiveness of treatment and decrease the frequency of limb amputation. Cytosine deaminase (CD)-based gene therapy has been shown to be effective in decreasing growth of solid tumors when animals with CD-expressing tumor cells are treated with 5 fluorocytosine (5FC), an inert prodrug that is converted to 5-fluorouracil (5FU) by CD. In this investigation, we used a novel CD-containing fusion gene to determine whether CD-based gene therapy affected soft tissue or bone sarcomas. The novel fusion gene (NGFR-CD) encodes for a protein with extracellular and transmembrane domains of human nerve growth factor receptor (NGFR) and cytoplasmic CD. Murine 2472 (2) sarcoma cells were transduced with fusion genes containing either the bacterial (NGFR-(b)CD) or yeast (NGFR-(y)CD) CD gene. 5FC treatment killed NGFR-(b)CD- and NGFR-(y)CD-transduced sarcoma cells in vitro through direct and bystander effects (P < 0.01). In contrast, 5FC treatment of mice with s.c. 2NGFR-(b)CD or 2NGFR-(y)CD tumors affected only 2NGFR-(y)CD tumors. 5FC had no effect on growth of NGFR-(b)CD tumors but caused significant decrease in the size of 2NGFR-(y)CD tumors (51 +/- 60 versus 938 +/- 767 mm(3), treated versus control, P < 0.01). Evaluation of bystander killing in vivo revealed significant tumor killing, with a 5-fold reduction in s.c. tumor volume evident in saline versus 5FC-treated mice when tumors were comprised of 90% 2472 cells and 10% 2NGFR-(y)CD selected for fluorescence-activated cell sorting (P < 0.01). Bone sarcomas were eliminated in 9 of 10 5FC-treated mice, compared with 11.8 +/- 6.0 mm(2) in saline-treated mice (P < 0.002). In addition, 5FC treatment of bone sarcomas caused a significant reduction in cancer-induced bone destruction (P < 0.002) and resulted in a reduction in the number of osteoclasts. Finally, 5FC treatment had no effect on animal weight or survival, whereas doses of 5FU providing equivalent tumor reduction as 5FC resulted in treatment-associated deaths and significant weight loss (P < 0.001).
Cancer Res 2003 Oct 15
PMID:Direct and bystander killing of sarcomas by novel cytosine deaminase fusion gene. 1458 82

Adenosine modulates the proliferation, survival and apoptosis of many different cell types, ranging from epithelial, endothelial and smooth muscle cells, to cells of the immune and neural lineages. In this review, we critically discuss the available in vitro and in vivo data which support a role for adenosine in both development-associated apoptosis, and in diseases characterized by either pathologically increased cell death (e.g., ischemia, trauma and aging-associated neurodegeneration) or abnormally reduced spontaneous apoptosis (e.g., cancer). Particular emphasis is given to the possible role of extracellular adenosine receptors, since these may represent novel and attractive molecular targets for the pharmacological modulation of apoptosis. In some instances, adenosine-induced cell death has been demonstrated to require entry of the nucleoside inside cells; however, in many other cases, activation of specific adenosine extracellular receptors has been demonstrated. Of the four G protein-coupled adenosine receptors so far identified, the A2A and the A3 receptors have been specifically implicated in modulation of cell death. For the A3 receptor, results obtained by exposing both cardiomyocytes and brain astrocytes to graded concentrations of selective agonists suggest induction of both cell protection and cell death. Such opposite effects, which likely depend on the degree of receptor activation, may have important therapeutic implications in the pharmacological modulation of cardiac and brain ischemia. For the A2A receptor, recent intriguing data suggest a specific role in immune cell death and immunosuppression, which may be relevant to both adenosine-deaminase-immunodeficiency syndrome (a pathology characterized by accumulation of adenosine to toxic levels) and in tumors where induction of apoptosis via activation of specific extracellular receptors may be desirable. Finally, preliminary data suggest that, in a similar way to the adenosine-deaminase-immunodeficiency syndrome, the abnormal accumulation of adenosine in degenerative muscular diseases may contribute to muscle cell death. Although the role of adenosine receptors in this effect still remains to be determined, these data suggest that adenosine-induced apoptosis may also represent a novel pathogenic pathway in muscular dystrophies.
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PMID:Adenosine-induced cell death: evidence for receptor-mediated signalling. 1463 82

