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: UMLS:C0596263 (carcinogenesis)
64,820 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Evidence from both experimental carcinogenesis and studies in human cirrhotic liver suggest that defective repair of the promutagenic DNA base lesion, O6-methylguanine, is a factor in the multistep process of hepatocellular carcinogenesis. Ubiquitous environmental alkylating agents such as N-nitroso compounds can produce O6-methylguanine in cellular DNA. Unrepaired, O6-methylguanine can lead to the formation of G --> A transition mutations, a known mechanism of human oncogene activation and tumour suppressor gene inactivation. Combined treatment of rodents with an agent producing O6-methylguanine in DNA, and an agent promoting cell proliferation, leads to development of hepatic nodules and hepatocellular carcinoma (HCC), cell division, hence DNA replication, being required for the propagation of tumorigenic mutation(s) in hepatocyte DNA. The paramount importance of O6-methylguanine in hepatocellular carcinogenesis is indicated by the observation that transgenic mice engineered to have increased hepatic levels of repair enzyme O6-methylguanine-DNA methyltransferase (MGMT) are significantly less prone to hepatocellular carcinogenesis following alkylating agent treatment. Cirrhosis is a universal risk factor for development of human HCC, and a condition that is characterized by increased hepatocyte proliferation as a result of tissue regeneration. Levels of the human repairing enzyme for O6-methylguanine were found to be significantly lower in cirrhotic liver than in normal tissue. In accord with findings from animal models, this suggested a mechanism in which persistence of O6-methylguanine due to defective DNA repair by MGMT, together with increased hepatocyte proliferation, might lead to specific gene mutation(s) and hepatocellular carcinogenesis. Screening for the presence and persistence of O6-methylguanine in human DNA presently involves formidable technical difficulty. Indications are that such limitations might be overcome by the use of an ultrasensitive method such as immuno-polymerase chain reaction (PCR). This approach should allow parallel measurement of DNA adduct and repair enzyme in routine liver biopsy samples. It might also enable investigation of O6-methylguanine in human genes specifically associated with hepatocellular carcinogenesis. Given the wide variation in human MGMT levels observed between individuals, tissues, and cells, this technology should be adapted to permit the ultrasensitive localisation and measurement of adducts and repairing enzyme in liver biopsy tissue sections. Ability to ultrasensitively measure O6-methylguanine, and its repair enzyme, should prove valuable in the risk assessment of cirrhotic patients for developing hepatocellular carcinoma.
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PMID:Repair of DNA lesion O6-methylguanine in hepatocellular carcinogenesis. 993 84

Our studies of DNA damage and repair in autoimmune disease, lymphomagenesis, and carcinogenesis, require identification of an immunoassay approach that is capable of ultrasensitive detection in a routine human tissue biopsy of several physicochemically diverse antigens, some of which will be present at very low level. Immuno-polymerase chain reaction (immuno-PCR) is a recently described method for ultrasensitive antigen detection that combines the amplification power of PCR with a method similar to a standard antibody capture, enzyme-linked immunosorbent assay (ELISA). As a test of the universality of immuno-PCR, and as an assessment of the suitability of this method for our studies, we used a single immuno-PCR protocol to assay purified forms of the following physicochemically diverse antigens: oligomeric pyruvate dehydrogenase complex (PDC; Mr 8.5 x 10(6)), the promutagenic DNA base adduct O(6)-methylguanosine (Mr 298) and its monomeric repair enzyme, O(6)-methylguanine-DNA methyltransferase (MGMT; Mr 22,000), and a peptide from the N-terminus of MGMT (Mr 2310). We found that all antigens could be ultrasensitively assayed using the single immuno-PCR protocol. Assay limits observed using antigen-specific (primary) antibodies at 1 microg/ml, were in the approximate range of 10(2)-10(9) molecules, with O(6)-methylguanosine being detected most sensitively. Sensitivity of the antigen assay appeared to positively correlate with primary antibody titres determined by ELISA. Furthermore, we observed a substantial increase in detection sensitivity for all antigens by the use of primary antibodies at the higher level of 10 microg/ml. The latter approach permitted antigen assay within the approximate range of 10(0)-10(7) molecules. The combination of higher titre primary antibodies and their use at higher input level, produced an increase of immuno-PCR assay sensitivity of up to four orders of magnitude greater than those previously reported through the use of this assay to measure other antigens. This represents up to a nine order of magnitude increase in immunoassay sensitivity compared to ELISA. Our findings provide compelling evidence that immuno-PCR is indeed a universal ultrasensitive antigen detection method. Using the indicated assay enhancements. immuno-PCR performed as detailed here can offer greatly increased sensitivity for antigen measurement compared to other methods. Thus, our findings suggest that parallel quantitation of several different antigens in very small samples of human tissue will be readily attainable using immuno-PCR.
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PMID:Enhanced ultrasensitive detection of structurally diverse antigens using a single immuno-PCR assay protocol. 1003 37

