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:3.2.1.23 (beta-galactosidase)
14,648 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Among the appealing features of adenoviruses as vectors for transfer of genes into the central nervous system (CNS) are that they are not neurotoxic, they can accommodate the insertion of several large genes, they are not associated with the hazards of insertional mutagenesis, and they can be concentrated to a high-titer preparation. The authors evaluated the feasibility of using adenovirally mediated gene transfer into cultured human glioma cells and in rat models of solid brain tumors and meningeal cancer. Replication-deficient adenoviral vector particles carrying a nuclear-localizing lacZ gene were injected into established 9L cerebral gliomas in Fischer rats. In addition, the adenoviral vector was injected into the subarachnoid space, either simultaneously with intrathecal tumor inoculation or after establishing leptomeningeal cancer. The brains and spinal cords were removed at various intervals for histochemical evaluation for beta-galactosidase activity using X-Gal staining. Additional rats received a stereotactic intracerebral injection of the vector into normal brain. No clinical abnormalities were observed in the injected rats. Injection of the adenoviral vector into normal brain resulted in diffuse transduction of astrocytes, microglia, neurons, and endothelial cells at the injection site. Injection of a high-concentration vector preparation into cerebral gliomas resulted in effective tumor transduction. Intrathecal injection of the vector in rats with meningeal cancer resulted in transduction of the infiltrating tumor in the subarachnoid space when injections were given simultaneously with, or 7 days after, tumor inoculation. Transduction rates of both solid and leptomeningeal tumors correlated with the number of injected particles. These results suggest that adenoviral vectors can efficiently transduce solid brain tumors and that the vectors survive in the cerebrospinal fluid for a sufficient period of time to allow leptomeningeal tumor transduction. Adenoviral vector should be evaluated for its potential use in therapeutic gene transfer approaches in malignancies of the CNS.
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PMID:Adenovirally mediated gene transfer into experimental solid brain tumors and leptomeningeal cancer cells. 781 37

Factors that influence early events of primary tumor development have been cumbersome to evaluate because of the need to either wait for tumor palpability after experimental manipulation or use of radiolabel to evaluate cell clearance. To facilitate these and similar analyses of cells in vivo, new methods are described that utilize histochemical marker genes to quantitate tumor cell number in a target tissue through the use of luminescent, enzymatic assays for these gene products. 3T3 Cells transfected with either human placental alkaline phosphatase or bacterial lacZ genes were injected subcutaneously into athymic nude mice. Using luminescent substrates designed for marker gene enzymes, extracts from homogenized tumor cell-bearing skins were assayed for the corresponding marker enzyme activities, which were optimized for recovery from skin extracts and correlated to cell number. The homogenization buffer used for these assays was designed to accommodate the optimal and simultaneous recovery of cytosolic beta-galactosidase and membrane-linked alkaline phosphatase from the skin, as well as from cultured cells. These assays provide an inexpensive, sensitive method for quantitatively monitoring the fate of cells genetically tagged with marker genes in various in vivo environments.
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PMID:Quantitation of two histochemical markers in the same extract using chemiluminescent substrates. 781 4

Numerous cell types, including fibroblasts, vascular smooth muscle cells, chondroblasts, monocytes, neutrophils, and several tumor cells express the 67-kD galactolectin, homologous to the alternatively spliced variant of beta-galactosidase. The 67-kD protein resides on the cell surfaces and is capable of interacting with elastin, laminin and collagen type IV. This peripheral membrane protein binds its matrix ligands but only in the absence of galactosugars, whereas binding of galactosugar-containing moieties to its lectin site changes its molecular folding which causes discharge of the ligand and release of the receptor from the cell surface. This review will address the functional significance of the single receptor that interacts with multiple matrix proteins and can be shed from cell surfaces by galactosugars. I will emphasize the role of the 67-kD protein in divergent cellular processes, such as cell-matrix attachment, matrix assembly, cellular chemotaxis, and active migration through the vascular walls.
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PMID:Nature and the multiple functions of the 67-kD elastin-/laminin binding protein. 782 55

