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:6.5.1.2 (DNA ligase)
2,749 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Resistance to alkylating agents is partly due to the presence of the DNA repair enzyme, termed O6 alkyltransferase (O6AT). Preclinical evidence of the transient restoration of sensitivity of cells resistant to nitrosoureas by pretreatment with a methylating agent, whose role is to deplete cells of O6AT activity and clinical evidence of such a depletion in patients lymphocytes, led us to test the sequential administration of dacarbazine 3 h prior to fotemustine, a chloroethylnitrosourea derivative. 24 patients with measurable advanced melanoma entered the trial and are evaluable. Toxicity was mainly haematological with early neutropenia and/or thrombocytopenia. Clinical activity (33%) was impressive especially on lung metastases with high complete response rate for that site (7/14). Unfortunately, the occurrence of a rapidly fatal pulmonary toxicity precludes further use of the regimen before a plausible explanation for this unexpected toxicity is obtained. Indeed, similar cases have been reported in other trials using the sequential schedule while no lung toxicity was reported in single agent or alternated administrations. Preclinical studies are ongoing to test the hypothesis of a glutathione depletion and the possibility of a rescue treatment.
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PMID:Sequential administration of dacarbazine and fotemustine in patients with disseminated malignant melanoma--an effective combination with unexpected toxicity. 159 Oct 62

Different biological aspects of a novel 2-chloroethyl nitrosourea derived from cysteamine, N'-(2-chloroethyl)-N-[2-(methylsulfinyl)ethyl]-N'- nitrosourea (CMSOEN2), were studied. Drug-induced cytotoxic effects, uptake kinetics, DNA damage, and O6-alkylguanine-DNA alkyltransferase activity were determined in 3 melanoma cell lines: the murine B16 and 2 human metastatic-derived cell lines (M4 Beu and M3 Dau). We found that radioactivity uptake and incorporation in acido-precipitable material was inversely proportional to cell drug viability. The highly CMSOEN2-sensitive B16 line showed the lowest total radioactivity uptake. In fact, among the melanoma cell parameters studied, 3 of them were well correlated: (a) cytotoxicity as reflected by the colony-forming assay; (b) DNA cross-link frequency estimated by the alkaline elution technique; and (c) O6-alkylguanine-DNA alkyltransferase activity (Mer phenotype), defined as the ability of cell extracts to remove O6-methylguanine from N-methyl-N-nitrosourea-alkylated DNA. The 2 human cell lines (M4 Beu and M3 Dau), the most resistant to the cytostatic drug effects, showed little or no ability to form DNA lethal cross-links. These results correspond to the higher O6-alkylguanine-DNA alkyltransferase activity found in human-derived cell lines compared with that present in murine B16 cell lines. This study confirms that the cell content in this repair DNA protein is certainly one of the important factors implicated in the variability of response to 2-chloroethyl nitrosourea treatment observed in a number of established malignant cell lines. It has been shown that pretreatment of derived cell lines with methylating agents (N-methyl-N-nitrosourea, N-methyl-N'-nitro-N-nitrosoguanidine) or O6-methylguanine used as a free base, increased cytotoxic effects of this class of anticancer agents, likely by saturating receptor sites (sulfhydryl groups) of this specific DNA repair enzyme. Nevertheless, in preliminary Phase I and II clinical trials, 2 patients who had been treated with multiple chemotherapies including alkylating agents [1-(2-chloroethyl)-3- cyclohexyl-1-nitrosourea, 5-(3,3-dimethyl-1-triazeno)-imidazole-4-carboxamide, platinum derivatives], presented complete or partial remission after CMSOEN2 treatment. Our results raise the question of the exact relation between the Mer phenotype determined in derived murine or human cultured cells and that directly observed on surgically excised tumors in cancer patients. The original Mer phenotype could be modified by cell culture conditions since it has been shown that O6-alkylguanine-DNA alkyltransferase activity is widely distributed between normal and tumoral tissues without any real difference.
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PMID:DNA damage induced by a new 2-chloroethyl nitrosourea on malignant melanoma cells. 239 61