Human methylguanine-DNA methyltransferase (MGMT) transgenic mice expressing high levels of O6-alkylguanine-DNA alkyltransferase (AGT) in lung were crossbred to A/J mice that are susceptible to pulmonary adenoma to study the impact of O6-methylguanine (O6mG)-DNA adduct repair on NNK-induced lung tumorigenesis. Expression of the chimeric human MGMT transgene in lung was identified by northern and western blot analysis, immunohistochemistry assay and enzymatic assay. AGT activity was 17.6 +/- 3.2 versus 1.2 +/- 0.4 fmol/microg DNA in lung of MGMT transgenic mice compared with non-transgenic mice. Immunohistochemical staining with anti-human AGT antibody showed that human AGT was expressed throughout the lung. However, some epithelial cells of bronchi and alveoli did not stain for human AGT, suggesting that the human MGMT transgene expression was heterogeneous. After 100 mg/kg NNK i.p. injection in MGMT transgenic mice, lung AGT activity remained much higher and levels of lung O6mG-DNA adducts in MGMT transgenic mice were lower than those of non-transgenic mice. In the tumorigenesis study, mice received 100 mg/kg NNK at 6 weeks of age and were killed 44 weeks later. Ten of 17 MGMT transgenic mice compared with 16 of 17 non-transgenic mice had lung tumors, P < 0.05. MGMT transgenic mice had lower multiplicity and smaller sized lung tumors than non-transgenic mice. Moreover, a reduction in the frequency of K-ras mutations in lung tumors was found in MGMT transgenic mice (6.7 versus 50% in non-transgenic mice). These results indicate that high levels of AGT expressed in mouse lung reduce lung tissue susceptibility to NNK-induced tumorigenesis due to increased repair capacity for O6mG, subsequently, decreased mutational activation of K-ras oncogene. Heterogeneity in the level of AGT expressed in different lung cell populations or other forms of carcinogenic DNA damage caused by NNK may explain the residual incidence of lung tumors in MGMT transgenic mice.
Carcinogenesis 1999 Feb
PMID:Reduced lung tumorigenesis in human methylguanine DNA--methyltransferase transgenic mice achieved by expression of transgene within the target cell. 1006 65

