PURPOSE: The growth regulatory effect of retinoid derivatives could be mediated by the transcriptional inactivation of AP-1 oncogenic transcription factor. By using ovarian cancer cell lines we were to investigate the cross-regulation mechanism between retinoids and AP-1. MATERIALS AND METHODS: Cell proliferation assays were performed in 4 ovarian cancer cells (A2774, PA-1, OVCAR-3, SKOV-3) by increasing the concentrations of all-trans retinoic acid (ATRA), 9-cis retinoic acid (9RA), 13-cis RA (13RA), 4-hydroxyphenyl retinamide (4-HPR). Transient transfection and CAT ELISA were done to determine the selective activity of each retinoid on the RAR (alpha, beta, gamma), RXR (alpha, beta, gamma). and the negative activity on AP-1 (c-Jun). RESULTS: Antiproliferative effect of 4-HPR (IC50; 0.7~2.7 micrometer) was more potent than those of other retinoid derivatives (IC50; 2.7~9.0 micrometer). To assess the anticancer mechanism, we examined the effect of 4-HPR on the transriptional activity of retinoic acid receptors (RAR/RXR) and of c-jun. Contrary to other retinoid derivatives that are active for RAR and RXR with some different levels, 4-HPR showed weak activity only for RARgamma. However, 4-HPR exerted the strongest suppression on AP-1 (c-Jun) activity. CONCLUSION: Based on our results showing much 4-HPR's potent antiproliferative activity coupled with the most effectively inhibiting activity on AP-1 and minimum activity on RA receptor (selective for RARgamma) than other retinoid derivatives, we suggest that 4-HPR may be a novel, and very effective anticancer drugs for ovarian cancer.
Animals ; Cats ; Cell Line ; Cell Proliferation ; Enzyme-Linked Immunosorbent Assay ; Fenretinide ; Ovarian Neoplasms* ; Receptors, Retinoic Acid ; Retinoid X Receptors ; Retinoids ; Transcription Factor AP-1* ; Transcription Factors ; Transfection ; Tretinoin

Retinoids play an important role in growth, reproduction and differentiation. Recently, retinoids have been used to both protect and treat from various animal models of carcinogenesis. In this study the effect of N-(4-hydroxyphenyl) retinamide (fenretinide) on viability of human neuroblastoma cell lines were evaluated. For the evaluation of apoptosis of human neuroblastoma cell lines by fenretinide. MTT assay, cytoplasmic DNA fragmentation, TUNEL stain, and Western blot analysis were performed. In MTT assay, fenretinide inhibited the proliferation of CHP134, IMR32 and SH-SY5Y but not in PC12 cells. Cytoplasmic DNA fragmentation was induced by treament of fenretinide (10 micrometer) for 48 h in IMR32 cells. PARP cleavage was detected by Western blot analysis after 16 h of treatment of fenretinide in CHP134, IMR32 and SH-SY5Y. These fenretinide effects on growth inhibition and increased apoptosis followed to the time dependent manner. The fenretinide treatment did not affect the phosphorylation of MAP kinases (ERK, JNK, p38). There was no change of Bcl-x and Bad expression after treatment of fenretinide (1 micrometer) in neroblastoma cell lines. Pretreatement of PD98059, SB203580, LY294002, or genistein also did not affect fenretinide-induced PARP cleavage in neuroblastoma cell lines. From these results, the fenretinide-induced apoptosis is due to the PARP cleavage which occured MAP kinase signal cascades independently.
Animals ; Apoptosis* ; Blotting, Western ; Carcinogenesis ; Cell Line* ; Cytoplasm ; DNA Fragmentation ; Fenretinide* ; Genistein ; Humans* ; In Situ Nick-End Labeling ; Models, Animal ; Neuroblastoma* ; PC12 Cells ; Phosphorylation ; Phosphotransferases ; Reproduction ; Retinoids

Animals ; Antineoplastic Agents ; therapeutic use ; Breast Neoplasms ; drug therapy ; genetics ; metabolism ; Cyclin-Dependent Kinase Inhibitor p27 ; Fenretinide ; therapeutic use ; Hepatocyte Nuclear Factor 3-alpha ; genetics ; metabolism ; Humans ; Intracellular Signaling Peptides and Proteins ; metabolism ; Prognosis ; Receptors, Estrogen ; metabolism

