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:3.5.4.17 (adenosine deaminase)
5,206 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Two new large animal models, non-human primates and fetal sheep, have been developed in an effort to determine the feasibility of using retroviruses for gene therapy. The retroviral vectors N2 and SAX have been used to introduce the genes for neomycin phosphotransferase (neoR, conferring resistance to the antibiotic G418) and human adenosine deaminase (ADA; EC 3.5.4.17), respectively. Varying levels of human ADA activity have been detected in six of the eight SAX-treated monkeys analysed. In the monkey with the greatest activity, human ADA levels approximately 0.5% of endogenous monkey ADA levels were detected. By in situ hybridization, roughly one in 100 bone marrow cells were found to express vector DNA. Sheep have been used for studies of the infectability of fetal blood progenitors in vivo. Blood cells were treated with the N2 vector at the 96th day of gestation, and marrow cells were assayed for the presence of G418-resistant haematopoietic progenitors, starting from one week after birth (62 days after treatment). Up to 33% of colony-forming progenitors were drug resistant initially and, although the proportion of resistant colony-forming units declined, a level of 10% has been found 153 days after transplantation. Human bone marrow has also been treated with the N2 vector, resulting in 1-2% G418-resistant progenitors.
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PMID:Gene therapy: efforts at developing large animal models for autologous bone marrow transplant and gene transfer with retroviral vectors. 332 64

Soon American researchers are expected to request approval for human gene therapy trials in individuals suffering from Lesch-Nyhan syndrome or ADA (adenosine deaminase) deficiency. Walters, who chairs the Working Group on Human Gene Therapy of the National Institutes of Health's Recombinant DNA Advisory Committee, summarizes the ethical dilemmas raised by these new genetic techniques. He addresses the issues of risk assessment, selection of patients, informed consent, and confidentiality, and reviews the framework for ethical evaluation of human gene therapy that has been developed in the United States. Walters also comments on genetic screening for late-onset disorders, the lack of commercial interest in gene therapy, the future possibility of altering human germlines, and the economics of genetic therapy.
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PMID:The ethics of human gene therapy. 345 68

Deficiency of the enzyme adenosine deaminase (adenosine aminohydrolase, EC 3.5.4.4; ADA) leads to severe combined immunodeficiency, a disorder that potentially could be corrected by gene transfer into hematopoietic cells. We have constructed retroviruses containing human ADA cDNA and a dominant selectable marker, a mutated dihydrofolate reductase gene (DHFR*) encoding methotrexate resistance. Human ADA cDNA was inserted alone (DHFR*-ADA) or with a simian virus 40 (SV40) promoter (DHFR*-SVADA). Although NIH 3T3 cells infected with either construct produced human ADA activity, substantially greater levels were attained with DHFR*-SVADA. Infection of murine lymphoid cells in culture with DHFR*-SVADA led to expression of human enzyme at a level well above the mouse endogenous level. ADA activity was also increased after infection of a human ADA-deficient B-cell line. Lethally irradiated mice that were reconstituted with syngeneic marrow infected with the DHFR*-SVADA virus contained unrearranged, integrated proviral DNA in total spleen DNA or in spleen hematopoietic stem cell (CFU-S)-derived colonies. Nevertheless, no human ADA was detectable. RNA analysis showed relatively low and variable expression from the retroviral long terminal repeat, and no detectable expression from the internal SV40 promoter. These data suggest that intrinsic biologic differences exist between cultured cells and CFU-S in vivo.
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PMID:Retrovirus-mediated transfer of human adenosine deaminase gene sequences into cells in culture and into murine hematopoietic cells in vivo. 345 18

A patient with adenosine deaminase-deficient severe combined immunodeficiency is described whose defect is secondary to deletion of a portion of the ADA structural gene. In Southern analyses, DNA from this patient does not hybridize to a genomic probe that includes the 3' end of exon 1. This implies that both his parents are heterozygous for deletions of exon 1 sequences. Consistent with this finding, the patient has no detectable adenosine deaminase mRNA by Northern analysis. This is the first report of a deletion mutation as the cause of adenosine deaminase deficiency.
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PMID:Identification of a deletion in the adenosine deaminase gene in a child with severe combined immunodeficiency. 357 74

