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:1.6.5.2 (NQO1)
6,196 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

NAD(P)H:quinone oxidoreductase 1 (NQO1) is a flavoenzyme that catalyzes two-electron reductive metabolism and detoxification of quinones and their derivatives leading to protection of cells against redox cycling and oxidative stress. To examine the in vivo role of NQO1, a NQO1-null mouse was produced using targeted gene disruption. Mice lacking NQO1 gene expression showed no detectable phenotype and were indistinguishable from wild-type mice. However, NQO1-null mice exhibited increased toxicity when administered menadione compared with wild-type mice. These results establish a role for NQO1 in protection against quinone toxicity. The NQO1-null mice are a model for NQO1 deficiency in humans and can be used to determine the role of this enzyme in sensitivity to toxicity and carcinogenesis.
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PMID:Disruption of the DT diaphorase (NQO1) gene in mice leads to increased menadione toxicity. 951 35

We investigated two of the major proposed modes of action of the benzoquinoid ansamycin geldanamycin using a pair of human colon-carcinoma cell lines, BE and HT29. One potential mechanism of action in colorectal cancer is the inhibition of c-Src kinase activity, since this proto-oncogene is hyperexpressed in human large-bowel tumours. Our results show that despite the 9-fold higher level of c-Src kinase activity found in HT29 cells, there was only a 1.4-fold difference in cytotoxicity as compared with BE cells, the latter being the most sensitive. Moreover, even at concentrations of geldanamycin that resulted in cell kill of 80% or more after a 24-h period of exposure, there was no effect on c-Src kinase activity in HT29 cells, although c-Src protein was depleted at supralethal levels of exposure. We also investigated the metabolism of the quinone moiety of geldanamycin by DT-diaphorase, an enzyme that activates certain quinone antibiotics such as mitomycin C and is hyperexpressed in colorectal cancer cells. Geldanamycin was shown to be a substrate for DT-diaphorase present in HT29 cells. However, the lack of a major differential in cytotoxicity between HT29 and BE cells indicates that this is unlikely to be pharmacologically significant, since the former contains high levels of enzyme activity, whereas BE cells have no significant activity due to a point mutation in the DT-diaphorase (NQO1) gene. Although reduction of geldanamycin was also catalysed by non-DT-diaphorase reductases in HT29 and BE cells, providing the potential for free radical induction, this is unlikely to be significant since studies previously reported by us elsewhere showed that cells exposed to geldanamycin exhibited no evidence of DNA damage. Thus, as far as the mode of action of geldanamycin in human colon-carcinoma cells is concerned, the present results rule out two major possibilities, namely, the involvement of c-Src tyrosine kinase inhibition and DT-diaphorase metabolism.
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PMID:Geldanamycin-induced cytotoxicity in human colon-cancer cell lines: evidence against the involvement of c-Src or DT-diaphorase. 952 39

Maintenance of cellular homeostasis is a critical survival trait when cells are exposed to electrophilic chemicals. Because conjugation and elimination of these toxins is dependent upon sequential and coordinated metabolic pathways, acquired resistance through a gradual adaptive response would rarely be expected to be the consequence of changes in one gene product. Human HT29 colon cancer cells chronically exposed to EA have acquired resistance to the drug. Commensurate with resistance, EA is more effectively conjugated to GSH and effluxed from the resistant cells. Using directed and random (differential display) approaches, a number of detoxification and/or protective gene products have been shown to be expressed at elevated levels. These include gamma-GCS (approximately 3-fold), GST-pi (approximately 3-fold), MRP (approximately 3-fold), NQO1 (approximately 3-fold), DDH (20-fold), and SSP 3521, a transcriptional regulator (approximately 3-fold). Multiple mechanisms contribute to these increases, including enhanced transcriptional rate and prolonged mRNA and protein half lives. Further indications for the involvement of transcriptional regulators is found in HL60 adriamycin-resistant cells which overexpress MRP, GST-pi and gamma-GCS and also have 15-20-fold more DNA-dependent protein kinase. It is possible that this enzyme serves as an early stress response gene which may activate downstream transcription factors. Intriguingly, the catalytic subunit of DNA-dependent protein kinase has a high avidity for [35S]azidophenacyl-GSH. High levels of GSH conjugates indicate cell stress and it would seem reasonable to speculate that DNA-dependent protein kinase may serve as a receiver and transmitter of signals which contribute to drug resistance and maintain cell viability.
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PMID:Coordinate changes in expression of protective genes in drug-resistant cells. 967 55