The enzyme O6-methylguanine-DNA methyltransferase (MGMT) protects cells from the cytotoxic and mutagenic effects of alkylating agents. Approximately 20% of tumor cell lines lack MGMT activity and are highly sensitive to alkylating agents. In established cancer cell lines, MGMT expression appears to be correlated with methylation of residues in both the promoter and the body of the gene. The effect of methylation of the MGMT promoter on gene expression and carcinogenesis in primary tumors is unknown. We investigated methylation of the MGMT promoter region in primary colorectal cancers and normal colonic mucosa. We used five methylation-sensitive restriction enzymes (BssHII, SacII, Eagl, Nael, and Smal) and Southern blot analysis to assess methylation in 46 cancers and 22 controls. Methylation of Eagl and Nael sites was seen in 12 tumors but in none of the 22 normal colorectal mucosa specimens. This difference was statistically significant (P<0.01). Methylation-sensitive single-nucleotide primer extension analysis of four additional cytosine residues confirmed methylation of the promoter region in the tumors identified by Eagl and Nael digestions and served to further quantitate the extent of methylation. Western blot analysis of 21 tumors revealed statistically significant lower MGMT expression in the eight tumors with methylation of the Eagl and Nael sites and nt -128 than in the 13 tumors lacking the methylation pattern (P<0.05). MGMT activity was lower in tumors with methylation than in tumors that were not methylated. The difference was not, however, statistically significant. We conclude that a subset of colorectal tumors is characterized by a specific methylation pattern in the MGMT promoter associated with reduced MGMT expression.
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PMID:A specific CpG methylation pattern of the MGMT promoter region associated with reduced MGMT expression in primary colorectal cancers. 1007 36

Mice deficient in the DNA mismatch repair (MMR) gene, PMS2, develop spontaneous thymic lymphomas and sarcomas. We have previously shown that PMS2(-/-) mice were hypersensitive to a single i.p. injection of 50 mg/kg of N-methyl-N-nitrosourea (MNU) for thymic lymphoma induction. We postulated that MNU sensitivity was due to formation of O(6)-methylguanine (O(6)-mG), which, if unrepaired by O(6)-alkylguanine DNA alkyltransferase (AGT), leads to apoptosis in MMR competent cells and O(6)-mG:T mismatches in MMR deficient cells. Tumor induction is less in MMR(+/+) mice because cells with residual DNA adducts die, whereas mutagenized cells survive in MMR(-/-) mice. Overexpression of AGT (encoded by the methylguanine DNA methyltransferase-MGMT-gene) is known to block MNU induced tumorigenesis in mice with functional MMR. To further determine the sensitivity of PMS2(-/-) mice to MNU and the protective effect of hAGT overexpression, a low dose of MNU (25 mg/kg) was studied in PMS2(-/-) mice and PMS2(-/-)/hMGMT(+) mice. No thymic lymphomas were found in MNU-treated PMS2(+/+) and PMS2(+/-) mice. At 1 year, 46% of the MNU-treated PMS2(-/-) mice developed thymic lymphoma, compared with an incidence of 25% in both untreated PMS2(-/-) mice and MNU treated PMS2(-/-)/hMGMT(+) mice. In addition, a significantly shorter latency in the onset of thymic lymphomas was seen in MNU-treated PMS2(-/-) mice. K-ras mutations were detected almost equally in the thymic lymphomas induced by MNU in both PMS2(-/-) and PMS2(-/-)/hMGMT(+) mice, but not in the spontaneous lymphomas. These data suggest that PMS(-/-) mice are hypersensitive to MNU, that there are different pathways responsible for spontaneous and MNU induced thymic lymphomas in PMS2(-/-) mice, and that overexpression of hMGMT protects the mice by blocking non-K-ras pathways.
Carcinogenesis 1999 Sep
PMID:Transgenic expression of human MGMT blocks the hypersensitivity of PMS2-deficient mice to low dose MNU thymic lymphomagenesis. 1046 9