The retinoid N-4-hydroxyphenyl retinamide (4-HPR also known as fenretinide), a synthetic derivative of all trans retinoic acid (ATRA), has shown as an efficient chemopreventive, chemotherapeutic agent and a potent inducer of apoptosis in various cancer cell types in vitro, including leukemic cells. However the mechanisms by which 4-HPR has the apoptotic effects is not completely elucidated. This study was aimed to investigate the effect of 4-HPR on several leukemic cells and explore its mechanisms of effect on U937 cells. The cell growth and proliferation experiments were performed [corrected] cell apoptosis was detected by annexin V; reactive oxygen species (ROS) and mitochondrial transmembrane potential (DeltaPsim) were determined; protein [corrected] expression was detected by Western blot. The results showed that 4-HPR inhibited the proliferation of U937 cells in a dose- and time-dependent manner. 4-HPR markedly [corrected] induced apoptosis in U937 cells, triggered the generation of ROS, induced the loss of mitochondrial transmembrane potential, decreased the expression of procaspase-8 and procaspase-3. Pretreatment of L-ascorbic acid suppressed the generation of ROS, disruption of mitochondrial potential, activation of caspases and apoptosis. It is concluded that the generation of ROS followed by the disruption of mitochondrial transmembrane potential plays an important role on 4-HPR-induced apoptosis in leukemic cells, suggesting that 4-HPR may be one of mitochondrial-targeted agents with clinical potential in treating cancer.
Antineoplastic Agents ; pharmacology ; Apoptosis ; drug effects ; Blotting, Western ; Caspases ; metabolism ; Dose-Response Relationship, Drug ; Fenretinide ; pharmacology ; HL-60 Cells ; Humans ; K562 Cells ; Leukemia ; metabolism ; pathology ; Membrane Potential, Mitochondrial ; drug effects ; Reactive Oxygen Species ; metabolism ; U937 Cells

OBJECTIVETo study the mechanism of the apoptosis induced by N-[4-hydroxyphenyl] retinamide (4-HPR) in bladder cancer cell line T24, and the involvement of DNA damage and repair.

METHODST24 cells were treated with 4-HPR at the concentration of 2.5, 5.0 and 10.0 micromol/L, and the cell grow inhibition was measured by cell counting assay. The fluorescent intensity of reactive oxygen species (ROS) was determined by spectrofluorometer. The apoptosis was measured by flow cytometry and DNA fragment assay. The expression of XRCC1 protein and activation of caspase-3 were detected by Western blot.

RESULTS4-HPR induced apoptosis in T24 cell. A dose-dependent increase in the percentage of apoptosis cells was observed (1.8%, 4.0% and 10.5% respectively at 2.5, 5.0, 10.0 micromol/L 4HPR). In the meantime, ROS level in the cell was increased (peaked at 3 fold). It also caused down-regulation of the expression of XRCC1, and activation of caspase-3. Vitamin C effectively inhibited ROS rise induced by 4-HPR, and also partially inhibited cell growth, apoptosis, and down-regulation of the expression of XRCC1.

CONCLUSIONThe generation of ROS and DNA damage may be the major mechanism of the apoptosis of bladder cancer cell line T24 induced by 4-HPR.

Antineoplastic Agents ; pharmacology ; Cell Line, Tumor ; DNA Damage ; DNA Repair ; DNA-Binding Proteins ; metabolism ; Fenretinide ; pharmacology ; Humans ; Reactive Oxygen Species ; metabolism ; Urinary Bladder Neoplasms ; metabolism ; pathology ; X-ray Repair Cross Complementing Protein 1

The growth inhibitory effects of four retinoic acid (RA) derivatives, 9-cis RA, 13-cis RA, N-(4-hydroxyphenyl) retinamide (4-HPR), and all-trans retinoic acid (ATRA) were compared. In addition, the effects of various combinations of these four agents were examined on non-small cell lung carcinoma (NSCLC) cell-lines, and on the expressions of retinoic acid receptors (RARs) and retinoid X receptors (RXRs) on these cells. At the clinically achievable concentration of 1 micrometer, only 4-HPR inhibited the growths of H1299 and H460 cells-lines. However, retinoic acid receptor beta(RAR beta) expression was up-regulated on H460 and H1299 cells treated with 1 micrometer of ATRA, 13-cis RA, or 9-cis RA. All NSCLC cell lines showed growth inhibition when exposed sequentially to 1 micrometer ATRA and 0.1 micrometer 4-HPR. In particular, sequential treatment with 1 micrometer ATRA or 13-cis RA and 4-HPR markedly inhibited H1703 cell growth; these cells exhibited no basal RAR beta expression and were refractory to 4-HPR. However, in NSCLC cell lines that expressed RAR beta, the expressional levels of RAR beta were up-regulated by ATRA alone and by sequential treatment with ATRA and 4-HPR. 4-HPR was found to be the most active of the four agents in terms of NSCLC growth-inhibition. Moreover, sequential treatments with ATRA or 13-cis RA followed by 4-HPR were found to have synergistic growth-inhibitory effects and to regulate RAR expression.
Base Sequence ; Carcinoma, Non-Small-Cell Lung/*drug therapy/genetics/metabolism ; Cell Line, Tumor ; DNA Primers/genetics ; Drug Therapy, Combination ; Fenretinide/administration & dosage ; Gene Expression/drug effects ; Humans ; Isotretinoin/administration & dosage ; Lung Neoplasms/*drug therapy/genetics/metabolism ; Receptors, Retinoic Acid/genetics ; Retinoid X Receptors/genetics ; Tretinoin/administration & dosage/*analogs & derivatives