This study examined the role of adenosine in regulating coronary arteriolar tone under basal conditions in the normal coronary circulation. Measurements of hemodynamics and flow (microspheres) were made in eight closed-chest, sedated pigs at 1) control and 2) after 10 min of infusion of adenosine deaminase (ADA, 10 U X kg-1 X min-1) into the left anterior descending (LAD) coronary artery. Heart rate was held constant by atrial pacing. Transmural flow in the distal LAD zone did not change versus control (1.81 +/- 0.36) with ADA (1.78 +/- 0.46). However, in comparison with control the distal LAD:circumflex zone transmural flow ratio (0.96 +/- 0.04) declined (P less than 0.01) during ADA infusion (0.93 +/- 0.04). Similarly, the distal LAD:circumflex zone transmural flow resistance ratio increased significantly (P less than 0.01) versus control (1.04 +/- 0.05) in response to intracoronary ADA infusion (1.08 +/- 0.04). Regional myocardial oxygen consumption in the distal LAD zone did not change versus control (16.9 +/- 3.3 3.3 ml X min-1 X 100 g-1) during ADA (16.9 +/- 4.5). Additional studies in 15 open-chest swine given intracoronary infusion of ADA demonstrated that 1) the enzyme penetrates the interstitial fluid (ISF) and 2) attains ISF levels which are adequate to reduce basal adenosine concentration 10 fold even if substantial increase in adenosine production occurs in response to ADA. Thus, since destruction of adenosine by ADA caused only very modest relative reduction in regional flow, it is likely that the nucleoside plays only a limited role in regulation of arteriolar tone under basal conditions in the normal coronary circulation.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Adenosine's role in regulating basal coronary arteriolar tone. 371 57

The possibility that the mutant mouse wasted (wst/wst) may serve as an animal model for studies of severe combined immunodeficiency disease (SCID) and the role of adenosine deaminase (ADA, EC 3.5.4.4) in adenosine metabolism were investigated. The specific activity of ADA in wst/wst compared with control mice was significantly lower by 26% in thymus, but significantly higher by 18% in spleen and 32% in cerebellum. Vmax values of ADA in spleens were 43% higher in wst/wst mice and no changes were observed in Km values. In contrast, the Vmax of ADA was unchanged in erythrocytes from wst/wst mice, but the Km for adenosine was significantly elevated. Thus, based on ADA measurements alone, it may be premature to consider wst/wst mice as a model for ADA deficiency and SCID in humans.
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PMID:Lack of adenosine deaminase deficiency in the mutant mouse wasted. 378 Sep 80

Human adenosine deaminase (ADA; adenosine aminohydrolase, EC 3.5.4.4) was expressed at high levels in cultured mouse cells using a transmissable murine retrovirus vector system. A cDNA clone encoding ADA has been inserted into a plasmid vector containing retroviral transcription and packaging signals as well as a selectable gene for G418 resistance. The constructions were transfected into psi 2 cells, which package the recombinant retroviral genomes into replication-defective virus particles. Isoenzyme analysis for ADA in G418-selected psi 2 cells showed at least 20-fold more human ADA activity than endogenous mouse ADA activity. A mouse T-cell lymphoma line, BL/VL3, was cocultured with transformed psi 2 cells producing human ADA, and some of the cocultured cells were selected for resistance to G418. Both G418-selected and unselected cocultured cells expressed human ADA activity at 25%-50% the level of the endogenous enzyme. Thus, efficient retroviral transduction of ADA expression was obtained.
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PMID:Expression of human adenosine deaminase using a transmissable murine retrovirus vector system. 385 23

