vitamin-k-semiquinone-radical and Colonic-Neoplasms

It is proposed that bile acids (deoxycholic acid), the K vitamins, iron(II) complexes and oxygen interact to induce an oncogenic effect in the colon by the generation of free radicals. In the relatively low oxidising/reducing conditions of the colonic lumen the K vitamins exist in the reduced form; however, if absorbed into the mucosa they have the capacity to be chemically oxidised and to enter into a redox cycle yielding oxygen radicals. The semiquinone radical of K(1) (phylloquinone) has been stabilised in bile acid mixed micelles and investigated by electron paramagnetic resonance spectroscopy and quantum chemical calculations. The estimated half-life of the radical was about 30 min which confirms a remarkably high stability in aqueous micellar solution. A model is presented in which the reduced K vitamins may initiate superoxide radical, O2(-*) generation leading to Fe(II) mediated Fenton reactions in the stem colon cells.

Topics: Bile Acids and Salts; Colon; Colonic Neoplasms; Diet; Electron Spin Resonance Spectroscopy; Half-Life; Humans; Intestinal Mucosa; Micelles; Models, Chemical; Molecular Structure; Oxidation-Reduction; Quinones; Superoxides; Vitamin K; Vitamin K 1

The human colon carcinoma cell lines Caco-2 and HT-29 were exposed to three structurally related naphthoquinones. Menadione (MEN), 1,4-naphthoquinone (NQ), and 2,3-dimethoxy-1,4-naphthoquinone (DIM) redoxcycle at similar rates, NQ is a stronger arylator than MEN, and DIM does not arylate thiols. The Caco-2 cell line was particularly vulnerable to NQ and MEN and displayed moderate toxic effects of DIM. The HT-29 cell line was only vulnerable to NQ and MEN after inhibition of DT-diaphorase (DTD) with dicoumarol, whereas dicoumarol did not affect the toxicity of quinones to Caco-2 cells. DTD activity in the HT-29 and Caco-2 cell lines, as estimated by the dicoumarol-sensitive reduction of 2,6-dichlorophenolindophenol, was 393.7 +/- 46.9 and 6.4 +/- 2.2 nmol NADPH x min(-1) x mg protein(-1), respectively. MEN depleted glutathione to a small extent in the HT-29 cell line, but a rapid depletion similar to Caco-2 cells was achieved when dicoumarol was added. The data demonstrated that the DTD-deficient Caco-2 cell line was more vulnerable to arylating or redoxcycling quinones than DTD-expressing cell lines. Exposure of the Caco-2 cell line to quinones produced a rapid rise in protein disulphides and oxidised glutathione. In contrast to NQ and DIM, no intracellular GSSG was observed with MEN. The relatively higher levels of ATP in MEN-exposed cells may account for the efficient extrusion of intracellular GSSG. The reductive potential of the cell as measured by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide reduction was only increased by MEN and not with NQ and DIM. We conclude that arylation is a major contributing factor in the toxicity of quinones. For this reason, NQ was the most toxic quinone, followed by MEN, and the pure redoxcycler DIM elicited modest toxicity in Caco-2 cells.

Topics: Adenosine Triphosphate; Cell Survival; Colonic Neoplasms; Dicumarol; Glutathione; Glutathione Disulfide; Humans; Kinetics; NAD(P)H Dehydrogenase (Quinone); Naphthoquinones; Neoplasm Proteins; Sulfhydryl Compounds; Tumor Cells, Cultured; Vitamin K

To investigate the resistant mechanisms against MMC in human tumor cells, we isolated an MMC-resistant variant (HT-29/MMC) of HT-29 human colon carcinoma cells. HT-29/MMC cells showed 5-fold resistance to MMC as compared with the parental cell line but did not show cross-resistance to Adriamycin, vincristine, ACNU, bleomycin, or cisplatin. Treatment of the cells with dicoumarol, an inhibitor of DT-diaphorase, reduced the cytotoxicity of MMC in DT-diaphorase proficient HT-29 cells but not in HT-29/MMC cells. HT-29/MMC cells were 5 times more sensitive than HT-29 cells to menadione, which is detoxified by DT-diaphorase, DT-diaphorase was deficient in HT-29/MMC cells as determined by the enzyme activity and immunoblot analysis of the cytoplasmic proteins. Levels of cytochrome P-450 reductase and glutathione S-transferase, however, were comparable in both cell lines. The amount of [3H]-MMC found covalently bound to chromosomal DNA in HT-29/MMC cells was one-fourth that detected in HT-29 cells. Treatment with dicoumarol reduced the DNA-bound MMC in HT-29 cells but not in HT-29/MMC cells. These results indicate that the deficiency in DT-diaphorase, an activating enzyme of MMC, is one of the mechanisms of resistance in HT-29/MMC cells.

