ascorbic-acid and motexafin-gadolinium

Redox regulation has been shown to be an important component of malignant cell survival. Tipping the cellular redox balance through pharmacologic regulation in favor of increasing intracellular reactive oxygen species (ROS) and/or depleting protective reducing metabolites (such as glutathione and nicotinamide adenine dinucleotide phosphate) may lead to oxidative stress and resultant induction of apoptosis for the treatment of cancer. We review the biology and importance of ROS with regard to malignant and normal cells. Moreover, we discuss pre-clinical and clinical data regarding novel therapeutic agents that modulate the cellular redox system including buthionine sulfoximine, ascorbic acid, arsenic trioxide, imexon, and motexafin gadolinium as single-agents and in combination. Continued research is needed to better understand the mechanisms and specific apoptotic pathways involved in ROS-induced cell death, as well as, to determine the most rationale and effective combination of redox-active agents.

Topics: Animals; Antimetabolites, Antineoplastic; Antineoplastic Combined Chemotherapy Protocols; Apoptosis; Arsenic Trioxide; Arsenicals; Ascorbic Acid; Buthionine Sulfoximine; Cell Death; Dose-Response Relationship, Drug; Glutathione; Hexanones; Humans; Metalloporphyrins; Models, Biological; Models, Chemical; Neoplasms; Oxidation-Reduction; Oxidative Stress; Oxides; Reactive Oxygen Species

Motexafin gadolinium (MGd) sensitizes malignant cells to ionizing radiation, although the underlying mechanisms for uptake and sensitization are both unclear. Here we show that MGd is endocytosed by the clathrin-dependent pathway with ensuing lysosomal membrane permeabilization, most likely via formation of reactive oxygen species involving redox-active metabolites, such as ascorbate. We propose that subsequent apoptosis is a synergistic effect of irradiation and high MGd concentrations in malignant cells due to their pronounced endocytic activity. The results provide novel insights into the mode of action of this promising anti-cancer drug, which is currently under clinical trials.

Topics: Antineoplastic Agents; Ascorbic Acid; Cell Line; Endocytosis; Flow Cytometry; Humans; Lysosomes; Metalloporphyrins; Microscopy, Fluorescence; Reactive Oxygen Species

Loss of p53 renders cells more susceptible to acute oxidant stress induced by oxidant-generating agents such as motexafin gadolinium (MGd). We hypothesized that reactive oxygen species (ROS)-generating MGd results in low-level p53 expression, making cells more susceptible to oxidant stress.. Lymphoma cells were incubated with different concentrations of MGd with or without zinc (Zn) and ascorbate, and ROS, apoptosis, proteins, and oxidant genes were measured.. MGd, with ascorbate and Zn, induced apoptosis in lymphoma cells. This was accompanied by reduction of p53 protein but not message, and by reduction of p53 downstream targets p21, glutathione peroxidase 1 (GPx1), and p53 up-regulated modulator of apoptosis (PUMA). p53 protein reduction was reversed by MG132, and nutlin-3.. Our data are consistent with a pathway of cell death that is independent of p53-mediated induction of PUMA; the cellular response to reduce p53 represents a cell survival adjustment to ROS-mediated stress.

Topics: Apoptosis; Ascorbic Acid; Burkitt Lymphoma; Cell Line, Tumor; Gene Expression; Humans; Imidazoles; Leupeptins; Lymphoma, Follicular; Metalloporphyrins; Piperazines; Proto-Oncogene Proteins c-mdm2; Reactive Oxygen Species; Tumor Suppressor Protein p53; Zinc

