flavin-adenine-dinucleotide and Ovarian-Neoplasms

In the present work, metabolomic and redox proteomic analyses were carried out on an untreated- and gossypol-treated ovarian cancer cell line, SKOV3. Gossypol treatment resulted in cell death through oxidative stress. Metabolite analysis showed that gossypol induces a decrease of the cellular levels of GSH, aspartic acid, and FAD. Using a combination of double labeling and LC-MS-MS, we identified changes in thiol-redox states of 545 cysteine-containing peptides from 356 proteins. The frequently occurring amino acid residue immediately before or after the cysteine in these peptides is the non-polar and neutral leucine, valine, or alanine. These redox sensitive proteins participate in a variety of cellular processes. We have characterized the redox-sensitive cysteine residues in PKM2, HSP60, malate dehydrogenase and other proteins that play important roles in metabolism homeostasis and stress responses. The three cysteine residues of HSP60 exhibit different responses to gossypol treatment: an increase of thiol/disulfide ratio for the Cys447 residue due to a decrease of the cellular GSH level, and a decrease of thiol/disulfide ratios for Cys442 and Cys237 residues due to oxidation and sulfation. This study suggests that thiol/disulfide ratios are dependent on the level of cellular GSH. Our data provide a valuable resource for deciphering the redox regulation of proteins and for understanding gossypol-induced apoptosis in ovarian cancer cells.

Topics: Apoptosis; Aspartic Acid; Carrier Proteins; Cell Line, Tumor; Chaperonin 60; Contraceptive Agents, Male; Disulfides; Female; Flavin-Adenine Dinucleotide; Gene Expression Profiling; Glutathione; Gossypol; Humans; Membrane Proteins; Metabolomics; Mitochondrial Proteins; Ovarian Neoplasms; Oxidation-Reduction; Oxidative Stress; Proteome; Proteomics; Sulfhydryl Compounds; Thyroid Hormone-Binding Proteins; Thyroid Hormones

The objective of this study was to compare the effects of chemopreventive agents on natural fluorescence emission of ovarian cells in a cell culture and in a primate model as a feasibility trial to monitor drug activity.. Fluorescence emission spectra were collected from normal (NOE) and immortalized ovarian surface epithelial cells at 290, 360, and 450 nm excitation. Redox potentials were calculated and compared to % apoptosis and cell survival. Fluorescence emission spectra were collected from 18 female rhesus macaques receiving fenretinide [N-(-hydroxyphenyl)retinamide (4-HPR)] orally and/or oral contraceptive pills (OCP) or no medication. Fluorescence intensities and redox ratios were compared using a two-tailed Student's t test.. Apoptosis and cell survival correlated with fluorescence emission consistent with metabolically active proteins [flavin adenine dinucleotide (FAD) and nicotinamide adenine dinucleotide (NAD(P)H)] and the resulting redox ratio in cells grown with 4-HPR. The 4-HPR consistently inhibited cell survival in a dose dependent manner. Degree of correlation varied between different cell lines. In primates receiving 4-HPR, fluorescence emission was increased at 450 nm excitation, 550 nm emission consistent with FAD presence, whereas those receiving OCP showed decreased emission at 350 nm excitation, 450 nm emission consistent with decreased NAD(P)H presence. Redox ratios were increased by both drugs.. Fluorescence intensity and redox ratio appear to be altered by 4-HPR treatment in vivo and in cell culture and by OCP in vivo. Fluorescence intensity may be useful to monitor chemopreventive agents in clinical trials.

Topics: Animals; Anticarcinogenic Agents; Apoptosis; Biomarkers, Tumor; Cell Survival; Cells, Cultured; Contraceptives, Oral; Dose-Response Relationship, Drug; Epithelial Cells; Feasibility Studies; Female; Fenretinide; Flavin-Adenine Dinucleotide; Macaca mulatta; NADP; Ovarian Neoplasms; Ovary; Oxidation-Reduction; Reference Values; Spectrometry, Fluorescence