DNA mismatch repair (MMR) defects bring about a strong mutator phenotype and microsatellite instability (MSI). In an attempt to exploit MSI in cancer therapy, we constructed expression vectors carrying a thymidine kinase/blasticidin deaminase fusion gene downstream from a (C)(12) or an (A)(26) microsatellite and stably transfected these constructs into human cells in which the MMR status could be regulated by doxycycline. We now show that ganciclovir-resistant clones arising through frameshifts in the (C)(12) microsatellite were 20 times more frequent in cells in which MMR was inactivated. This difference may be exploited in gene therapy of tumors with MSI, which represent a substantial proportion of cancers of many different tissues.
Cancer Res 2003 Dec 01
PMID:Differential killing of mismatch repair-deficient and -proficient cells: towards the therapy of tumors with microsatellite instability. 1467 62

A 56-year-old man was admitted to our hospital because of bilateral pleural effusion. Computed tomography revealed solitary mediastinal lymphadenopathy, splenomegaly and a small amount of ascites. No lung parenchymal lesion was seen. Although lymphocyte predominance without atypia and a high adenocine deaminase concentration in the pleural fluid were compatible with tuberculous pleurisy, no mycobacteria could be detected either with Ziehl-Nielsen stain or with PCR. Because the serum soluble interleukin 2 receptor (sIL-2 R) level was unexpectedly high (> 8,000 U/ml), and a level not previously reported in benign diseases, we performed thoracoscopy- and mediastinoscopy-assisted biopsies, both of which eventually confirmed the diagnosis of tuberculosis. After a 4-drug anti-tuberculous regimen was initiated, pleural effusion and ascites subsided, with a marked decrease in the sIL-2R level. This case indicates that in tuberculous pleurisy, serum sIL-2R can rise to a level suggestive of hematological malignancies, it and also illustrates the validity of thoracoscopy-assisted pleural biopsy in such situations.
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PMID:[A case of tuberculosis pleuritis with high serum soluble IL-2 receptor]. 1500 22

Suicide gene therapy of cancer is a method whereby cancerous tumors can be selectively eradicated while sparing damage to normal tissue. This is accomplished by delivering a gene, encoding an enzyme capable of specifically converting a nontoxic prodrug into a cytotoxin, to cancer cells followed by prodrug administration. The Escherichia coli gene, codA, encodes cytosine deaminase and is introduced into cancer cells followed by administration of the prodrug 5-fluorocytosine (5-FC). Cytosine deaminase converts 5-FC into cytotoxic 5-fluorouracil, which leads to tumor-cell eradication. One limitation of this enzyme/prodrug combination is that 5-FC is a poor substrate for bacterial cytosine deaminase. The crystal structure of bacterial cytosine deaminase (bCD) reveals that a loop structure in the active site pocket of wild-type bCD comprising residues 310-320 undergoes a conformational change upon cytosine binding, making several contacts to the pyrimidine ring. Alanine-scanning mutagenesis was used to investigate the structure-function relationship of amino acid residues within this region, especially with regard to substrate specificity. Using an E. coli genetic complementation system, seven active mutants were identified (F310A, G311A, H312A, D314A, V315A, F316A, and P318A). Further characterization of these mutants reveals that mutant F316A is 14-fold more efficient than the wild-type at deaminating cytosine to uracil. The mutant D314A enzyme demonstrates a dramatic decrease in cytosine activity (17-fold) as well as a slight increase in activity toward 5-FC (2-fold), indicating that mutant D314A prefers the prodrug over cytosine by almost 20-fold, suggesting that it may be a superior suicide gene.
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PMID:Alanine-scanning mutagenesis reveals a cytosine deaminase mutant with altered substrate preference. 1524 53

Cytosine deaminase (CD) is currently being used as a suicide gene for cancer gene therapy. The premise of this therapy is the preferential deamination of 5-fluorocytosine (5FC) to 5-fluorouracil by cancer cells expressing cytosine deaminase. However, a lack of efficient gene transfer to tumors combined with inefficient 5FC turnover currently limits the clinical applications of this gene therapy approach. We have used random mutagenesis to create novel bacterial cytosine deaminases that demonstrate an increased preference for 5FC over cytosine. Among the 15 mutants isolated, one conferred sensitivity to Escherichia coli in a negative selection system at a concentration of 5FC that was 10-fold lower than a sublethal dose for wild-type CD. Evaluation of individual substitutions found in this double mutant (Q102R, D314G) demonstrated that the substitution at residue D314 was solely responsible for the observed increase in sensitivity to 5FC. Additional mutagenesis at D314 resulted in the identification of two more substitutions with the ability to confer enhanced 5FC sensitivity to E.coli. Structure determinations of the three CD variants in the presence and absence of a transition state 5FC analogue provide insights to the determinants of substrate binding specificity at the 5' position of the pyrimidine ring. CD mutant D314A is a promising candidate for further gene therapy studies.
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PMID:Random mutagenesis and selection of Escherichia coli cytosine deaminase for cancer gene therapy. 1538 61


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