Topics: Aerobiosis; Alkylation; Biotransformation; Cell Survival; Colonic Neoplasms; Dicumarol; DNA, Neoplasm; Drug Resistance; Glutathione Transferase; Humans; Mitomycin; NAD(P)H Dehydrogenase (Quinone); NADPH-Ferrihemoprotein Reductase; Oxidation-Reduction; Tumor Cells, Cultured; Vitamin K

Reactive oxygen metabolites have been associated with gastrointestinal injury and may play a role as mediators of inflammation. The effect of oxygen metabolites on Caco-2 cell viability (trypan blue exclusion and 51Cr release), hexose monophosphate shunt activity, and glutathione was assessed. Caco-2 cells were incubated with amino acid oxidase, xanthine oxidase, menadione, and t-butylhydroperoxide in the presence and absence of superoxide dismutase, catalase, mannitol, and butylated hydroxytoluene. With amino acid oxidase, trypan blue exclusion decreased (P < 0.01) and 51Cr release, oxidized glutathione, and shunt activity increased (P < 0.05). The addition of catalase attenuated these changes. Trypan blue exclusion decreased (P < 0.005) and 51Cr release, oxidized glutathione, and shunt activity increased (P < 0.01) with xanthine oxidase. The addition of superoxide dismutase caused a further increase in 51Cr release, oxidized glutathione, and shunt activity (P < 0.01), which was prevented by the addition of catalase or mannitol. t-Butylhydroperoxide did not effect 51Cr release or trypan blue exclusion, but oxidized glutathione and shunt activity increased (P < 0.01). The increase in shunt activity was prevented by preincubation with butylated hydroxytoluene (P < 0.01). Menadione did not alter trypan blue exclusion or 51Cr release, but caused an increase in oxidized glutathione and shunt activity (P < 0.001). The increase in shunt activity was attenuated by preincubation with butylated hydroxytoluene (P < 0.001). Menadione also caused a depletion of total glutathione.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: Chromium Radioisotopes; Colonic Neoplasms; Digestive System; Glutathione; Humans; Oxidants; Pentose Phosphate Pathway; Peroxides; Reactive Oxygen Species; tert-Butylhydroperoxide; Trypan Blue; Tumor Cells, Cultured; Vitamin K

The effects of menadione (Vit K3) administered at 20 or 40 mg.kg-1 ig 3 times a week for both 24 and 28 wk on 1,2-dimethylhydrazine (DMH)-induced mouse colon adenocarcinomas were investigated. At the 24th wk, the number of colon tumors in Vit K3 20 or 40 mg.kg-1 group (0.3 +/- 0.5 and 0.5 +/- 0.8, respectively) was less than that of DMH controls (2.1 +/- 2.5, P < 0.05), but the difference in incidence of colon tumors in these 3 groups was not significant (P > 0.05). After 28 wk, the tumor incidence of both Vit K3 groups (each 8 of 13) was lower than that of DMH controls (13 of 13, P < 0.05); the number of colon tumors of Vit K3 40 mg.kg-1 group (1.3 +/- 1.3, P < 0.05) was decreased, whereas the Vit K3 20 mg.kg-1 group (3.0 +/- 5.1, P > 0.05) was not different from the DMH controls (7.3 +/- 9.3). Determination of the nuclear DNA content of cells from DMH-induced mouse colon mucosa (24 wk) indicated that Vit K3 20 or 40 mg.kg-1 group showed lower DNA content (1.92 +/- 0.12 C and 1.91 +/- 0.10 C, respectively) decreased values of percent-over-3C and -4C and narrow distribution range. Besides, the colon mucosa of DMH-treated mice (28 wk) showed higher superoxide dismutase (SOD) activity (70 +/- 28 U/mg protein, P < 0.05) than the normal controls (30 +/- 20 U/mg protein). Vit K3 40 mg.kg-1 reduced the elevated SOD activity markedly (44 +/- 23 U/mg protein, P < 0.05).

Topics: Adenocarcinoma; Animals; Antineoplastic Agents; Colonic Neoplasms; Dimethylhydrazines; DNA, Neoplasm; Female; Intestinal Mucosa; Mice; Mice, Inbred ICR; Superoxide Dismutase; Vitamin K; Vitamin K 3

