epidermal-growth-factor and 13-hydroxy-9-11-octadecadienoic-acid

Androgens as well as monohydroxy-fatty acids are implicated in the pathogenesis of prostate cancer. Like a huge variety of endo- and xenobiotics, they are eliminated as glucuronide conjugates formed by uridine diphosphate-glucuronosyltransferase (UGT) enzymes. In the present study, we observe that treatment of the prostate cancer cells LNCaP with natural and synthetic androgens, IL-1alpha, or epidermal growth factor (EGF) differently modulates the glucuronidation of androgen and bioactive lipid metabolites. Indeed, glucuronidation of 5alpha-androstane-3alpha,17beta-diol and 13-hydroxyoctadecadienoic acid was drastically reduced, whereas 12-hydroxyeicosatetraenoic acid conjugation by UGT was increased after androgen treatment. These effects reflected the reduction of UGT2B10, -B15, and -B17 enzyme expression, and the activation of the UGT2B11 gene. In human prostate epithelial cells, only UGT2B11 and -B15 mRNAs are detected and are regulated by androgens in a similar manner as in LNCaP cells. In LNCaP cells, IL-1alpha and EGF also regulate UGT2B expression in an isoform-specific manner; IL-1alpha induced UGT2B10 and reduced UGT2B17, while having no effects on UGT2B11 mRNA levels. EGF treatment resulted in a decreased UGT2B17 expression, whereas UGT2B10 and -B11 mRNA remained at their basal levels. Overall, these results demonstrate that in the human prostate, androgens do not only affect their own inactivation but also influence the levels of monohydroxy-fatty acids by regulating the expression of UGT2B enzymes in an isoform-specific manner. These differential effects of androgens, IL-1alpha, and EGF on lipid metabolism likely constitute an additional mechanism by which these endogenous factors promote prostate cancer development.

Topics: 12-Hydroxy-5,8,10,14-eicosatetraenoic Acid; Androstane-3,17-diol; Cells, Cultured; Dose-Response Relationship, Drug; Epidermal Growth Factor; Glucuronides; Glucuronosyltransferase; Humans; Hydroxyeicosatetraenoic Acids; Interleukin-1; Linoleic Acids; Lipid Metabolism; Male; Metribolone; Minor Histocompatibility Antigens; Prostate; Receptors, Androgen; RNA, Messenger

Human prostate tumors have elevated levels of 15-lipoxygenase-1 (15-LOX-1) and data suggest that 15-LOX-1 may play a role in the development of prostate cancer. In contrast, 15-LOX-2 expression is higher in normal rather than in tumor prostate tissue and appears to suppress cancer development. We recently reported that 13-(S)-HODE, the 15-LOX-1 metabolite, up-regulates the MAP kinase signaling pathway and subsequently down-regulates PPARgamma in human colorectal carcinoma cells. To determine whether this mechanism is applicable to prostate cancer and what the effects of 15-LOX-2 are, we investigated the effect of 15-LOX-1, 15-LOX-2, and their metabolites on epidermal growth factor (EGF)- and insulin-like growth factor (IGF)-1 signaling in prostate carcinoma cells. In PC3 cells, 13-(S)-HODE, a 15-LOX-1 metabolite, up-regulated MAP kinase while in contrast 15-(S)-HETE, a 15-LOX-2 metabolite, down-regulated MAP kinase. As a result, 13-(S)-HODE increased PPARgamma phosphorylation while a subsequent decrease in PPARgamma phosphorylation was observed with 15-(S)-HETE. Thus, 15-LOX metabolites have opposing effects on the regulation of the MAP kinase signaling pathway and a downstream target of MAP kinase signaling like PPARgamma. In addition to the EGF signaling pathway, the IGF signaling pathway appears to be linked to prostate cancer. 13-(S)-HODE and 15-(S)-HETE up-regulate or down-regulate, respectively, both the MAPK and Akt pathways after activation with IGF-1. Thus, the effect of these lipid metabolites is not solely restricted to EGF signaling and not solely restricted to MAPK signaling. These results provide a plausible mechanism to explain the apparent opposing effects 15-LOX-1 and 15-LOX-2 play in prostate cancer.

