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)

The telomerase complex is responsible for telomere maintenance and represents a promising cancer therapeutic target. We describe herein the antitelomerase and antitumor properties of a small-molecule compound designed by computer modeling to interact with and stabilize human G-quadruplex DNA, a structure that may form with telomeric DNA, thereby inhibiting access to telomerase. The 3,6,9-trisubstituted acridine 9-[4-(N,N-dimethylamino)phenylamino]-3,6-bis(3-pyrrolodinopropionamido) acridine (BRACO19) represents one of the most potent cell-free inhibitors of human telomerase yet described (50% inhibitory concentration of 115 +/- 18 nM). Moreover, in contrast to G-quadruplex interactive agents described previously, BRACO19 did not cause nonspecific acute cytotoxicity at similar concentrations to those required to completely inhibit telomerase activity. There exists a 90-fold differential (mean 50% inhibitory concentration for acute cell kill across seven human tumor cell lines of 10.6 +/- 0.7 microM). The exposure of 21NT human breast cancer cells, which possess relatively short telomeres, to nonacute cytotoxic concentrations of BRACO19 (2 microM) resulted in a marked reduction in cell growth after only 15 days. This was concomitant with a reduction in intracellular telomerase activity and onset of senescence as indicated by an increase in the number of beta-galactosidase positive-staining cells. Intraperitoneal administration of nontoxic doses of BRACO19 (2 mg/kg) to mice bearing advanced stage A431 human vulval carcinoma subcutaneous xenografts and previously treated with paclitaxel induced a significant increase in antitumor effect compared with that observed with paclitaxel alone. BRACO19 thus represents the first of a "second generation" of G-quadruplex-mediated telomerase/telomere-interactive compounds. It possesses nanomolar potency against telomerase but low nonspecific cytotoxicity, growth inhibitory effects, and induction of senescence in a human breast cancer cell line and, moreover, significant antitumor activity in vivo when administered post paclitaxel to mice bearing a human tumor xenograft carcinoma.
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PMID:A G-quadruplex-interactive potent small-molecule inhibitor of telomerase exhibiting in vitro and in vivo antitumor activity. 1196 Nov 34

Elevated insulin-like growth factor binding protein-related protein 1 (IGFBP-rP1) mRNA in senescent human mammary epithelial cells suggested that the IGFBP-3 gene product may inhibit cell proliferation. To test this hypothesis, we used a retroviral vector to express IGFBP-rP1 cDNA in the IGFBP-rP1-deficient MCF-7 breast cancer cell line. Compared with control vector-transduced cells, cumulative cell numbers for IGFBP-rP1-transduced polyclonal or clonal cell cultures were reduced by 39 and 74%, respectively, after 1 week. Medium conditioned by IGFBP-rP1-producing cultures reduced cumulative cell numbers in parental MCF-7 cultures by 20% compared with medium from cultures of a control vector-transduced cell line. Nuclear fragmentation analysis and cell proliferation assays completed in the presence of the pan-caspase inhibitor benzyloxycarbonyl-Val-Ala-Asp(OMe)-fluoromethylketone excluded apoptosis as the responsible mechanism. The percentage of cells containing senescence-associated beta-galactosidase activity was doubled compared with control cell cultures. Flow cytometry analysis indicated that twice as many noncycling cells were present in the IGFBP-rP1-transduced MCF-7 cell cultures compared with controls. These findings indicate that IGFBP-rP1 is an inhibitor of MCF-7 breast cancer cell proliferation and may act via a cellular senescence-like mechanism.
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PMID:Insulin-like growth factor binding protein-related protein 1 inhibits proliferation of MCF-7 breast cancer cells via a senescence-like mechanism. 1206 44