Tanning is a protective response of ultraviolet (UV)-irradiated skin that decreases damage from subsequent sun exposures by increasing the epidermal content of melanin, a brown-black pigment that absorbs light energy throughout the UV and visible portions of the electromagnetic spectrum. The melanin pigment is made by epidermal melanocytes and transferred to surrounding keratinocytes. The action spectrum, time course, and histologic features of tanning are well studied, but the initiating molecular events are unknown. Previous work has shown that T4 endonuclease V, a prokaryotic DNA repair enzyme that catalyzes the first and rate-limiting step in repair of UV-induced pyrimidine dimers, delivered in carrier liposomes (T4N5), enhances repair of UV-induced DNA damage in cultured human cells and protects against photocarcinogenesis in an animal model. We now report that T4N5 treatment enhances UV-induced melanogenesis, as measured by melanin content, tyrosinase activity, 14C-dopa incorporation, and visual assessment in both S91 murine melanoma cells and human melanocytes. T4N5 treatment also increases cell yields following UV irradiation. These data suggest that tanning can be stimulated through enhanced DNA repair.
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PMID:Treatment of human melanocytes and S91 melanoma cells with the DNA repair enzyme T4 endonuclease V enhances melanogenesis after ultraviolet irradiation. 822 26

Mammalian DNA polymerase beta is a DNA repair enzyme expressed constitutively at a low level. In vitro, purified DNA polymerase (Pol) beta incorporates the nucleotide analogues 2'-3' deoxycytidine (ddC)-triphosphate and 3'-azido-3'-deoxythymidine (AZT)-triphosphate into DNA, causing chain termination. We have tested the possibility of enhancing the cytotoxicity of these chain terminators against mammalian cells by increasing the level of Pol beta. Chinese hamster ovary AA8 and murine melanoma B16 cell lines were stably transfected with rat pol beta cDNA under the control of a viral enhancer/promoter. We found that overexpression of Pol beta sensitized the cells to ddC and AZT. To confirm the role of this polymerase in this process, we prepared cell extracts from the control and Pol beta overexpressing Chinese hamster ovary cell lines and tested in vitro their capacity to incorporate ddC-triphosphate and AZT-triphosphate into DNA. We found that inhibition of DNA replication by both chain terminators was more pronounced when extracts from pol beta-transfected cells were used, providing a direct evidence of the involvement of Pol beta in the sensitization process. In addition, we showed that cotransfection with bacterial or viral thymidine/thymidylate kinase genes enhanced the Pol beta-mediated cytotoxicity of AZT, suggesting that phosphorylation and polymerization activities might be combined to potentiate their respective effects. These observations may be useful for improving therapeutic efficiency of DNA chain terminators.
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PMID:Overexpression of DNA polymerase beta sensitizes mammalian cells to 2',3'-deoxycytidine and 3'-azido-3'-deoxythymidine. 898 50

We investigated the effect of oestrogens, anti-oestrogens and flavonoids on the growth of a human melanoma cell line (SK-Mel-28) and, at the same time, the presence of both type I oestrogen receptors (ERs) and type II oestrogen binding sites (type II EBS) to gain a fuller picture of the relationship between melanoma cell proliferation and receptor status. 17beta-Oestradiol (E2) and the flavonoid quercetin (Q) produced a marked inhibition of proliferation, but only at the highest dose used (10(-5) M) and only when added daily to the medium. Diethylstilboestrol (DES) (10(-5) M) was effective in inhibiting cell growth when the medium was renewed every 3 days and produced a more pronounced reduction when added daily to the medium. Tamoxifen (TAM) inhibited cell proliferation at a dose starting from 10(-7) M when the medium was renewed every 3 days. When added daily to the medium, it did not induce a greater inhibitory effect and it was cytotoxic at 5 x 10(-6) M and 10(-5) M. The antiproliferative effect of E2, DES and Q did not seem to be dependent on their interaction with ERs, which were minimally detected in SK-Mel-28 in both immunocytochemical and biochemical assays. Our model revealed, through a biochemical assay, a large number of type II EBSs which could be involved in the anti-oestrogen action, but this does not exclude the involvement of other mechanisms. Finally, TAM (10(-5) M) appeared to reduce the activity of the DNA repair enzyme O6-alkylguanine-DNA alkyltransferase, an effect that could be interesting from the point of view of the therapeutic efficacy of alkylating agents.
Melanoma Res 1998 Aug
PMID:Sensitivity of human melanoma cells to oestrogens, tamoxifen and quercetin: is there any relationship with type I and II oestrogen binding site expression? 976 6