Carcinogens present in cigarette smoke and diet have been associated with pancreatic cancer. We hypothesized that heterocyclic and aromatic amines implicated in these exposures could be involved as causative agents and that therefore genetic variation in enzymes metabolizing these carcinogens could modify the risk of developing malignant and benign pancreatic disease. The effect of the genetic polymorphism of acetyltransferases (NAT1) and NAT2), glutathione S-transferase M1 (GSTM1) and NAD(P)H: quinone oxidoreductase 1 (NQO1) on the risk of pancreatic diseases (cancer, pancreatitis) was examined in a case-control study. PCR-based assays were used for genotype analysis of genomic DNA from whole blood cells. Samples collected from Caucasian patients with diagnosed pancreatic cancer (n = 81), with non-alcoholic (n = 41) and alcoholic pancreatitis (n = 73) and from asymptomatic control subjects (n = 78) were analysed. The prevalence of GSTM1 null genotype and of NAT2 fast and slow acetylator genotypes and the distribution of frequencies for NQO1 genotypes did not differ in subjects with pancreatic diseases vs controls. For NAT1 slow acetylators a non-significant excess (P = 0.18) was found among pancreatic cancer cases vs controls. There was a significant over-representation of the GSTM1 AB or B genotype in all pancreatic disease cases combined (OR = 2.6; P < 0.05). When concurrent controls were pooled with literature controls (n = 1427), OR was 1.4 (P = 0.08). The results of this study, requiring confirmation, suggest that the polymorphism of GSTM1 and NAT1 enzymes may be associated with a modest increase in susceptibility to pancreatic diseases.
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PMID:Genetic polymorphism of N-acetyltransferases, glutathione S-transferase M1 and NAD(P)H:quinone oxidoreductase in relation to malignant and benign pancreatic disease risk. The International Pancreatic Disease Study Group. 988 82

Benzo(a)pyrene and benzene are human carcinogens. The metabolic activation of these compounds into ultimate mutagenic and carcinogenic metabolites is prerequisite for their carcinogenic effects. In this report, the mutagenicity and carcinogenicity of hydroquinones of benzo(a)pyrene and benzene was investigated to address two important questions: (1) do hydroquinones contribute to benzo(a)pyrene and benzene carcinogenicity; and (2) how safe is it to increase the levels of NAD(P)H:quinone oxidoreductase 1 (NQO1), a key enzyme in the generation of hydroquinone. The supF tRNA of the plasmid pSP189 was used as the mutational target in a cell-free and Chinese hamster ovary (CHO) cell system to study hydroquinone mutagenicity. RNA and protein-free pSP189 DNA was incubated in a cell-free system with benzo(a)pyrene-3,6-quinone and purified NQO1 or with benzoquinone hydroquinone to generate adducted pSP189 DNA. The adducted pSP189 DNA was transfected in human embryonic kidney cells Ad293. In the CHO cell system, monolayer cultures of CHO cells and CHO cells overexpressing NQO1 or P450 reductase were transfected with pSP189 vector DNA, treated with benzo(a)pyrene-3,6-quinone. The adducted and replicated pSP189 DNA was rescued from transfected Ad293 (cell-free system) and CHO cells (CHO cell system), digested with the restriction enzyme Dpn1 to remove unreplicated DNA followed by transformation in Escherichia coli MBM7070. The mutant colonies [white/pale blue on 5-bromo-4-chloro-3-indolyl beta-D-galactoside/isopropyl beta-D-thiogalactoside (X-gal/IPTG) plates] were selected, regrown and analysed by DNA sequencing. Mutagenesis experiments demonstrated that hydroquinones cause sequence-specific frameshift mutations involving deletion of a single cytosine from the DNA sequence 5'-172-CCCCC176-3' or a single guanosine from the complementary strand sequence 5'-GGGGG-3' in the supF tRNA gene. This mutation was specific to the hydroquinones, as it was not observed with quinones and other components of the redox cycling (semiquinones and reactive oxygen species). Exposure of BALBc/3T3 cells to hydroquinones resulted in cellular transformation leading to the loss of contact inhibition and regulation of cell growth. The transformation efficiency of BALBc/3T3 cells exposed to hydroquinones was significantly increased by the tumour promoter 12-O-tetradecanoylphorbol-13-acetate (TPA), indicating that hydroquinones are excellent initiators that require additional co-carcinogens or promoters to exert an effect. The hydroquinone + TPA as well as hydroquinone-transformed BALBc/3T3 cells, when injected s.c. in severe combined immunodeficient (SCID) mice, produced tumours at 100% frequency. These results establish that hydroquinones lead to mutagenicity and carcinogenicity.
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PMID:Hydroquinones cause specific mutations and lead to cellular transformation and in vivo tumorigenesis. 970 76

