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:3.2.1.31 (beta-glucuronidase)
7,680 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The anti-pan carcinoma monoclonal antibody (MAb) 323/A3, linked to E. coli-derived beta-glucuronidase (GUS) was used to study the tumour-site-selective activation of the prodrug Epirubicin-glucuronide (Epi-glu). Epi-glu was isolated from the urine of patients treated with Epirubicin (Epi) by reversed phase chromatography on a silica-C18 column. Epi-glu was stable in human blood and was not converted into Epi by A2780, MCF-7, or OVCAR-3 cancer cells, despite the presence of intracellular GUS. The stability of the prodrug was confirmed in BALB/c mice. MAb 323/A3 and GUS were linked through a stable thioether bond. The conjugate (1:1) was purified by ion exchange and gel filtration chromatography. Binding to target cells revealed an immunoreactivity of at least 60% and good retention of enzyme activity. A protein dye (sulforhodamine B) assay was used to analyse cytotoxicity. Epi (IC50 of 0.003-0.2 microM) was 100-1,000 times more toxic than Epi-glu (IC50 of greater than 20 microM), when cancer cells were exposed for 4 or 24 h to the drugs. The low cytotoxicity of Epi-glu was most likely due to the reduced cellular uptake rate of the prodrug (2.7 pmol 10(-6) cells min-1) as compared to that of the parent compound (25 pmol 10(-6) cells min-1). Pretreatment of antigen-positive cells with the 323/A3-GUS conjugate prior to prodrug exposure completely restored cytotoxicity as a result from hydrolysis of Epi-glu into Epi. Our results demonstrate that the 323/A3-GUS conjugate can specifically activate the stable non-toxic prodrug Epi-glu at the tumour cell level.
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PMID:A monoclonal antibody-beta-glucuronidase conjugate as activator of the prodrug epirubicin-glucuronide for specific treatment of cancer. 152 May 85

The inhibitory effects of N-(4-hydroxyphenyl)retinamide (HPR) and its glucuronide derivative on the growth of MCF-7 human breast cancer cells in vitro were compared. The results indicate that the glucuronide had slightly greater potency and much less cytotoxicity than the free retinoid. At a concentration of 10(-6) M, HPR inhibited MCF-7 cell growth by approximately 25%, whereas an equimolar concentration of the glucuronide caused a 40% growth inhibition. Higher concentrations of HPR were highly cytotoxic. At a 10(-5) M concentration of the glucuronide, cell viability was 77%, and 65% of the cells were able to resume growth. On the other hand, at 10(-5) M HPR, cell viability dropped to 49%, and only 15% of the cells were capable of resuming growth. The lower cytotoxicity and higher potency of the retinoid glucuronide compared to the parent retinamide suggest that the conjugate may have a chemotherapeutic advantage over the parent compound. The apparent higher efficacy of HPR in combination with glucarate (GT) compared to the single agents could be due to increased net formation of HPR glucuronide conjugate following conversion of GT to the beta-glucuronidase inhibitor, D-glucaro-1,4-lactone. However, HPLC analysis of the cell metabolites did not show any detectable levels of the retinoid glucuronide upon treatment of MCF-7 cells with HPR and GT.
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PMID:Growth suppression of human breast carcinoma cells in culture by N-(4-hydroxyphenyl)retinamide and its glucuronide and through synergism with glucarate. 182 40

Platelets contain mitogenic activities for MCF-7 human breast cancer cells when assayed under serum-free chemically defined conditions. Purification from outdated human platelets identified insulinlike growth factor I (IGF-I) as the most potent breast cancer cell mitogen in lysates (Karey KP, Sirbasku DA: see accompanying article, this issue). In this study the release and subcellular localization of IGF-I was investigated. Degranulation of platelets by thrombin treatment caused release of lysosomal enzymes (beta-glucuronidase and N-acetyl-D-glucosaminidase), alpha-granule proteins (beta-thromboglobulin and fibrinogen) as well as mitogenic activity for MCF-7 cells and IGF-I as measured by radioimmunoassay (RIA) and radioreceptor assay. Release of mitogenic activity and immunologically identified IGF-I was induced tenfold over controls by thrombin and was nearly complete as compared to platelets disrupted by repeated freezing and thawing. Disruption of platelets by nitrogen cavitation followed by separation of the organelles by sucrose density gradient sedimentation showed that IGF-I and mitogenic activity localized predominantly to fractions containing alpha-granules rather than soluble cellular components, lysosomes, or dense granules. The morphology of MCF-7 cells in serum-free medium supplemented with supernatants from thrombin-treated platelets also indicated the release of important cell-adhesion factors for human breast cancer cells.
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PMID:Human platelet-derived mitogens. II. Subcellular localization of insulinlike growth factor I to the alpha-granule and release in response to thrombin. 275 54