A mitomycin C (MMC)- and porfiromycin (PFM)-resistant subline of the HCT 116 human colon-cancer cell line was isolated after repeated exposure of HCT 116 cells to increasing concentrations of MMC under aerobic conditions. The MMC-resistant subline (designated HCT 116-R30A) was 5 times more resistant than the parent cells to MMC and PFM under aerobic conditions. Both the MMC-resistant cells and the parent HCT 116 cells accumulated similar amounts of PFM by passive diffusion, but levels of macromolecule-bound PFM were about 50% lower in the resistant cell line, implying a decrease in PFM reductive activation in the resistant cells. The finding that microsomes from either sensitive or resistant cells showed an equal ability to reduce MMC and PFM indicated that the activity of NADPH cytochrome P-450 reductase (EC 1.6.2.4) was not changed in the resistant subline. Soluble extracts of HCT 116 cells reduced MMC and PFM more effectively at pH 6.1, and NADH and NADPH were utilized equally well as electron donors under both aerobic and anaerobic conditions. These data suggest that quinone reductase (EC 1.6.99.2; DT-diaphorase) in soluble extracts is responsible for the reduction of MMC. Quinone reductase activities in soluble extracts of HCT 116-R30A cells for the reduction of dichlorophenol indophenol (DCPIP) and menadione-cytochrome c at optimal pHs were decreased by 95% as compared with those obtained in parent cells. However, the MMC-reducing activity of HCT 116-R30A soluble extracts was only 50% lower than that of the parent cell extracts. The kinetic constants (Km, Vmax) found for quinone reductase in the two cell lines with respect to the substrates DCPIP and menadione differed. Two species of mRNA for quinone reductase (2.7 and 1.2 kb) were detected in both cell lines, and there was no detectable difference between parent and resistant cells in the steady-state level of either of these mRNA species. Furthermore, incubation with the quinone reductase inhibitor dicoumarol rendered HCT 116 cells more resistant to MMC. Alteration of the quinone reductase activity in HCT 116-R30A cells appears to be the mechanism responsible for their resistance to MMC and PFM.

Topics: 2,6-Dichloroindophenol; Carbon Radioisotopes; Cell Extracts; Cell Size; Cell Survival; Colonic Neoplasms; Drug Resistance; Drug Screening Assays, Antitumor; Humans; Intracellular Fluid; Microsomes; Mitomycin; NAD(P)H Dehydrogenase (Quinone); Oxidation-Reduction; Porfiromycin; RNA, Messenger; Tissue Distribution; Transcription, Genetic; Tumor Cells, Cultured; Vitamin K

EO9 [3-hydroxymethyl-5-aziridinyl-1-methyl-2-(H-indole-4, 7-indione)-propenol] is a novel indoloquinone structurally related to mitomycin C, a quinone anticancer drug that requires reductive bioactivation. NAD(P)H: (quinone-acceptor) oxidoreductase (quinone reductase, DT-diaphorase, EC 1.6.99.2) is an obligate 2-electron donating enzyme that can reduce a variety of quinones resulting either in bioactivation or bioprotection. Using quinone reductase (QR) preparations from rat Walker 256 mammary tumor cells and human HT29 colon carcinoma cells, we have characterized the role of this enzyme in EO9 reductive metabolism. QR activity was assayed under optimal conditions by following cytochrome c reduction at 550 nm in the presence of enzyme, quinone substrate, NADH, and bovine albumin, and confirmed by loss of EO9 absorbance at 550 nm. Both the rat and human tumor cell enzymes catalyzed reduction of the benchmark quinone menadione with a similar Km of 1.4-3.1 microM, although the Vmax was 7 to 8-fold lower for the human preparation. EO9 was readily reduced by the rat Walker QR. The mean Km was about 5-fold higher than for menadione at around 15 microM and the Vmax was 6-fold lower at around 2.5 mumol of cytochrome c reduced mg-1 of protein. EO9 was also metabolized by QR from HT29 human colon carcinoma cells but rather less efficiently than by the rat tumor enzyme. For example, the rate was 6-fold lower than that for the Walker tumor enzyme at 100 microM substrate concentration after correcting for the 7- to 8-fold difference in specific activity for the two preparations.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: Animals; Antineoplastic Agents; Aziridines; Biotransformation; Carcinoma 256, Walker; Colonic Neoplasms; Dicumarol; DNA Damage; DNA, Bacterial; Humans; Indolequinones; Indoles; Kinetics; Mitomycin; Mitomycins; NAD(P)H Dehydrogenase (Quinone); Oxidation-Reduction; Quinone Reductases; Quinones; Rats; Superoxide Dismutase; Tumor Cells, Cultured; Vitamin K

Levels of glutathione (GSH) in tumour tissue may be important in determining the clinical response to certain anticancer agents. Recent reports have suggested that D,L-buthionine-S,R-sulphoximine (BSO), a specific inhibitor of GSH synthesis, may be used to deplete tumour cell GSH and thus increase the therapeutic ratio of these agents. We have previously shown that 1-naphthol is a potential antitumour agent, and that its possible metabolite 1,4-naphthoquinone is thiol reactive and capable of redox cycling. It was therefore of interest to investigate the effect of pretreatment with BSO, on the toxicity of these agents, to tumour cells. For comparison we included three other cytotoxic agents, melphalan, helenalin and menadione, the toxicities of which are reported to be modulated by intracellular GSH. Depletion of GSH using BSO did not effect the toxicity of 1-naphthol, or 1,4-NQ but did produce slight potentiation of the cytotoxicities of menadione, helanalin and melphalan. The lack of effect of BSO on 1-naphthol and 1,4-NQ is not easily explained but if one also considers the modest potentiation of cytotoxicity+ achieved with the other agents studied, the potential use of BSO in combined chemotherapy is at best rather modest.