Six mouse monoclonal antibodies (mAbs) defining separate systems of cell surface antigens of cultured human renal cancer were tested for reactivity with normal fetal and adult tissues and with neoplastic tissues. Five of the mAbs identified glycoproteins of Mr 160,000 (designated S4), Mr Mr 140,000 (F23), Mr 120,000 (S23 and S27), and Mr 115,000 (S22). The glycoprotein component of Mr 120,000 has been shown recently to be the adenosine deaminase binding protein (ADA-BP) and mAbS23 and mAbS27 define two distinct epitopes on ADA-BP. S22 was not detected on any normal fetal or adult tissues but was found on a subset of renal cancers. S4, F23, S23, and S27 defined distinct domains of the nephron: glomerulus (S4), proximal tubules (S4, F23, S23, and S27), and portions of Henle's loop (S23 and S27). mAbS4 also reacted with the interstitial matrix in the renal medulla and of other tissues, and mAbF23 reacted with fetal and adult fibroblasts. The S23 epitope of ADA-BP was expressed by placental trophoblasts and epithelial cells of breast, prostate, lung, and colon, whereas the S27 epitope was detected on a more limited range of cell types (trophoblasts and prostate epithelium). A panel of 20 renal cell carcinomas was typed for expression of these antigens; 7 phenotypes could be distinguished, with the S4+/F23+/S23+/S27+/S22+ or - phenotype (15 cases) being most common. The other antigenic system, V1, identified a heat-stable antigen that was widely expressed on cultured cell types but showed a restricted pattern of reactivity in tissues. V1 expression was limited to the adrenal cortex, Leydig cells, and the theca of ovarian follicles, and to adrenal cortical carcinomas.
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PMID:Specificity analysis of mouse monoclonal antibodies defining cell surface antigens of human renal cancer. 385 26

The human adenosine deaminase cDNA has been cloned in a lambda-vector. Contained within a sequence of over 1500 nucleotides is an open reading frame of 1089 nucleotides that encodes the amino acids of ADA. The functional ADA gene contains at least six kilobases and has at least two introns. Using in vitro translation, molecular hybridization to ADA cDNA, and S1 nuclease mapping, ADA mRNA has been characterized in lymphoblast lines from seven different ADA-deficient children. All of the lines contain substantial amounts of RNA, which hybridizes specifically to the ADA cDNA. Four of the cell lines contain translatable mRNAs with small defects such as single base substitutions that are not detectable by S1 mapping. Deficiency of ADA activity in these lines appears secondary to synthesis of structurally altered proteins containing simple amino acid substitutions. Three of the lines contain mRNAs with S1 nuclease detectable defects. Some or all of these defective mRNAs are postulated to result from anomalous RNA processing. In these cases the causes of the ADA deficiency may be more complex than simple amino acid substitutions in the protein and could include small insertions or deletions of amino acids as well as changes in the efficiency of translation of the mRNAs.
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PMID:Molecular biology of the adenosine deaminase gene and messenger RNA. 386 73

Cloned cDNA sequences of human adenosine deaminase (ADA; adenosine aminohydrolase, EC 3.5.4.4) have been isolated from a cDNA library constructed in bacteriophage lambda gt10. The cDNA for the library was prepared from poly(A)+ RNA isolated from a human T-lymphoblast cell line, CCRF-CEM. The library was initially screened by differential plaque hybridization to labeled cDNA prepared from human T- and B-lymphoblast cell lines with a 21-fold difference in levels of translatable ADA mRNA. Two recombinants containing cloned cDNA sequences for ADA were identified by hybridization-selected translation. Both recombinants contained approximately 1,600 base pairs of inserted human DNA. Restriction maps of the two inserts were not identical. One contained approximately 40 base pairs of additional DNA toward the center of the cDNA. The cloned cDNA specifically hybridized to five fragments generated by HindIII digestion of human genomic DNA. It also hybridized to human lymphoblast RNA species 1.6 and 5.8 kilobases in length. The cDNA was used as a probe to estimate ADA mRNA levels in human lymphoblast cell lines. ADA mRNA levels correlate closely with levels of ADA catalytic activity and ADA protein in cell lines containing structurally normal ADA. A leukemic T-lymphoblast line produced 6 to 9 times as much ADA protein and ADA mRNA as transformed B-lymphoblast lines. Two mutant B-lymphoblast lines from patients with hereditary ADA deficiency contained unstable ADA protein but had 3 to 4 times the normal level of ADA mRNA.
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PMID:Cloning of cDNA sequences of human adenosine deaminase. 620 Aug 75


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