Cyclophosphamide and its isomer ifosfamide are cell cycle-nonspecific alkylating agents that undergo bioactivation catalyzed by liver cytochrome P-450 enzymes. The therapeutic efficacy of these oxazaphosphorine anticancer drugs is limited by host toxicity resulting from the systemic distribution of activated drug metabolites formed in the liver. Since tumor cells ordinarily do not have the capacity to activate oxazaphosphorines, we examined whether introduction into tumor cells of a cDNA encoding CYP2B1, a major catalyst of oxazaphosphorine activation, sensitizes the cells to the cytotoxic effects of cyclophosphamide and ifosfamide. Here we show that 9L gliosarcoma cells stably transfected with a cDNA encoding rat CYP2B1 are highly sensitive to cyclophosphamide and ifosfamide cytotoxicity as compared to parental 9L cells or 9L cells transfected with an Escherichia coli beta-galactosidase gene. The CYP2B1 enzyme inhibitor metyrapone protects the CYP2B1-expressing 9L cells from oxazaphosphorine cytotoxicity, demonstrating that the chemosensitivity of these cells is a direct consequence of intracellular prodrug activation. Moreover, CYP2B1-expressing 9L cells potentiate the cytotoxic effects of cyclophosphamide and ifosfamide toward cocultured CYP2B1-negative 9L tumor cells. This "bystander effect" does not require cell-cell contact, and therefore may have the therapeutic advantage of distributing cytotoxic drug metabolites to a wide area within a solid tumor mass. In vivo experiments using Fischer 344 rats implanted s.c. with CYP2B1-expressing 9L tumor cells demonstrated that intratumoral expression of the CYP2B1 gene provides a substantial therapeutic advantage over that provided by liver cytochrome P-450-dependent drug activation alone; cyclophosphamide treatment resulted in complete growth inhibition of CYP2B1-positive tumors, whereas only a modest growth delay effect was obtained with CYP2B1-negative tumors. These studies establish that drug-activating CYP genes may be useful for the development of novel combined chemotherapy/gene therapy strategies for cancer treatment utilizing established cancer chemotherapeutic agents.
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PMID:Intratumoral activation and enhanced chemotherapeutic effect of oxazaphosphorines following cytochrome P-450 gene transfer: development of a combined chemotherapy/cancer gene therapy strategy. 783 28

The dismal results of conventional therapy for primary malignant brain tumors has justified exploring gene therapy approaches for this disease. Transduction of animal brain tumor models in vivo has been reported previously with retroviruses and herpes viruses. Because adenoviruses have the advantage of transducing quiescent and actively dividing tumor cells, they may prove to be more effective in such therapy. We used a replication-deficient recombinant adenovirus bearing the Escherichia coli beta-galactosidase gene in a rat C6 glioma tumor model. Transduced cells were detected by X-5-bromo-4-chloro-3-indolyl beta-D-galactoside staining to reveal beta-galactosidase activity. Initial experiments in vitro showed 50% and 90% transduction at vector titers of approximately 10(7) and 10(8) plaque-forming units/ml, respectively. Although no cytopathic effects were seen at 10(7) plaque-forming units/ml, more than 50% reduction in tumor cell growth was noted at 10(8) plaque-forming units/ml both in vitro and in vivo. Stereotactic delivery of the recombinant adenovirus into the frontal lobe of normal rat brains resulted in intense staining of all cell types, that is, neurons, astrocytes, and ependymal cells. Stereotactic injection into C6 glioma brain tumors in rats stained 25 to 30% of the tumor cells. We conclude that adenovirus vectors can be used to transfer genes to central nervous system tumors in vivo. Using stereotactic delivery, adenovirus vectors can transfer genes into the central nervous system intended for tumor therapy.
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PMID:Stereotactic delivery of a recombinant adenovirus into a C6 glioma cell line in a rat brain tumor model. 783 41

To assess the potential of an in vivo, adenovirus-mediated gene therapy approach for the treatment of malignant melanoma, the efficacy of adenovirus-mediated herpes simplex virus thymidine kinase gene (HSV-Ek) transfer and administration of ganciclovir (GCV) was investigated using a nude mouse model. Initially, B16 murine melanoma cells were efficiently transduced in vitro by a recombinant replication-defective adenovirus containing the HSV-tk gene (ADV/RSVtk), and rendered sensitive to cell killing by 10 micrograms/ml GCV. A significant "bystander effect" was observed at low multiplicity of infection in comparison of cell killing to control B16 transduction by adenovirus containing the beta-galactosidase gene (ADV/RSV-beta-gal). In vivo, melanomas established from subcutaneous injection of 4 x 10(5) B16 cells were injected after 14 d with 1 x 10(10) ADV/RSV-tk viral particles. Subsequent treatment for 6 d with GCV resulted in an inhibition of melanoma growth, with an approximately 40-50% reduction in melanoma volume in comparison to controls in repeated experiments. These data demonstrate that adenovirus-mediated gene transfer can function as an efficient delivery system to reduce established tumor burden in vivo. This result may hold significant promise for the development of effective in situ gene therapy for melanoma in humans.
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PMID:Inhibition of melanoma growth by adenoviral-mediated HSV thymidine kinase gene transfer in vivo. 786 Sep 93