Carboxymethylating agents are potential sources of endogenous DNA damage that have been proposed as possible contributors to gastrointestinal carcinogenesis. The cytotoxicity of the model DNA carboxymethylating agent azaserine was investigated in human cells. Expression of the DNA repair enzyme O(6)-methylguanine-DNA methyltransferase (MGMT) did not affect sensitivity to the drug in two related Raji Burkitt's lymphoma cell lines. DNA mismatch repair-defective variants of Raji cells which display increased tolerance to DNA methylation damage were not selectively resistant to azaserine. Complementary results were obtained with a second carboxymethylating agent, potassium diazoacetate. In contrast, lymphoblastoid cell lines representative of each of the xeroderma pigmentosum complementation groups, including the variant, were all significantly more sensitive to azaserine than nucleotide excision repair-proficient cells. The hypersensitivity of XP cells was not due to systematic differences in the concentrations of intracellular thiol compounds or related thiol metabolizing enzymes. The data indicate that of the two types of potentially lethal DNA damage which azaserine introduces, carboxymethylated bases and O(6)-methylguanine, the former are repaired by nucleotide excision repair and are a more significant contributor to azaserine lethality in human cells.
Carcinogenesis 1999 Sep
PMID:The cytotoxicity of DNA carboxymethylation and methylation by the model carboxymethylating agent azaserine in human cells. 1046 34

Differential expression of DNA-O6MeG: protein-L-cysteine S-methyltransferase (MGMT) activity and posttranslational modification of the protein during liver regeneration and carcinogenesis were compared in Sprague-Dawley male rats after partial hepatectomy and/or single i.p injection of diethylnitrosamine (DEN, 200 mg/kg). Regenerating hepatocytes after partial hepatectomy induced MGMT transiently within 3 days; however, the induction of MGMT was persistent for 2 weeks after DEN injection, and the combined treatment of DEN and partial hepatectomy maintained the elevated MGMT level for up to 4 weeks. The increased activity was transcriptionally regulated, when analyzed by Northern blot hybridization. The major active form of MGMT protein in the partially hepatectomized or DEN-treated rats was a 26-kDa or 24-kDa species respectively, which was confirmed by Western blot analysis and gel slice assay. The biological significance of the differential induction of MGMT during partial hepatectomy or DEN-induced carcinogenesis is not obvious; however, further studies on possible posttranslational modifications of MGMT protein might shed some light on the functional aspect of MGMT induction.
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PMID:Differential expression of O6-methylguanine-DNA methyltransferase during diethylnitrosamine-induced carcinogenesis and liver regeneration in Sprague-Dawley male rats. 1048 Mar 42

O(6)-alkylguanine-DNA alkyltransferase (AGT) is a suicide protein that corrects DNA damage by alkylating agents and may also serve to activate environmental carcinogens. We expressed human wild-type and two active mutant AGTs in bacteria that lack endogenous AGT and are also defective in nucleotide excision repair, to examine the ability of the AGTs to protect Escherichia coli from DNA damage by different types of alkylating agents and, oppositely, to sensitize cells to the genotoxic effects of dibromoalkanes (DBAs). Control bacteria carrying the cloning vector alone were extremely sensitive to mutagenesis by low, noncytotoxic doses of N-methyl-N'-nitro-N-nitrosoguanidine (MNNG). Expression of human wild-type AGT prevented most of this enlarged susceptibility to MNNG mutagenesis. Oppositely, cell killing required much higher MNNG concentrations and prevention by wild-type AGT was much less effective. Mutants V139F and V139F/P140R/L142M protected bacteria against MNNG-induced cytotoxicity more effectively than the wild-type AGT, but protection against the less stringent mutagenesis assay was variable. Subtle differences between wild-type AGT and the two mutant variants were further revealed by assaying protection against mutagenesis by more complex alkylating agents, such as N-ethyl-N-nitrosourea and 1-(2-chloro- ethyl)-3-cyclohexyl-1-nitrosourea. Unlike wild-type and V139F, the triple mutant variant, V139F/P140R/L142M was unaffected by the AGT inhibitor, O(6)-benzylguanine. Wild-type AGT and V139F potentiated the genotoxic effects of DBAs; however, the triple mutant virtually failed to sensitize the bacteria to these agents. These experiments provide evidence that in addition to the active site cysteine at position 145, the proline at position 140 might be important in defining the capacity by which AGTs modulate genotoxicity by environmentally relevant DBAs. The ability of AGTs to activate dibromoalkanes suggests that this DNA repair enzyme could be altered, and if expressed in tumors might be lethal by enhancing the activation of specific chemotherapeutic prodrugs.
Carcinogenesis 1999 Nov
PMID:Human O(6)-alkylguanine-DNA alkyltransferase: protection against alkylating agents and sensitization to dibromoalkanes. 1054 10