To investigate the effects of motexafin gadolinium (MGd) on the levels of reactive oxygen species (ROS), glutathione (GSH), and DNA damage in EMT6 mouse mammary carcinoma cells. The ability of MGd to alter radiosensitivity and to inhibit DNA damage repair after X-ray irradiation was also evaluated.. Reactive oxygen species and GSH levels were assessed by 2,7-dichlorofluorescein fluorescence flow cytometry and the Tietze method, respectively. Cellular radiosensitivity was assessed by clonogenic assays. Deoxyribonucleic acid damage and DNA damage repair were assessed in plateau-phase EMT6 cells by the Comet assay and clonogenic assays.. Cells treated with 100 mumol/L MGd plus equimolar ascorbic acid (AA) had significantly increased levels of ROS and a 58.9% +/- 3.4% decrease in GSH levels, relative to controls. Motexafin gadolinium plus AA treatment increased the hypoxic, but not the aerobic, radiosensitivity of EMT6 cells. There were increased levels of single-strand breaks in cells treated with 100 mumol/L MGd plus equimolar AA, as evidenced by changes in the alkaline tail moment (MGd + AA, 6 h: 14.7 +/- 1.8; control: 2.8 +/- 0.9). The level of single-strand breaks was dependent on the length of treatment. Motexafin gadolinium plus AA did not increase double-strand breaks. The repair of single-strand breaks at 2 h, but not at 4 h and 6 h, after irradiation was altered significantly in cells treated with MGd plus AA (MGd + AA, 2 h: 15.8 +/- 3.4; control: 5.8 +/- 0.6). Motexafin gadolinium did not alter the repair of double-strand breaks at any time after irradiation with 10 Gy.. Motexafin gadolinium plus AA generated ROS, which in turn altered GSH homeostasis and induced DNA strand breaks. The MGd plus AA-mediated alteration of GSH levels increased the hypoxic, but not aerobic, radiosensitivity of EMT6 cells. Motexafin gadolinium altered the kinetics of single-strand break repair soon after irradiation but did not inhibit potentially lethal damage repair in EMT6 cells.

Topics: Animals; Ascorbic Acid; Cell Hypoxia; Cell Line, Tumor; Cell Survival; Comet Assay; DNA Damage; DNA Repair; Glutathione; Metalloporphyrins; Mice; Photosensitizing Agents; Radiation Tolerance; Reactive Oxygen Species; Time Factors

Motexafin gadolinium (MGd) oxidizes ascorbate, in neutral buffer and in cell culture, forming reactive oxygen species and a coordination polymer with oxalate.

Topics: Ascorbic Acid; Metalloporphyrins; Molecular Structure; Oxalic Acid; Photosensitizing Agents; Polymers; Reactive Oxygen Species; Tumor Cells, Cultured

To examine the mechanism of radiation enhancement by motexafin gadolinium (Gd-Tex) in vitro.. Oxidation of ascorbate and NADPH by Gd-Tex was evaluated in a neutral buffer. Growth inhibition of human uterine cancer cell line MES-SA was measured using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) dye. Clonogenic assays were used to measure radiation response in MES-SA, A549 human lung carcinoma, E89, a CHO cell line variant deficient in glucose-6-phosphate dehydrogenase activity, and murine lymphoma cell lines LYAR and LYAS.. Gd-Tex catalyzed the oxidation of NADPH and ascorbate under aerobic conditions, forming hydrogen peroxide. Decreased viability was observed in MES-SA cells incubated with Gd-Tex in media containing NADPH or ascorbate. Gd-Tex and ascorbate increased fluorescence in dichlorofluorescin acetate-treated cultures. Synergistic effects on the aerobic radiation response in MES-SA and A549 were seen using Gd-Tex in combination with L-buthionine-(S,R)-sulfoximine (BSO). Incubation with Gd-Tex in the presence of ascorbate increased the aerobic radiation response of E89 and the apoptosis-sensitive B-cell line (LYAS).. Gd-Tex sensitizes cells to ionizing radiation by increasing oxidative stress as a consequence of futile redox cycling. Optimization of the concentration of ascorbate (or other reducing species) may be required when evaluating Gd-Tex activity in vitro.

Topics: Animals; Antineoplastic Agents; Ascorbic Acid; CHO Cells; Cricetinae; Female; Humans; Hydrogen Peroxide; Metalloporphyrins; NADP; Oxidation-Reduction; Reactive Oxygen Species; Tumor Cells, Cultured; Uterine Neoplasms