Direct experimental evidence implicates telomere erosion as a primary cause of cellular senescence. Using a well characterized model system for breast cancer, we define here the molecular and cellular consequences of adriamycin treatment in breast tumor cells. Cells acutely exposed to adriamycin exhibited an increase in p53 activity, a decline in telomerase activity, and a dramatic increase in beta-galactosidase, a marker of senescence. Inactivation of wild-type p53 resulted in a transition of the cellular response to adriamycin treatment from replicative senescence to delayed apoptosis, demonstrating that p53 plays an integral role in the fate of breast tumor cells treated with DNA-damaging agents. Stable introduction of hTERT, the catalytic protein component of telomerase, into MCF-7 cells caused an increase in telomerase activity and telomere length. Treatment of MCF-7-hTERT cells with adriamycin produced an identical senescence response as controls without signs of telomere shortening, indicating that the senescence after treatment is telomere length-independent. However, we found that exposure to adriamycin resulted in an overrepresentation of cytogenetic changes involving telomeres, showing an altered telomere state induced by adriamycin is probably a causal factor leading to the senescence phenotype. To our knowledge, these data are the first to demonstrate that the mechanism of adriamycin-induced senescence is dependent on both functional p53 and telomere dysfunction rather than overall shortening.
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PMID:Adriamycin-induced senescence in breast tumor cells involves functional p53 and telomere dysfunction. 1210 Nov 84

We have examined whether inhibition of phosphatidylinositol-3 kinase (PI3K) and its target, the serine/threonine kinase Akt, play a role in the antitumor effect of the HER2 antibody Herceptin. Herceptin inhibited colony formation, down-regulated cyclin D1, and increased p27 protein levels in the HER2 gene-amplified BT-474 and SKBR-3 human breast cancer cells. These effects were temporally associated with the inhibition of PI3K activity in vitro as well as Akt function as measured by steady-state levels of phospho-Ser473 Akt and kinase activity against glycogen synthase kinase (GSK)-3beta. These responses were not observed in MDA-361 and MDA-453 cells, which do not exhibit HER2 gene amplification and are relatively resistant to Herceptin. Treatment of BT-474 cells with Herceptin inhibited the constitutive tyrosine phosphorylation of HER3 and disrupted the basal association of HER3 with HER2 and of HER3 with p85alpha potentially explaining the inhibition of PI3K. Treatment with either Herceptin or the PI3K inhibitor LY294002 increased the levels of p27 in the nucleus>cytosol, thus increasing the ratio of p27:Cdk2 in the nucleus and inhibiting Cdk2 activity and cell proliferation. Antisense p27 oligonucleotides abrogated the increase in p27 induced by Herceptin and prevented the antibody-mediated reduction in S phase. Transduction of BT-474 cells with an adenovirus-encoding active (myristoylated) Akt (Myr-Akt), but not with a beta-galactosidase control adenovirus, prevented the Herceptin- or LY294002-induced down-regulation of cyclin D1 and of phosphorylated GSK-3beta and prevented the accumulation of p27 in the nucleus and cytosol. In addition, Myr-Akt prevented Herceptin-induced inhibition of the cell proliferation of BT-474 cells and Herceptin-induced apoptosis of SKBR-3 cells. These data suggest that (a) changes in cell cycle- and apoptosis-regulatory molecules after HER2 blockade with Herceptin result, at least in part, from the inhibition of Akt; and (b) disabling PI3K and Akt is required for the antitumor effect of HER2 inhibitors.
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PMID:Herceptin-induced inhibition of phosphatidylinositol-3 kinase and Akt Is required for antibody-mediated effects on p27, cyclin D1, and antitumor action. 1212 52

Orthotopic or intracardiac injection of human breast cancer cell lines into immunocompromised mice allows study of the molecular basis of breast cancer metastasis. We have established a quantitative real-time PCR approach to analyze metastatic spread of human breast cancer cells inoculated into nude mice via these routes. We employed MDA-MB-231 human breast cancer cells genetically tagged with a bacterial beta-galactosidase (Lac-Z) retroviral vector, enabling their detection by TaqMan real-time PCR. PCR detection was linear, specific, more sensitive than conventional PCR, and could be used to directly quantitative metastatic burden in bone and soft organs. Attesting to the sensitivity and specificity of the PCR detection strategy, as few as several hundred metastatic MDA-MB-231 cells were detectable in 100 microns segments of paraffin-embedded lung tissue, and only in samples adjacent to sections that scored positive by histological detection. Moreover, the measured real-time PCR metastatic burden in the bone environment (mouse hind-limbs, n = 48) displayed a high correlation to the degree of osteolytic damage observed by high resolution X-ray analysis (r2 = 0.972). Such a direct linear relationship to tumor burden and bone damage substantiates the so-called 'vicious cycle' hypothesis in which metastatic tumor cells promote the release of factors from the bone which continue to stimulate the tumor cells. The technique provides a useful tool for molecular and cellular analysis of human breast cancer metastasis to bone and soft organs, can easily be extended to other cell/marker/organ systems, and should also find application in preclinical assessment of anti-metastatic modalities.
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PMID:Correlation between extent of osteolytic damage and metastatic burden of human breast cancer metastasis in nude mice: real-time PCR quantitation. 1219 65