Genomic DNA for genetic analyses has traditionally been derived from blood samples. With the availability of PCR techniques requiring only minute amounts of DNA and the current demand for high-volume testing, a less invasive, simpler to perform, and cheaper method to obtain DNA is desirable. We developed a method to obtain high-quality genomic DNA from buccal cells that has high acceptability and allows for a large number of PCR assays from a single sample. Sixty subjects vigorously swished 10 ml of undiluted commercial mouthwash in the mouth for 60 s and expelled the liquid into a collection container. DNA was isolated from the buccal cells with a rapid method using proteinase K digestion, phenol-chloroform extraction, and ethanol precipitation. Electrophoretic analysis of the extracted DNA showed detectable levels of high molecular weight genomic DNA in all samples. The DNA yields ranged from 0.2 to 134.0 microg, for an average of 49.7 microg. Using these samples, all 60 subjects were successfully genotyped by PCR-based assays for polymorphisms in the CYP1A1 (MspI and exon 7), CYP2E1 (RsaI), GSTM1, GSTT1, and NQO1 genes, confirming that the quality of DNA isolated from mouthwash samples was sufficient to reliably support PCR amplification. Storage of the (unprocessed) specimens at room temperature or at 37 degrees C for 1 week (temperature conditions that may be encountered when mailing samples) or at -20 degrees C for at least 6 months did not affect the DNA yield or ability to PCR amplify the samples. The results suggest that this mouthwash procedure may be suitable for large community-based studies of genetic susceptibility to disease in which samples can be collected by the participants themselves, mailed back to the study center, and stored for months prior to DNA analysis.
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PMID:A simple mouthwash method for obtaining genomic DNA in molecular epidemiological studies. 971 25

NAD(P)H:quinone oxidoreductase (NQO1) catalyses the two-electron reduction of quinone compounds. NQO1 is involved in the reductive bioactivation of cytotoxic antitumour quinones such as mitomycin C, but also plays a protective role against the carcinogenicity and mutagenicity of quinones, their precursors and metabolites. Three alleles have been identified in the human population: the functional Arg139/Pro187 allele (which we have termed NQO1*1); the nonfunctional allele Arg139/Ser187 (NQO1*2) and the Trp139/Pro187 allele (NQO1*3), which is associated with a diminished activity. We applied polymerase chain reaction-based genotyping assays to characterize interethnic variability in the frequency of NQO1 alleles in Caucasian (n = 575), Canadian Native Indian (n = 110), Canadian Inuit (n = 83) and Chinese (n = 86) populations. The NQO1*2 allele was found at significantly higher frequencies in Chinese (0.49) and Native North American populations (Inuit 0.46; Canadian Native Indians 0.40) compared with Caucasians (0.16). The NQO1*3 allele was not observed in Inuit individuals, and occurred at a lower frequency than the NQO*2 allele in Caucasians (0.05), Chinese (0.04) and Canadian Native Indians (0.01). Our results predict that a greater proportion of Orientals and related ethnic groups lack, or have reduced, NQO activity relative to Caucasians. Affected individuals may not only exhibit resistance to quinone-based cancer therapy because of a decreased production of cytotoxic drug metabolites, but may also be more susceptible to toxicities associated with toxicants.
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PMID:NAD(P)H:quinone oxidoreductase: polymorphisms and allele frequencies in Caucasian, Chinese and Canadian Native Indian and Inuit populations. 973 17

