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)

To examine the potential of steroid hormones to serve as putative regulators of aortic cell function, we defined hormone receptor content and distribution in intact baboons. Total androgen receptor content in baboon aortic arch, thoracic arch, and abdominal aorta of young mature males was indistinguishable from that of proestrus females. However, 30% to 40% of male aortic androgen receptors were in the nuclear fraction, whereas all aortic androgen receptors of proestrus females were in the cytoplasmic fraction. Cytoplasmic fraction estrogen receptor content of aortic arch and thoracic aorta of intact males was indistinguishable from that of proestrus females. However, cytoplasmic fraction estrogen receptor content of abdominal aorta of proestrus females was significantly greater than that of males. Nuclear fraction estrogen receptors were not detectable in either male or proestrus female baboon aortas. To assess effects of endogenous estrogen on aortic progesterone receptor content, we quantified cytoplasmic fraction progesterone receptors and found that content of proestrus female aortic arch was not significantly different from that of males. However, cytoplasmic fraction progesterone receptor content of thoracic and abdominal aorta of proestrus females was significantly higher than that of males. To determine whether differences in aortic receptor content or distribution were associated with changes in aortic cell function, we quantified the activity of two enzymes of glycosaminoglycan metabolism. Aortic beta-glucuronidase activity was not different in male or proestrus female baboons.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Gender and baboon aortic steroid hormone receptors. 359 71

Lack of tumor selectivity is a severe limitation of cancer chemotherapy. Consequently, reducing dose-limiting organ toxicities such as the cardiac toxicity of doxorubicin (Dox) is of major clinical relevance. Approaches that would facilitate a more tumor-selective anticancer therapy by using nontoxic prodrugs that are converted to active anticancer agents at the tumor site have been the subject of intensive research. One potential method to overcome the cardiac toxicity of Dox is to apply a nontoxic, glucuronide prodrug (HMR 1826) from which Dox is released by the action of beta-glucuronidase, an enzyme present at high levels in many tumors. Using a recently developed, isolated, perfused human lung model, we compared the uptake of Dox into normal lung and lung tumors after a 2.5-h lung perfusion with doxorubicin (n = 8) and with the novel doxorubicin glucuronide prodrug (n = 8). Dox showed a poor uptake into lung tumors as compared with normal lung [mean Dox concentration at the end of perfusion, 1.78 +/- 3.11 (median, 0.66) microg/g versus 22.03 +/- 10.4 (median, 18.5) microg/g; P < 0.001]. However, after perfusion with HMR 1826, the level of Dox in tumor tissue was about 7-fold higher than after perfusion with Dox itself [14.04 +/- 12.9 (median, 12.9) microg/g versus 1.78 +/- 3.11 (median, 0.66) microg/g, P < 0.05, n = 8]. In vitro experiments showed a significantly higher beta-glucuronidase expression and activity in the tumors. The extent of in vitro cleavage of HMR 1826 by homogenized lung tissue was closely related to the content of beta-glucuronidase (r = 0.9834, P < 0.0001). When D-saccharolactone, a specific inhibitor of beta-glucuronidase, was added to the perfusate containing HMR 1826, no accumulation of Dox in lung tissue was seen. These data indicate that the high Dox levels achieved in the tumors with HMR 1826 resulted from cleavage of the prodrug by beta-glucuronidase at the tumor site. Thus, the problem of poor Dox uptake into lung tumors could be circumvented by applying the doxorubicin glucuronide prodrug. Several lines of evidence based on both ex vivo and in vitro results indicate that the approach described using a glucuronide prodrug may be useful in facilitating more selective delivery of chemotherapy to tumors in humans.
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PMID:Enhanced uptake of doxorubicin into bronchial carcinoma: beta-glucuronidase mediates release of doxorubicin from a glucuronide prodrug (HMR 1826) at the tumor site. 919 23