Metabolism of tamoxifen by rat hepatocytes and hydrolysis of the resulting polar metabolites corresponding to conjugates with beta-glucuronidase gave a major component which was identified as 1-[4-(2-hydroxyethoxy)phenyl]-1-(4-hydroxyphenyl)-2-phenyl-1-butene by comparison of mass spectral properties with those of synthetic material. This compound, which was not observed as a phase I metabolite, is believed to have been found previously in rat bile and in human faeces (metabolite F) but its structure had been incorrectly assigned. Its binding affinity for the estrogen receptor was greater than that of tamoxifen but less than that of 4-hydroxytamoxifen, and it possessed a corresponding degree of antitumour activity against the MCF-7 breast cancer cell line. By carrying out the hepatocyte incubation separately under oxygen and air, it has been shown that the N-oxidation of tamoxifen is favoured by a high concentration of oxygen during in vitro metabolism but that the rate of 4-hydroxylation is not dependent on oxygen availability.
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PMID:Metabolism of tamoxifen by isolated rat hepatocytes. Identification of 1-[4-(2-hydroxyethoxy)phenyl]-1-(4-hydroxyphenyl)-2-phenyl-1-butene and the dependence of N-oxidation on oxygen availability. 357 87

D-Glucarate has shown modest chemopreventive and synergistic chemopreventive effects with retinoids in a number of tumor models as well as a similar antiproliferative effect in MCF-7 human tumor cells in culture. It has been postulated that D-glucarate exerts some of its effects by equilibrium conversion to D-glucarolactone, a potent beta-glucuronidase inhibitor. In the present study, D-glucarate and a number of its analogues, including D-glucarolactone, were evaluated as antiproliferatives in the MCF-7 model with and without added retinoid. Results suggest that the effects of glucarate are reasonably specific for its structure and may not require conversion to glucarolactone.
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PMID:Activity of D-glucarate analogues: synergistic antiproliferative effects with retinoid in cultured human mammary tumor cells appear to specifically require the D-glucarate structure. 819 1

The O-glucuronide analog of N-(4-hydroxyphenyl)retinamide (4-HPROG) has shown a greater chemopreventive activity than the parent N-(4-hydroxyphenyl)retinamide (4-HPR). However, this compound is relatively unstable. In order to improve stability and efficacy, we have prepared a number of stable C-linked analogs of 4-HPROG (C-phenyl and C-benzyl glucuronosyl, glucosyl, and xylosyl analogs). These analogs are stable toward acid hydrolysis and the glucuronosyl analogs resist the actions of beta-glucuronidase. The analogs were prescreened for their antiproliferative potential in vitro using cultured human MCF-7 breast cancer cells. Selected analogs were then evaluated for their ability to inhibit the development and growth of tumors in the 7,12-dimethylbenzanthracene-induced rat mammary tumor model. Although the stable C-linked analogs bound poorly to the nuclear retinoic acid receptors, many showed more potency than the less stable 4-HPROG in inhibiting tumor incidence and multiplicity in vivo. The glucuronide/glucoside analogs are more potent than the xylosides, and the C-benzyl more effective than the C-phenyl analogs. The higher potency of at least two C-linked analogs (retinamidobenzyl glucuronide and retinamidobenzyl glucose) suggests that these analogs may have a chemopreventive advantage over the parent retinamide and its natural O-glucuronide.
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PMID:Chemopreventive activities of C-glucuronide/glycoside analogs of retinoid-O-glucuronides against breast cancer development and growth. 868 25