Azoxymethane (AOM) causes O(6)-methylguanine adduct formation which leads to G-->A transitions. Their repair is carried out by O(6)-methylguanine-DNA methyltransferase (MGMT). To evaluate the importance of this repair event in AOM-induced carcinogenesis, we examined the effect of O(6)-benzylguanine (BG), a potent inhibitor of MGMT, on colonic tumor development. Rats were treated weekly for 2 weeks at 0 and 24 h with BG (60 mg/kg body wt i.p.) or vehicle (40% polyethylene glycol, PEG-400), followed 2 h after the first dose of BG with AOM (15 mg/kg body wt) or vehicle (saline) i.p. Rats were killed 35 weeks later and tumors harvested and DNA extracted. In the AOM-treated groups, BG caused a significant increase in tumor incidence with tumors in 65.9%, versus 30.8% in the AOM/PEG-treated group (P < 0.05). In the BG/AOM group there was also a significant increase in tumor multiplicity, with 2.3 tumors/tumor-bearing rat, versus 1.6 tumors/tumor- bearing rat in the AOM/PEG group (P < 0.05). Since O(6)-methylguanine adducts can cause activating mutations in the K-ras and beta-catenin genes, we examined the effects of BG on these mutations. In the BG group there were seven mutations in codon 12 or 13 of exon 1 of the K-ras gene in 51 tumors examined, compared with no K-ras mutations in 17 tumors analyzed in the AOM/PEG group (P = 0.12). In the BG/AOM group there were 10 mutations in exon 3 of the beta-catenin gene among 48 tumors evaluated, compared with six mutations in 16 tumors analyzed in the PEG/AOM group (P = 0.16). In summary, MGMT inhibition increases AOM-induced colonic tumor incidence and multiplicity in rats.
Carcinogenesis 1999 Dec
PMID:Inhibition of O(6)-methylguanine-DNA methyltransferase increases azoxymethane-induced colonic tumors in rats. 1059 Feb 33

We observed previously that wild-type p53 rendered neonatal mouse astrocytes resistant to 1,3-bis(2-chloroethyl)-1-nitrosourea (BCNU) in a gene dose-dependent fashion. This effect of p53 appeared to be unrelated to its cell cycle regulation or apoptotic functions. Because in many cell types O(6)-methylguanine-DNA methyltransferase (MGMT)-mediated DNA repair is an important mechanism of resistance to nitrosoureas, we measured MGMT activity in wild-type, heterozygous and p53 knockout neonatal mouse astrocytes. Wild-type p53 astrocytes had significantly greater MGMT activity than either heterozygous or p53 knockout astrocytes: MGMT activity was approximately 5-fold greater in wild-type p53 astrocytes than in p53 knockout cells. However, despite successful depletion of MGMT activity in wild-type astrocytes by O(6)-benzylguanine (BG), resistance to BCNU persisted unchanged. Moreover, we excluded the possibility that continued resistance to BCNU at the concentrations used could be explained by a compensatory induction of MGMT triggered by exposure to either BCNU or BG. Although these studies support a role for p53 regulation of MGMT in neonatal mouse astrocytes, BCNU resistance in wild-type cells appears to be mediated by a non-MGMT mechanism. Nevertheless, regulation of DNA repair by MGMT may be another mechanism by which alterations of the p53 gene promote tumor initiation or progression.
Carcinogenesis 1999 Dec
PMID:O(6)-methylguanine-DNA methyltransferase activity, p53 gene status and BCNU resistance in mouse astrocytes. 1059 Feb 34


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