Topics: Arachidonate 15-Lipoxygenase; Epidermal Growth Factor; Humans; Hydroxyeicosatetraenoic Acids; Isoenzymes; Linoleic Acids; Male; Mitogen-Activated Protein Kinases; Phosphorylation; Prostate; Prostatic Neoplasms; Receptors, Cytoplasmic and Nuclear; Signal Transduction; Transcription Factors; Tumor Cells, Cultured

Epidermal growth factor (EGF) stimulates the lipoxygenase metabolism of linoleic acid to 13(S)-hydroxyoctadecadienoic acid (HODE) in Syrian hamster embryo (SHE) fibroblasts. 13(S)-HODE is a potent and specific enhancer of EGF-dependent DNA synthesis in normal phenotypic SHE cells (supB+), but is inactive in variant SHE cells that have lost tumor suppressor gene function (supB-). EGF activation of quiescent SHE cells results in increased levels of 13-HODE esterified in cellular phospholipid and triglyceride. Steric analyses suggest that this metabolite is generated in part by direct oxygenation of membrane lipids by an n-6 lipoxygenase. In studies on the uptake and mobilization of 13-HODE in SHE cells, we observed EGF to stimulate a time- and dose-dependent incorporation and reacylation of the mono-hydroxy linoleate metabolite. The level of 13-HODE uptake in supB+ cells is twice that of supB-. Among classes of phospholipids, radiolabeled 13-HODE is esterified predominantly into phosphatidylcholine and this distribution pattern is similar for both SHE cell lines. Pretreatment of cells with the tyrosine kinase inhibitor methyl-2,5-dihydroxycinnamate blocks EGF-stimulated HODE incorporation. Inhibition of tyrosine phosphatase activity with vanadate potentiates HODE uptake in supB+ but not supB- cells. Moreover, activation of protein kinase C with phorbol ester stimulates HODE incorporation in the supB+ line only. The differential effects of EGF on 13-HODE uptake and mobilization in supB+ and supB- cells appear to be related to loss of the tumor suppressor phenotype. EGF-stimulated generation of esterified 13-HODE may be an important biological process in determining the mechanism and site of HODE interaction with the mitogenic signaling pathway.

Topics: Animals; Cell Line; Chemical Fractionation; Cricetinae; Embryo, Mammalian; Epidermal Growth Factor; Esterification; Fibroblasts; Genes, Tumor Suppressor; Linoleic Acids; Lipids; Mesocricetus; Phenotype; Phospholipids; Protein-Tyrosine Kinases; Stimulation, Chemical

Epidemiological and experimental data suggest a role for polyunsaturated fatty acids in the etiology of breast cancer. In this report we have studied arachidonic acid and linoleic acid metabolism in the human breast carcinoma cell line BT-20 which overexpresses both EGF receptor and the homologous erbB-2 oncogene product. EGF and TGF alpha stimulated DNA synthesis in these cells which was attenuated by the addition of a lipoxygenase inhibitor, NDGA. The addition of a prostaglandin H synthase inhibitor did not alter DNA synthesis. Analytical studies reveal little arachidonic acid metabolism while linoleic acid was metabolized to 13-hydroxyoctadecadienoic acid (13-HODE). The formation of 13-HODE was inhibited by the addition of NDGA and was dependent on EGF or TGF alpha. These results suggest the metabolism of linoleic acid by a n-6 or 15-lipoxygenase regulated by EGF/TGF alpha, RT-PCR was used to isolate a clone, and sequenced the cDNA for this enzyme and it was found to be identical to the human 15-lipoxygenase previously characterized from human pulmonary tissue. EGF/TGF alpha did not alter the expression of this enzyme suggesting a potential post-translational regulation of activity. This study provides a link between metabolism of linoleic acid and growth factor regulation of cell proliferation in a human breast carcinoma cell line.