Trans-resveratrol, a polyphenol present in red wines and various human foods, is an antioxidant also with reported chemopreventive properties. However, whether resveratrol may exert different effects in malignant cells with a common anatomical origin yet displaying different invasive characteristics is not known. Since invasiveness and metastasis are considered to be the most insidious and life-threatening aspects for all cancers, we compared the ability of resveratrol to control growth and cell cycle transition in the highly invasive MDA-MB-435 with the minimally invasive MCF-7 breast carcinoma cells. The data revealed that resveratrol exerted a greater inhibitory effect on the MDA-MB-435 cells. A diminution of percentage of cells in G1 phase and a corresponding accumulation of cells in S phase of the cell cycle was observed. We also studied the effect of resveratrol on a panel of MDA-MB-435 cells transfected with nm23-H1 and nm23-H2 genes, which have been suggested to play a role in controlling metastasis in breast cancer cells. These cells are designated as Vbeta, 1beta, 1Tbeta, 2beta, and 2Tbeta, respectively. The control Vbeta consists of MDA-MB-435 cells transfected with bacterial beta-glucuronidase. Cells labeled 1beta and 1Tbeta correspond to those carrying beta-glucuronidase and overexpressed wild-type (His118) or mutant (Tyr118, catalytically inactive) nm23-H1 genes. The 2beta and 2Tbeta refer to cells transfected with wild-type and mutant nm23-H2 genes. The responses of these cells to resveratrol were assessed by measuring proliferation, cell cycle phase distribution, and changes in expression of several genes. These studies have shown that resveratrol (25 microM, 3 days) reduced growth of all cell types by 60-80%. Overexpression of both wild-type and catalytically inactive nm23-H1 (1beta, 1Tbeta) but not nm23-H2 (2beta, 2Tbeta) reduced the proportion of cells in G1 phase, compared to the Vbeta control cells. Little changes in expression of PCNA, Rb, p53, and bcl-2 were observed in the five cell types treated with resveratrol, compared to untreated cells. Noted exceptions included reduced expression of Rb protein and increased expression of p53 in 2beta and 2Tbeta cells, and increased expression of bcl-2 in 2beta cells, treated with resveratrol. In contrast, resveratrol upregulated expression of cathepsin D by 50-100% in all cell lines except 1beta. These results suggest that the intrinsic metastatic potential of cancer cells may affect their responses to chemopreventive agents such as resveratrol.
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PMID:Cell cycle effects and control of gene expression by resveratrol in human breast carcinoma cell lines with different metastatic potentials. 1040 33

In a previous study of nine human breast-derived cell lines, rates of metabolism of 17beta-estradiol (E(2)) were greatly enhanced when cultures were exposed to the aromatic hydrocarbon receptor agonist, 2,3,7,8-tetrachlorodibenzo-p-dioxin. Elevated rates of E(2) hydroxylation at the C-2, -4, -6alpha and -15alpha positions were observed concomitant with the induction of cytochromes P450 1A1 and 1B1. In each cell line, 2- and 4-hydroxyestradiol (2- and 4-OHE(2)) were converted to 2- and 4-methoxyestradiol (2- and 4-MeOE(2)) by the action of catechol O:-methyltransferase. In this study, conjugation of these estrogen metabolites was investigated. A comparison of the levels of metabolites determined with and without prior treatment of the media with a crude beta-glucuronidase/sulfatase preparation showed that most of the 2-MeOE(2) present was in conjugated form, whereas 4-MeOE(2), 6alpha-OHE(2) and 15alpha-OHE(2) were minimally conjugated. Inhibitor studies suggested that it was the sulfatase activity of the preparation that hydrolyzed the 2-MeOE(2) conjugates in MCF-7 cell media; the presence of 2-MeOE(2)-3-sulfate in MCF-7 culture media was confirmed by electrospray ion-trap mass spectrometry. To identify the enzyme catalyzing this conjugation, the expression of mRNAs encoding five sulfotransferases (SULT1A1, SULT1A2, SULT1A3, SULT1E1 and SULT2A1) was evaluated in the nine cell lines by use of the reverse transcription-polymerase chain reaction. Only expression of SULT1A1 mRNA correlated with the observed conjugation of nanomolar levels of 2-MeOE(2) in these cell lines. Cloning and sequencing of SULT1A1 cDNA from MCF-7 cells revealed that mRNAs encoding two previously identified allelic variants, SULT1A1*1 ((213)Arg) and SULT1A1*2 ((213)His), were expressed in these cells. Heterologous cDNA-directed expression of either variant in MDA-MB-231 cells, which do not normally express SULT1A1, conferred 2-MeOE(2) sulfonation activity. The SULT1A1 allelic variants were also expressed in SF:9 insect cells, from which post-microsomal supernatants were used to determine K:(m) values of 0.90 +/- 0.12 and 0.81 +/- 0.06 microM for SULT1A1*1 and SULT1A1*2, respectively, with 2-MeOE(2) as substrate. These results show that SULT1A1 is an efficient and selective catalyst of 2-MeOE(2) sulfonation and, as such, may be important in modulating the anticarcinogenic effects of 2-MeOE(2) that have been described recently.
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PMID:SULT1A1 catalyzes 2-methoxyestradiol sulfonation in MCF-7 breast cancer cells. 1106 53