We have previously reported that antiestrogens stimulate quinone reductase (NAD(P)H:(quinone-acceptor) oxidoreductase (QR or NQO1); EC 1.6.99.2) enzymatic activity, an action that may provide protective effects against the toxicity and mutagenicity caused by quinones. We have now investigated the transcriptional regulation of the QR gene by antiestrogens. In transfection experiments employing the 5'-flanking (863-base pair) region of the human QR gene promoter with its electrophile/antioxidant response element (EpRE/ARE) or deleted or mutated constructs, we observe that antiestrogens induced an increase in QR gene promoter reporter activity in estrogen receptor (ER) negative breast cancer and endometrial cancer cells transfected with ER, and this induction by antiestrogens was repressed by estradiol. The stimulation of QR transcriptional activity required the 31-base pair electrophile-responsive region from the human QR gene promoter and a functional ER. Intriguingly, antiestrogens were stronger activators of the QR EpRE via the ER subtype ERbeta than ERalpha. Oligonucleotide gel mobility and antibody shift assays reveal that the ER binds to the EpRE but is only a minor component of the proteins bound to the EpRE in ER-containing MCF-7 breast cancer cells. While binding of ERbeta to the estrogen response element was weaker when compared with ERalpha, ERbeta and ERalpha showed similar binding to the EpRE. Together these findings provide evidence that QR gene regulation by the antiestrogen-occupied ER is mediated by the EpRE-containing region of the human QR gene and indicate that the ER is one of the complex of proteins that binds to the EpRE. In addition, that ERbeta is a more potent activator at EpRE elements than is ERalpha suggests that the different levels of these two receptors in various estrogen target cells could impact importantly on the antioxidant potency of antiestrogens in different target cells. These findings have broad implications regarding the potential beneficial effects of antiestrogens since EpREs mediate the transcriptional induction of numerous genes, including QR, which encode chemoprotective detoxification enzymes.
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PMID:Transcriptional regulation of the human quinone reductase gene by antiestrogen-liganded estrogen receptor-alpha and estrogen receptor-beta. 973 13

Transient transfection studies of human HepG2 and mouse Hepa hepatocarcinoma cells with a reporter gene construct regulated by a human antioxidant responsive element (ARE) from the NQO1 gene demonstrated that the element is responsive to low oxygen conditions. The antioxidant N-acetyl L-cysteine (NAC) strongly inhibited basal aerobic reporter gene activity in HepG2 cells without obviously affecting the hypoxic induction, as is consistent with ARE sensitivity to oxidative stress in aerobic cultures. Electrophoretic mobility shift (EMS) assays of nuclear extracts of HepG2 and Hepa cells lysed under aerobic or hypoxic conditions or after exposure to the phenolic compound 3-(2)-tert-butyl-4-hydroxyanisole (BHA), showed specific and constitutive protein binding to the ARE under all of these conditions. Taken together, these findings show that the ARE can mediate gene expression in response to low oxygen conditions. Co-ordinately regulated expression of ARE-dependent genes, such as phase II detoxification enzymes, may be an important phenotype of solid tumors containing significant regions of pathophysiological hypoxia.
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PMID:The redox-sensitive human antioxidant responsive element induces gene expression under low oxygen conditions. 974 25

NAD(P)H:quinone oxidoreductase (NQO1) is a flavoenzyme that catalyzes the two-electron reduction of quinones and related compounds. With the use of biochemical assays, NQO1 has been shown to be overexpressed in many types of cancer, including non-small cell lung cancer (NSCLC). NQO1 can bioactivate antitumor quinones such as mitomycin C, and new quinone-based drugs are currently being developed to target this enzyme in tumors such as NSCLC. Because there is no information on the cell-specific expression of NQO1 in lung, the purpose of this study was to examine the expression of NQO1 in human NSCLC, small cell lung cancer, carcinoid lung tumors, and normal lung using immunohistochemistry. A high level of NQO1 protein expression was detected by immunohistochemistry in NSCLC (adenocarcinoma, squamous cell carcinoma, and bronchoalveolar carcinoma), but no NQO1 protein could be detected in small cell lung cancer or carcinoid lung tumors. In addition, NQO1 protein expression was examined by immunohistochemistry in normal lung tissue. A high level of NQO1 protein expression was detected by immunohistochemistry in normal lung respiratory epithelium, with the highest levels of expression observed in ciliated columnar epithelial cells. Significant amounts of NQO1 protein were also detected in the vascular endothelium and adipocytes. These data demonstrate that NQO1 is overexpressed in NSCLC. Cells in normal lung also contain marked NQO1 protein and may be damaged by drugs activated by NQO1. These data validate NSCLC as a target for NQO1-directed agents and suggest that the potential for lung toxicity be considered in the preclinical development of quinone-based antitumor drugs.
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PMID:Immunohistochemical detection of NAD(P)H:quinone oxidoreductase in human lung and lung tumors. 974 20


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