Topics: Arachidonate 15-Lipoxygenase; Arachidonic Acid; Breast Neoplasms; Cyclooxygenase Inhibitors; DNA; Epidermal Growth Factor; Female; Humans; Indomethacin; Linoleic Acid; Linoleic Acids; Lipoxygenase Inhibitors; Masoprocol; Polymerase Chain Reaction; Prostaglandin-Endoperoxide Synthases; RNA, Messenger; Transforming Growth Factor alpha; Tumor Cells, Cultured

Topics: Animals; Calcium-Calmodulin-Dependent Protein Kinases; Cell Division; Cell Line, Transformed; Cell Transformation, Neoplastic; Cricetinae; Embryo, Mammalian; Enzyme Activation; Epidermal Growth Factor; ErbB Receptors; Fibroblasts; Hybrid Cells; Linoleic Acid; Linoleic Acids; Lipid Peroxides; Mesocricetus; Phosphorylation; Protein Processing, Post-Translational; Signal Transduction; Stimulation, Chemical

Topics: Animals; Arachidonic Acid; Cell Division; Cell Line, Transformed; Cell Transformation, Neoplastic; Cricetinae; Cyclooxygenase Inhibitors; Dexamethasone; DNA Replication; Epidermal Growth Factor; Fibroblasts; Hydroxyeicosatetraenoic Acids; Indomethacin; Linoleic Acid; Linoleic Acids; Lipoxygenase; Masoprocol; Mesocricetus; Receptor, ErbB-2; Recombinant Fusion Proteins; Transfection

Topics: Animals; Arachidonic Acid; Cell Division; Cells, Cultured; Cricetinae; Cyclooxygenase 1; Cyclooxygenase 2; Embryo, Mammalian; Epidermal Growth Factor; Fibroblasts; Isoenzymes; Linoleic Acid; Linoleic Acids; Linoleic Acids, Conjugated; Mesocricetus; Prostaglandin-Endoperoxide Synthases; RNA, Messenger; Transcription, Genetic

Topics: 12-Hydroxy-5,8,10,14-eicosatetraenoic Acid; Animals; Cell Division; Cell Line; Cricetinae; DNA; DNA Replication; Embryo, Mammalian; Epidermal Growth Factor; Hydroxyeicosatetraenoic Acids; Linoleic Acids; Linoleic Acids, Conjugated; Mesocricetus; Oxidation-Reduction; Thymidine

Metabolism of arachidonic and linoleic acid can be regulated by polypeptide growth factors in a variety of cell types. In Syrian hamster embryo (SHE) fibroblasts, epidermal growth factor (EGF) stimulates the conversion of exogenous linoleic acid to 13(S)-hydroxyoctadecadienoic acid (HODE). Inhibition of 13-HODE biosynthesis blocks the EGF-mitogenic response in SHE cells, and 13-HODE and its hydroperoxy precursor are potent and highly specific enhancers of EGF-dependent DNA synthesis. We demonstrated that EGF stimulates a biphasic production and release of endogenous 13-HODE. Through development of a stable isotope-dilution GC/MS assay for 13-HODE, we observed 13-HODE production as early as 5 min after EGF stimulation, and this initial phase peaked at 1 hr. A second rise in 13-HODE formation was seen at 2-4 hr, and this phase plateaued at 4-6 hr at a level of 30-40 ng/10(6) cells. EGF stimulation of 13-HODE biosynthesis is not mediated by transcriptional or translational regulation of the inducible form of prostaglandin H synthase. Based on enzyme inhibitor studies and structural characterization of products, the linoleate metabolite is apparently formed by an n-6 lipoxygenase that remains to be characterized. EGF stimulation of 13-HODE formation is linked with activation of the EGF receptor tyrosine kinase. Inhibition of EGF receptor tyrosine kinase activity with methyl-2,5-dihydroxycinnamate blocked EGF-dependent linoleic acid metabolism and EGF-regulated DNA synthesis. Potentiation of the EGF receptor tyrosine phosphorylation cascade through treatment of SHE cells with the tyrosine phosphatase inhibitor vanadate resulted in a 3-fold increase in EGF-stimulated 13-HODE production and a corresponding enhancement of the EGF mitogenic response. The coupling of EGF-regulated linoleic acid metabolism with the EGF receptor tyrosine kinase activity suggests the importance of specific linoleate compounds in mediating mitogenic signal transduction.