Tumor-specific activation of the glucuronide prodrug of doxorubicin, N-[4-doxorubicin-N-carbonyl(oxymethyl)phenyl]-O-beta-glucuronyl carbamate (DOX-GA3), by beta-glucuronidase present in necrotic tumor areas might be improved after transduction of tumor cells to secrete a targeted form of beta-glucuronidase. To that end, we constructed an adenovirus vector, designated Ad/C28-GUSh, encoding human beta-glucuronidase fused to a human single-chain Fv (scFv) against the epithelial cell adhesion molecule (EpCAM), C28, and preceded by a signal sequence for secretion. Antibody specificity and enzyme activity were retained in the fusion protein secreted by tumor cells infected with Ad/C28-GUSh. Diffusion of fusion protein from transduced tumor cells within MCF-7 multicellular spheroids was visualized by immunohistochemistry. Treatment of spheroids with Ad/C28-GUSh and DOX-GA3 resulted in growth inhibition comparable to treatment with doxorubicin alone. Treatment of well-established FMa human ovarian cancer xenografts with intravenous injection of DOX-GA3 (500 mg/kg) resulted in a tumor volume-doubling time of 23.8 days compared to 8.0 days for phosphate-buffered saline (PBS)-treated mice. Intratumoral administration of Ad/C28-GUSh before DOX-GA3 enhanced the growth inhibition and increased the tumor volume-doubling time to 43.1 days (p < 0.01), while virus alone had no effect. Thus, we have successfully shown that an adenovirus vector encoding a secreted, targeted form of human beta-glucuronidase can further improve DOX-GA3 monotherapy.
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PMID:Pronounced antitumor efficacy by extracellular activation of a doxorubicin-glucuronide prodrug after adenoviral vector-mediated expression of a human antibody-enzyme fusion protein. 1501 32

In an effort to develop potent anti-inflammatory and cancer chemopreventive agents, a series of chalcones were prepared by Claisen-Schmidt condensation of appropriate acetophenones with suitable aromatic aldehyde or prepared with appropriate dihydrochalcone reacted with appropriate alkyl bromide or prepared in one-pot procedure involving acetophenone and convenient aromatic aldehyde using ultrasonic agitation on basic alumina. The synthesized products were tested for their inhibitory effects on the activation of mast cells, neutrophils, macrophages, and microglial cells. The potent inhibitors of NO production in macrophages and microglial cells were further evaluated for their in vitro cytotoxic effects against several human cancer cell lines. 2'-Hydroxychalcones 1-3, and 2',5'-dihydroxychalcone 7 exhibited potent inhibitory effects on the release of beta-glucuronidase or lysozyme from rat neutrophils stimulated with formyl-Met-Leu-Phe (fMLP)/cytochalasin B (CB). Two 2'-hydroxychalcones (1 and 3) showed potent inhibitory effects on superoxide anion generation in rat neutrophils in response to fMLP/CB. The previously reported chalcone, 5, 6, and 12, exhibited potent inhibitory effect on NO production in lipopolysaccharide (LPS)/interferon-gamma (IFN-gamma)-activated N9 microglial cells or in LPS-activated RAW 264.7 macrophage-like cells. The potent inhibitors 5, 6, and 12 of NO production in macrophages or microglial cells revealed significant or marginal cytotoxic effects against several human cancer lines. Compound 12 manifested potent selective cytotoxicity against human MCF-7 cells and caused cell death by apoptosis. The present results demonstrated that 1-3, and 7 have anti-inflammatory effects and 5, 6, and 12 are potential anti-inflammatory and cancer chemopreventive agents.
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PMID:Synthetic chalcones as potential anti-inflammatory and cancer chemopreventive agents. 1564 15


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