Elucidation of the mechanism enabling tumor selective PMT in vivo with appropriate glucuronyl-spacer-doxorubicin prodrugs, such as HMR 1826, is important for the design of clinical studies, as well as for the development of more selective drugs. Enzyme histochemistry, immunohistochemistry, and the terminal deoxytransferase technique were applied using human cryopreserved cancer tissues, normal human, monkey, and mouse tissues, and human tumor xenografts to examine mechanisms underlying the selectivity of successful PMT with HMR 1826. It could unambiguously be shown by enzyme histochemistry that necrotic areas in human cancers are the sites in which lysosomal beta-glucuronidase is liberated extracellularly in high local concentrations. The cells responsible for the liberation of the enzyme are mainly acute and chronic inflammatory cells, as shown by IHC. Furthermore, it could be demonstrated that beta-glucuronidase liberated in necrotic areas of tumors can activate HMR 1826, resulting in increased doxorubicin deposition in human tumor xenografts or in human lung cancers subjected to extracorporal perfusion, compared to chemotherapy with doxorubicin. Additionally, the doxorubicin load to normal tissues was significantly reduced compared to chemotherapy with doxorubicin. Surprisingly, the increased doxorubicin deposition in tumors also resulted in strong antitumor effects also in cancers resistant to maximum tolerated doses of systemic doxorubicin. Finally, toxicity studies in mice and monkeys revealed an excellent tolerability of HMR 1826, up to a dose of 3 g/m2 (monkeys). These data suggest that HMR 1826 is a promising candidate for clinical development.
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PMID:Elucidation of the mechanism enabling tumor selective prodrug monotherapy. 951 5

A major problem of tumor gene therapy is the low transduction efficiency of the currently available vectors. One way to circumvent this problem is the delivery of therapeutic genes encoding intracellular enzymes for the conversion of a prodrug to a cytotoxic drug which can then spread to neighboring non-transduced cells (bystander effect). One possibility to improve the bystander effect could be the extracellular conversion of a hydrophilic prodrug to a lipophilic, cell-permeable cytotoxic drug. Toward this end, we have used a secreted form of the normally lysosomal human beta-glucuronidase (s-betaGluc) to establish an extracellular cytotoxic effector system that converts an inactivated glucuronidated derivative of doxorubicin (HMR 1826) to the cytotoxic drug. We demonstrate that s-betaGluc-transduced tumor cells convert HMR 1826 to doxorubicin which is taken up by both transduced and non-transduced cells. s-betaGluc in combination with HMR 1826 efficiently induces tumor cell killing both in cell culture and in vivo. This effect is mediated through a pronounced bystander effect of the generated cytotoxic drug. Most notably, this gene therapeutic strategy is shown to be clearly superior to conventional chemotherapy with doxorubicin. Gene Therapy (2000) 7, 224-231.
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PMID:Secreted human beta-glucuronidase: a novel tool for gene-directed enzyme prodrug therapy. 1069 99

Improvement of non-surgical strategies is a pivotal task in the treatment of pancreatic cancer. Response to treatment with most anticancer agents has been very poor, probably due to insufficient drug concentration in tumor tissue. Increased response rates during chemotherapy might be achieved by dose escalation; however, this approach is often hampered by severe side effects. One strategy to overcome these adverse effects is application of nontoxic glucuronide prodrugs from which the active moiety is released by beta-glucuronidase within or near the tumor. The use of glucuronide prodrugs in pancreatic cancer requires increased expression of the enzyme in the diseased tissue, a problem that has not been addressed so far. We therefore investigated function and expression of beta-glucuronidase in tissue samples from human healthy pancreas (n=7) and pancreatic adenocarcinoma (n=8), respectively. Comparing the ability of tissue homogenates to cleave the standard substrate 4-methylumbelliferyl-beta-D-glucuronide, we found a significantly increased specific beta-glucuronidase activity (P<0.05) in pancreatic cancer (median: 133; 75% percentile: 286; 25% percentile: 111 nmol/mg per h) as compared to healthy pancreas (median: 74; 75% percentile: 113; 25% percentile: 71 nmol/mg per h). Enzyme kinetic experiments with the model prodrug N-[4-beta-glucuronyl-3-nitrobenzyloxycarbonyl] doxorubicin (HMR 1826) demonstrated bioactivation of HMR 1826 by pancreatic beta-glucuronidase. Enzymatic activity was found to be closely related to enzyme contents (r=0.87) as assessed by Western blot analysis. Our data indicate that increased beta-glucuronidase activity in pancreatic cancer seems to be due to an elevated steady-state level of the protein. This may be the basis for new therapeutic strategies in treatment of pancreatic carcinoma by using glucuronide prodrugs of anticancer agents.
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PMID:Expression and function of beta-glucuronidase in pancreatic cancer: potential role in drug targeting. 1096 72