Mutations or loss of the APC tumor-suppressor gene is important for the development of colorectal polyps and cancers, but little is known about the function of this gene in normal tissue. To study the role of APC and other genes in colonocytes in vivo, a system was developed whereby transient expression of genes is established in normal rodent colonic epithelium, using liposomal gene delivery by rectal catheter infusion. Expression of a beta-galactosidase reporter gene and of the human APC gene under a constitutive promoter is demonstrated. A high efficiency of transfection is maintained, with close to 100% of epithelial cells expressing the introduced gene. Expression is transient and does not persist beyond 4 days, consistent with the normal turnover time of gut epithelium, but it can be maintained by repeated treatments. Human APC was expressed for three weeks under these conditions at approximately one-tenth the level of the endogenous APC gene, and no toxicity was observed beyond that attributed to repeated rectal enemas. These results reveal that in vivo expression of exogenous gene is feasible using a liposomal delivery system and suggest a method to further study the physiologic role of APC or other genes in the interrelated process of colonic epithelial proliferation and differentiation.
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PMID:Human APC gene expression in rodent colonic epithelium in vivo using liposomal gene delivery. 787 18

The efficacy of adenovirus (ADV)-mediated gene therapy to treat brain tumors was tested in a syngeneic glioma model. Tumor cells were transduced in situ with a replication-defective ADV carrying the herpes simplex virus thymidine kinase (HSV-tk) gene controlled by the Rous sarcoma virus promoter. Expression of the HSV-tk gene enables the transduced cell to convert the drug ganciclovir to a form that is cytotoxic to dividing cells. Tumors were generated in Fischer 344 rats by stereotaxic implantation of 9L gliosarcoma cells into the caudate nucleus. Eight days later, the tumors were injected either with the ADV carrying the HSV-tk (ADV-tk) gene or a control ADV vector containing the beta-galactosidase (ADV-beta gal) gene and the rats were treated with either ganciclovir or saline. Tumor size was measured 20 days after implantation of 9L cells or at death. Rats treated with ADV-beta gal and ganciclovir or with ADV-tk and saline had large tumors. No tumors were detected in animals treated with ADV-tk and with ganciclovir at doses > or = 80 mg/kg. An infiltrate of macrophages and lymphocytes at the injection site in animals treated with ADV-tk and ganciclovir indicated an active local immune reaction. In survival studies, all animals treated with ADV-tk and ganciclovir have remained alive longer than 80 and up to 120 days after tumor induction whereas all untreated animals died by 22 days. These results demonstrate that ADV-mediated transfer of HSV-tk to glioma cells in vivo confers sensitivity to ganciclovir, and represents a potential method of treatment of brain tumors.
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PMID:Adenovirus-mediated gene therapy of experimental gliomas. 788 26

Murine RSV-M glioma cells were genetically labeled with a retroviral BAG vector carrying the Escherichia coli beta-galactosidase gene. The X-gal-positive stable cell line RSV-M/BAG was obtained by the FDG-FACS method. To examine the behavior of glioma cells in the brain, we homografted RSV-M/BAG cells into the brain of C3H/HeN mice as cell suspensions. Individual grafted glioma cells were easily detected by histochemical staining for B-galactosidase (beta-gal). Three days after grafting, the beta-gal-positive cells were mainly found in the subependymal zone of the lateral ventricle. In addition, some solitary labeled cells were found at locations distant from the injection sites. On the seventh day after implantation, tumor masses were observed and graft-derived glioma cells were migrating bilaterally along the fibers in the corpus callosum. Other labeled cells extended into the brain parenchyma via the perivascular (Virchow-Robin) spaces. Rapid and extensive migration of individual glioma cells was thus clearly demonstrated by intracerebral transplantation of RSV-M/BAG cells.
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PMID:Migration of genetically labeled glioma cells after implantation into murine brain. 793 73

The delivery of therapeutic genes to primary brain neoplasms opens new opportunities for treating these frequently fatal tumors. Efficient gene delivery to tissues remains an important obstacle to therapy, and this problem has unique characteristics in brain tumors due to the blood-brain and blood-tumor barriers. The presence of endothelial mitogens and vessel proliferation within solid tumors suggests that genetically modified endothelial cells might efficiently transplant to brain tumors. Rat brain endothelial cells immortalized with the adenovirus E1A gene and further modified to express the beta-galactosidase reporter were examined for their ability to survive implantation to experimental rat gliomas. Rats received 9L, F98, or C6 glioma cells in combination with endothelial cells intracranially to caudate/putamen or subcutaneously to flank. Implanted endothelial cells were identified by beta-galactosidase histochemistry or by polymerase chain reaction in all tumors up to 35 days postimplantation, the latest time examined. Implanted endothelial cells appeared to cooperate in tumor vessel formation and expressed the brain-specific endothelial glucose transporter type 1 as identified by immunohistochemistry. The proliferation of implanted endothelial cells was supported by their increased number within tumors between postimplantation days 14 and 21 (P = 0.015) and by their expression of the proliferation antigen Ki67. These findings establish that genetically modified endothelial cells can be stably engrafted to growing gliomas and suggest that endothelial cell implantation may provide a means of delivering therapeutic genes to brain neoplasms and other solid tumors. In addition, endothelial implantation to brain may be useful for defining mechanisms of brain-specific endothelial differentiation.
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PMID:Endothelial cell implantation and survival within experimental gliomas. 793 75


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