Adjuvant hyperthermia can improve treatment outcome for locally recurrent breast cancer (LRBC). Previously, we demonstrated that infection of human breast cancer cells with a recombinant adenovirus expressing beta-galactosidase from the human hsp70b gene promoter (Ad.70b.betagal) results in 50- to 800-fold increases in reporter gene expression following heat treatment (30 minutes at 43 degrees C). Here, we describe a heat-directed suicide gene therapy strategy based on an adenoviral vector (Ad.70b.CDTK) in which expression of the dual prodrug-activating E. coli cytosine deaminase/herpes simplex virus thymidine kinase (CDTK) fusion gene is under the control of the hsp70b promoter. Treatment of T47D and MCF-7 breast cancer cells with mild hyperthermia (43 degrees C/30 minutes) and prodrugs (100 microg/ml 5-fluorocytosine and 10 microg/ml ganciclovir) following infection with Ad.70b.CDTK (10-100 PFU/cell) resulted in 30- to 60-fold decreases in clonogenic survival relative to control cultures treated with heat or prodrugs alone. Clonogenic survival declined even further (up to 240-fold) following heat treatment at 41.5 degrees C for 120 minutes. A decreased clonogenic survival was accompanied by tumor cell apoptosis. These results demonstrate that this combined treatment strategy can be highly effective against heat- and radiation-resistant breast tumor cells and supports the continued development of heat-directed CDTK suicide gene therapy strategies for LRBC.
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PMID:Heat-directed suicide gene therapy for breast cancer. 1267 2

The tumor suppressor p53 is the most frequently mutated gene in human tumors. In response to DNA damage, aberrant growth signals, or chemotherapeutic drugs, p53 is stabilized and induces apoptosis and/or cell cycle arrest. While the mechanisms of p53-dependent apoptosis are not well understood, p53-dependent cycle arrest is primary mediated by the CDK inhibitor p21. p53 is a transcriptional activator and it is not surprising that a majority of p53 mutations occur in the core DNA binding domain and affect DNA binding and transactivation of p53 targets in tumors. We used the capability of p53 to activate transcription for developing a new assay that permits rapid determination of the status of p53 in cancer cell lines of different origin. Our strategy involved using a retrovirus containing a p53-regulated lacZ reporter gene that was introduced into colon and breast tumor cell lines to determine p53 status. Simple staining for beta-galactosidase allowed us to confirm that the colon cancer cell lines LIM1215 and HCT116, as well as the breast cancer cell line MCF7. have wild-type p53, and the colon cancer cell line Caco-2 as well as breast cancer cell lines MDA-MB-435 and MDA-MB-231 have mutant p53. This method may be applied to novel cell lines of any origin with unknown status of p53.
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PMID:A new method for determining the status of p53 in tumor cell lines of different origin. 1272 31