A new glucuronylated prodrug of nornitrogen mustard, incorporating the same spacer group as the doxorubicin prodrug HMR 1826, has been prepared. Upon exposure to E. coli beta-glucuronidase, fast hydrolysis occurs but a lower cytotoxicity against LoVo cancer cells is observed compared to the nornitrogen mustard alone. This is explained by cyclization of the intermediate carbamic acid to the inactive chloroethyl oxazolidinone.
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PMID:Synthesis and cytotoxic activity of a glucuronylated prodrug of nornitrogen mustard. 1096 80

HMR 1826 (N-[4-beta-Glucuronyl-3-nitrobenzyl-oxycarbonyl]doxorubicin) is a nontoxic glucuronide prodrug from which active doxorubicin is released by beta-glucuronidase. Preclinical studies aimed at dose optimization of HMR 1826, based on intratumoral pharmacokinetics, are important to design clinical studies. Using an isolated perfused human lung model, the uptake of doxorubicin into normal tissue and tumors after perfusion with 133 microg/ml (n = 6), 400 microg/ml (n = 10), and 1200 microg/ml (n = 6) HMR 1826 was compared. Extracellular tissue pH was measured, and enzyme kinetic studies were performed in vitro to investigate the effect of pH on the formation of doxorubicin. Extracellular pH was lower in tumors than in healthy tissue (6.46 +/- 0.35, n = 8 versus 7.30 +/- 0.33, n = 10; p < 0.001). In vitro, beta-glucuronidase activity was 10 times higher at pH 6.0 than at neutral pH. After perfusion with HMR 1826, there was a linear relationship between HMR 1826 concentrations in perfusate and normal lung tissue. After perfusion with 133, 400, and 1200 microg/ml HMR 1826, the final doxorubicin concentrations in normal and tumor tissue were 2.7 +/- 0.9, 11.1 +/- 5.4, and 21.8 +/- 8.4 microg/g (p < 0.05 for all comparisons), and 0.7 +/- 0.3, 8.6 +/- 2.0 microg/g (p < 0.01 versus 133 microg/g), and 8.7 +/- 4.9 microg/g, respectively. This agrees with the enzyme kinetic observations of saturation of beta-glucuronidase at 400 microg/ml HMR 1826 in the acidic environment of the tumor. Therefore, the escalation of the HMR 1826 dose most likely results in higher circulating concentrations than 400 microg/ml but does not increase the uptake of doxorubicin into tumors and, subsequently, antitumor efficacy. The isolated perfused human lung is an excellent model for preclinical investigations aimed at optimization of tissue pharmacokinetics of tumor-selective prodrugs.
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PMID:Dose optimization of a doxorubicin prodrug (HMR 1826) in isolated perfused human lungs: low tumor pH promotes prodrug activation by beta-glucuronidase. 1190 77

Antibody directed enzyme prodrug therapy (ADEPT) using glucuronide prodrugs is an experimental approach to reduce systemic toxicity of anti-cancer agents. Bioactivation of such prodrugs is achieved by fusion proteins consisting of targeting moieties (e.g. ligands of tumor specific antigens) and human beta-glucuronidase. In order to test a large panel of possible beta-glucuronidase fusion proteins for their applicability in ADEPT, an easy, rapid and high-yield expression system like the baculovirus/insect cell expression system would be needed. A prerequisite for using such fusion proteins is functional and biochemical characterization of human beta-glucuronidase expressed in baculovirus-infected insect cells. Therefore, recombinant human beta-glucuronidase was expressed in Sf9 insect cells and characterized at the protein and functional level. As shown by Western blot analysis the recombinant enzyme consists of dimers with their monomers being linked via disulfide bonds. Posttranslational modifications of the monomers seem to be different as compared with mammalian cells or tissues. The enzyme is functionally active in cleaving the substrates 5-bromo-4-chloro-3-indolyl-beta-D-glucuronic acid, 4-methylumbelliferyl-beta-D-glucuronide and the glucuronide prodrug HMR 1826, respectively, with similar enzyme kinetic parameters as those found in human tissues. Our data demonstrate that beta-glucuronidase expressed in Sf9 cells displays the same enzymatic features as the protein expressed in mammalian cells. Therefore, we suggest that beta-glucuronidase fusion proteins produced in this cell line will be valuable tools for testing a large panel of various targeting moieties in human tumor xenograft models or may be used for ADEPT in man.
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PMID:Expression of active human beta-glucuronidase in Sf9 cells infected with recombinant baculovirus. 1212 9

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