Mutations in the mismatch DNA repair gene human MutS homologen 2 (hMSH2) are causative for microsatellite instability and carcinogenesis in various human tumours, including hereditary nonpolyposis colorectal cancer. Because microsatellite instability has been detected in malignant melanoma, we have investigated hMSH2 in melanocytic tumours. We found strong nuclear immunoreactivity for hMSH2 that was elevated in malignant melanoma and melanoma metastases as compared to acquired nevi. These findings suggest that increased genomic instability in malignant melanoma is associated with elevated protein levels of this DNA repair enzyme. hMSH2 is not exclusively regulated by proliferative activity in melanocytes, because there was no correlation between staining patterns of hMSH2 and the proliferation marker Ki-67. In contrast, immunoreactivity scores for hMSH2 and p53 were both upregulated in malignant melanocytic tumours. These findings support the concept that hMSH2 gene expression may be regulated in melanocytes by the p53 protein, as has been reported previously in other tissues. Using the reverse transcription-polymerase chain reaction, we detected strong hMSH2 mRNA expression in each of 8 melanoma cell lines analysed (highest amounts in SK-MEL-25 cells, lowest amounts in MML-I cells). In conclusion, our findings indicate that hMSH-2 may be of importance for genetic stability, tumorigenesis and progression of malignant melanoma.
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PMID:DNA mismatch repair enzyme hMSH2 in malignant melanoma: increased immunoreactivity as compared to acquired melanocytic nevi and strong mRNA expression in melanoma cell lines. 1193 86

Deprivation of tyrosine (Tyr) and phenylalanine (Phe) inhibits growth and induces programmed cell death (apoptosis) of human A375 melanoma cells. Herein, we found that activation of caspases and release of mitochondrial cytochrome c are required for this process. Culturing A375 cells in Tyr/Phe-free medium, containing 10% dialyzed fetal bovine serum, results in activation of caspase-3-like activity. This is accompanied by decreased cell viability and increased apoptosis. Tyr/Phe deprivation also stimulates proteolytic cleavage of the DNA repair enzyme, poly(ADP-ribose) polymerase (PARP). Western blot analysis showed that caspases 3, 7, 8, and 9 are activated by deprivation of Tyr/Phe. Tyr/Phe deprivation decreases mitochondrial membrane potential, induces cleavage of Bid, increases translocation of Bax from the cytosol to mitochondria, and results in release of cytochrome c from the mitochondria to the cytosol. Apoptosis due to Tyr/Phe deprivation is almost completely inhibited by the broad-spectrum cell-permeable caspase inhibitor, benzyloxycarbonyl-Val-Ala-Asp-fluoromethyl ketone (Z.VAD.fmk). This inhibitor suppresses the cleavage of Bid, the release of cytochrome c from the mitochondria to the cytosol, and the cleavage of PARP. Decylubiquinone, a mitochondrial permeability transition pore inhibitor, does not suppress the activation of caspase 8 but suppresses release of cytochrome c, activation of caspase 9, and induction of apoptosis. These results indicate that activation of caspases, cleavage of Bid, and mitochondrial release of cytochrome c are required for apoptosis induced by Tyr/Phe deprivation.
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PMID:Activation of caspases and cleavage of Bid are required for tyrosine and phenylalanine deficiency-induced apoptosis of human A375 melanoma cells. 1206 1