The hypothesis that cross-talk between membrane-active beta-adrenergic agonists and estrogens includes beta-adrenergic modulation of estrogen receptor (ER)-regulated gene expression was investigated. Vascular smooth muscle-derived A7r5 cells were transfected with an ERalpha expression plasmid (pCR3.1-hERalpha), the estrogen response element (ERE)-linked reporter pERE-E1b-luciferase (ERE-Luc), and pCMV-beta-galactosidase using a lysine-conjugated adenovirus transfection method. Hormone or agonist treatment and harvest followed 6 hours and 24 hours later, respectively. Treatment with 17beta-estradiol (E(2), 1 nmol/L) significantly stimulated ERE-Luc activity. Isoproterenol (10-9 to 10-6 mol/L) treatment alone did not stimulate ERE-Luc activity. Cotreatment with both E(2) and isoproterenol resulted in complete inhibition of E(2)-stimulated ERE-Luc activity. This isoproterenol effect was prevented by the beta-adrenergic antagonist propanolol (10-6 mol/L). Adrenomedullin treatment in these cells (1-50 nmol/L) did not inhibit ER/ERE-Luc activity, whether in the presence or absence of E(2). Moreover, isoproterenol did not affect vitamin D-stimulated VDRE-Luc expression, indicating that the inhibitory effect of isoproterenol on E(2)-directed ERE-Luc expression is specific among nuclear transcription factor receptors. Moreover, in MCF-7 breast cancer cells, there was no effect of isoproterenol on ER/ERE-directed transcription in the absence or presence of E(2), demonstrating tissue specificity of this isoproterenol effect. These studies demonstrate cross-talk between the beta-adrenergic agonist isoproterenol and ER-directed reporter gene expression in A7r5 cells. Furthermore, this cross-talk is specific with respect to agonist, nuclear receptor species, and cell type. These observations may have important implications both for the use of beta-adrenergic agents to treat hypertension and for possible gender-related differences in cardiovascular regulation.
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PMID:Cross-talk between beta-adrenergic stimulation and estrogen receptors: isoproterenol inhibits 17beta-estradiol-induced gene transcription in A7r5 cells. 1288 32

The action of electromagnetic fields (EMF) on different pathways related to cell physiology, proliferation, toxicity of chemicals, gene expression, etc., are currently being investigated although the results are still not conclusive and even conflicting. In laboratory and animal studies, EMF has been found to produce a great variety of effects such as: increase in ornithine decarboxylase activity in breast, increase in beta-galactosidase gene expression and oncogene transcription after exposure to 50/60 Hz. Animal studies have shown that the use of EMF can enhance drug delivery across biological barriers (rat abdominal skin), using benzoic acid as the drug candidate. It has been reported by different authors that pulsed EMF (PEMF) can produce alterations in antineoplastic drugs potency. In the present study, we investigated the effects of PEMF on methotrexate cytotoxicity in MCF-7 breast cancer cells and the effects with simultaneous exposure to FeCl3. The data presented in the current report indicate that PEMF (25 Hz, 1.5 mT) do not induce modulation of the action of methotrexate (with and without iron-III) in MCF-7 cells when they are exposed to PEMF for 2 h/day during 3 days.
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PMID:Methotrexate cytotoxicity on MCF-7 breast cancer cells is not altered by exposure to 25 Hz, 1.5 mT magnetic field and iron (III) chloride hexahydrate. 1289 13

Tamoxifen has contributed to a dramatic reduction in breast cancer mortality and recent results indicate that aromatase inhibitors may further improve survival in some patients. Nevertheless, a substantial proportion of patients become resistant to treatment. To date, with the exception of estrogen receptor (ER) determination by ligand binding or immunohistochemical techniques, there has been no way of predicting which of several therapies is indicated in particular patients. We describe a novel assay using the adenoviral gene delivery system to assess ER function in breast cancer cells derived directly from patients. The purification and short-term culture of these cells has been recently described by our laboratory. Adenovirus containing an estrogen-regulated beta-galactosidase reporter gene (ERE-lacZ) was constructed and used to test ER activity in breast cancer cells derived from 18 patients with primary and 16 patients with metastatic cancer, under varying treatment schedules. The adenoviral assay enabled ER activity to be readily determined in purified cells from primary breast cancers and secondary sites. Breast cancers cells could be categorized on the basis of ER activity in the absence of ligand, the presence of estrogen or anti-estrogens. In primary breast cancers, our results correlated with ER determination by immunohistochemistry in 78% of cases. In patients who had become resistant to tamoxifen, however, we found some in whom reporter activity was stimulated by tamoxifen and others whose tumors were either still estrogen responsive or completely unresponsive, irrespective of the original ER content. Our findings indicate that this reporter assay could be useful in decisions regarding use of adjuvant endocrine therapies in breast cancer.
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PMID:Reporter gene assay demonstrates functional differences in estrogen receptor activity in purified breast cancer cells: a pilot study. 1456 18


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