Topics: Antimetabolites; Antineoplastic Agents, Phytogenic; Buthionine Sulfoximine; Cell Line; Cell Survival; Colonic Neoplasms; Drug Evaluation, Preclinical; Glutathione; Humans; Melphalan; Methionine Sulfoximine; Naphthols; Naphthoquinones; Sesquiterpenes; Sesquiterpenes, Guaiane; Vitamin K

Topics: Antineoplastic Agents; Colonic Neoplasms; Energy Transfer; Humans; Melanoma; Naphthols; Neoplasms; Pancreatic Neoplasms; Skin Neoplasms; Vitamin K

Topics: Animals; Cholesterol, Dietary; Colon; Colonic Neoplasms; Diet; Dietary Fats; Disease Models, Animal; Feeding Behavior; Humans; Rectal Neoplasms; Vitamin K

An attempt has been made to develop tritiated derivatives of Synkavit (menadiol sodium diphosphate, MNDP) of high specific activity as a radioactive drug.This paper summarizes the preliminary biological and physical studies, with emphasis on approximate radiation dosimetry and the necessary preliminary testing, and then gives an account of the clinical investigations and the trials carried out so far, which correspond essentially to Phases I and II trials for a chemotherapeutic agent.In all, 214 patients with different sites and types of advanced and recurrent, inoperable, histologically verified malignant tumours including reticuloses have been treated with doses of at least 1 Ci of the various preparations. Among the 203 evaluable treated cases, some form of response was observed in 23 out of 151 (15·2%) receiving the drug by intravenous injections and 13 out of 52 (25%) after intra-arterial injections. For the sites and types of malignant diseases which showed responses after either intravenous or intra-arterial administration among the 55 patients surviving at least 3 months after the first injection, some form of response was observed in 32 but only 5 of these showed either a "complete" or a "partial" response.It is concluded that further investigation is desirable. It is suggested that clinical trials with randomization should be carried out for inoperable cases of carcinoma of the colon and of the pancreas.

Topics: Adult; Aged; Animals; Breast Neoplasms; Carcinoma 256, Walker; Colonic Neoplasms; Culture Techniques; Dose-Response Relationship, Radiation; Female; Half-Life; Hodgkin Disease; Humans; Injections, Intra-Arterial; Injections, Intraperitoneal; Injections, Intravenous; Male; Methods; Middle Aged; Neoplasms; Ovarian Neoplasms; Palatal Neoplasms; Radionuclide Imaging; Rectal Neoplasms; Testicular Neoplasms; Tongue Neoplasms; Tritium; Vitamin K

Topics: Adult; Antineoplastic Agents; Arsenicals; Breast Neoplasms; Carcinoma, Bronchogenic; Carcinoma, Squamous Cell; Choriocarcinoma; Colonic Neoplasms; Drug Synergism; Female; Fluorides; Heparin; Humans; Lung Neoplasms; Male; Malonates; Melanoma; Neoplasms; Ovarian Neoplasms; Pharyngeal Neoplasms; Pregnancy; Rectal Neoplasms; Sarcoma; Stomach Neoplasms; Testicular Neoplasms; Thyroid Neoplasms; Time Factors; Vitamin K

Topics: Adenocarcinoma; Adult; Aged; Animals; Breast Neoplasms; Carcinoma; Colonic Neoplasms; Cystadenoma; Female; Humans; Iodine Radioisotopes; Kidney Neoplasms; Lymphoma, Large B-Cell, Diffuse; Male; Mice; Middle Aged; Mouth Neoplasms; Neoplasm Metastasis; Neoplasms; Neoplasms, Experimental; Peritoneal Neoplasms; Radiation-Sensitizing Agents; Radionuclide Imaging; Rats; Rectal Neoplasms; Sigmoid Neoplasms; Stomach Neoplasms; Vitamin K

Topics: Adenocarcinoma; Autoradiography; Colonic Neoplasms; Dysgerminoma; Female; Gallbladder Neoplasms; Hodgkin Disease; Humans; Injections, Intra-Arterial; Injections, Intravenous; Melanoma; Neoplasm Metastasis; Neoplasms; Ovarian Neoplasms; Radiotherapy Dosage; Skin Neoplasms; Tritium; Vitamin K