The prognosis of advanced melanoma is generally poor, because this tumor commonly exhibits intrinsic or acquired resistance to chemotherapy. In an attempt to identify the underlying causes of this resistance, we studied the roles played by the DNA repair enzyme O(6)-alkylguanine-DNA alkyltransferase (OGAT) and the mismatch repair (MMR) system in the sensitivity of melanoma cells to temozolomide (TMZ), 1,3-bis(2-chloroethyl)-1-nitrosourea (BCNU), or cis-diamminedichloroplatinum(II) (CDDP). To this end, OGAT levels and MMR efficiency of extracts of nine melanoma cell lines and selected clones derived from four of these lines were determined and correlated with the sensitivity of the respective cells to these drugs. The effectiveness of O(6)-benzylguanine (BG), a specific OGAT inhibitor, in potentiating TMZ- or BCNU-mediated cytotoxicity was also evaluated. Our results demonstrate that MMR efficiency and OGAT levels strongly affect melanoma cell sensitivity to TMZ. In MMR-proficient cells, a direct correlation between OGAT levels and TMZ IC(50) values was found. When OGAT activity was inhibited with BG, the sensitivity of these cells to TMZ increased and was then dictated largely by their MMR efficiency. MMR-deficient cells were highly resistant to the drug irrespective of their OGAT levels. Although OGAT activity and MMR status seemed to be the major determinants of melanoma sensitivity to TMZ, this was not the case for BCNU and CDDP; resistance to the latter drugs clearly involves processes other than the two DNA repair pathways analyzed in this study.
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PMID:The effect of O6-alkylguanine-DNA alkyltransferase and mismatch repair activities on the sensitivity of human melanoma cells to temozolomide, 1,3-bis(2-chloroethyl)1-nitrosourea, and cisplatin. 1253 19

The project focuses on cancer types with outstanding publich health importance (breast-, colorectalhead and neck cancers and childhood tumors). Epidemiological studies revealed significant regional differences in the mobidity/mortality of these cancer types in Hungary. Molecular epidemiological studies revealed characteristic BRCA1 mutation patterns of familiar breast cancer and DNA repair enzyme polymorphism in head and neck cancer. New methods have been developed for the screening (lactoferrin), prognostication (c-met expression) or the prediction of therapeutic sensitivity (TS expression) of colorectal cancer. In the pediatric oncology program alternative way of MRD monitoring (WT1 expression) and a potential new therapeutic modality (IFN-alpha) of ALL was developed. Experimental studies demonstrated that the tumoral matrix significantly influences the effects of the classic chemotherapeutic agents. We have identified several genes the expression of which could serve efficiently as markers or targets for therapy of the progression of melanoma (alphaIIbbeta3 integrin, CD44v3 and decorin proteoglycans, AMF receptor).
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PMID:[Report on the first year of the activity of the National Oncological RD Consortium]. 1256 51

Temozolomide is an alkylating cytostatic drug that finds increasing application in the treatment of melanoma, anaplastic astrocytoma and glioblastoma multiforme. The compound is a prodrug that decomposes spontaneously, independent of an enzymatic activation step. DNA methylation induces futile mismatch repair cycles and depletion of the DNA repair enzyme O(6)-methylguanine-DNA methyltransferase should then initiate programmed cell death. We show drug-dependent inhibition of tumour growth in a three-dimensional cell culture model of the glioma cell lines U87MG and GaMG. Migrational behaviour of the glioblastoma cells remained unaltered. However, coincubation of tumour spheroids with primary brain aggregates showed reduced tumour cell invasion into brain tissue in the presence of temozolomide. This was not achieved by slowing cellular migration, as temozolomide-treated cells displayed no reduced motility. By transferase-mediated dUTP nick-end labelling (TUNEL) of apoptotic nuclei, we found that the drug was able to induce apoptosis throughout the tumour cell spheroids. Apoptosis was highest in the core region of the spheroids. Repetitive application of sublethal doses of temozolomide to multicellular spheroids resulted in the development of drug resistance in GaMG cells. We suggest that temozolomide is a strong initiator of apoptosis in glioblastoma tumour cells in a spheroid cell culture system, when cells are already in a stressful environment.
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PMID:Temozolomide induces apoptosis and senescence in glioma cells cultured as multicellular spheroids. 1256 92


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