Topics: Animals; Arachidonate 5-Lipoxygenase; Cells, Cultured; Cricetinae; Epidermal Growth Factor; ErbB Receptors; Fibroblasts; Linoleic Acids; Mesocricetus

Topics: Animals; Cell Division; Cell Line, Transformed; Cricetinae; Embryo, Mammalian; Epidermal Growth Factor; ErbB Receptors; Fibroblasts; Linoleic Acid; Linoleic Acids; Mesocricetus; Phosphorylation; Phosphotyrosine; Signal Transduction; Tyrosine

Epidermal growth factor induces the oxygenation of linoleic acid in Syrian hamster embryo fibroblasts, and the lipoxygenase-derived products potentiate the mitogenic signal. We have further characterized the linoleate metabolites of growth factor-activated cells by chiral phase HPLC analysis. The primary product was identified as the pure (S) enantiomer of 13-hydroxyoctadecadienoic acid (HODE). In comparison to 13(R)-HODE isomer, only the biologically derived 13(S)-HODE was active in augmenting DNA synthesis as assessed by [3H]thymidine incorporation. To extend these investigations, we defined the structural requirements of analogous lipid compounds necessary for stimulation of mitogenesis in these cells. Carbon-chain length, degree of unsaturation, type of oxidized functionality, position of oxygenated moiety, double-bond geometry, and chirality were all identified as factors that modulate the mitogenic activity of related compounds. The results demonstrate a high degree of specificity for (S)-isomer hydro(pero)xylinoleic acid metabolites in stimulating DNA synthesis and further define the relationship between linoleic acid metabolism and growth-factor-dependent cell growth.

Topics: Animals; Cell Division; Cell Line; Chromatography, High Pressure Liquid; Cricetinae; DNA; Dose-Response Relationship, Drug; Embryo, Mammalian; Epidermal Growth Factor; Linoleic Acid; Linoleic Acids; Mesocricetus; Stereoisomerism; Structure-Activity Relationship; Thymidine; Tritium

Epidermal growth factor (EGF) stimulates DNA synthesis in quiescent Syrian hamster embryo (SHE) cells. Work in the present authors' laboratory has shown that the formation of 9- and 13-hydroxyoctadecadienoic acid (HODEs), 15-lipoxygenase-derived metabolites of linoleic acid, are involved in the mitogenic response to EGF in these cells (Glasgow et al. (1992) J. Biol. Chem. 267, 10771-10779). SHE cells also produce prostaglandin E2 (PGE2) and prostaglandin F2 alpha (PGF2 alpha). We now report the effects of HODEs and prostaglandins on EGF-dependent expression of the growth-related proto-oncogene c-myc in SHE cells. Treatment of cells with eicosatetraynoic acid (ETYA), which blocks EGF-dependent HODE formation, inhibited the mitogenic response to EGF, while exogenous 13-HODE potentiated EGF-dependent DNA synthesis. However, neither ETYA or 13-HODE altered the accumulation of c-myc mRNA in response to EGF. In contrast, PGE2 inhibited EGF-induced DNA synthesis and down-regulated EGF-stimulated c-myc mRNA accumulation in a dose-dependent manner, whereas PGF2 alpha had no effect on these responses. PGE2, but not PGF2 alpha, induced a rapid increase in cAMP formation, and both forskolin and 8-(4-chlorophenylthio)-cAMP mimicked the inhibitory effects of PGE2 on EGF-dependent DNA synthesis and c-myc mRNA accumulation, suggesting that the involvement of cAMP. The results indicate that the modulation of EGF-dependent DNA synthesis by PGE2, but not by HODEs, is associated with altered expression of the proto-oncogene c-myc in SHE cells.

Topics: Animals; Cell Line; Cricetinae; Cricetulus; DNA; Epidermal Growth Factor; Genes, myc; Linoleic Acids; Linoleic Acids, Conjugated; Prostaglandins; Proto-Oncogene Proteins c-myc

Topics: Animals; Cell Division; Cell Line; DNA; Epidermal Growth Factor; Fibroblasts; Linoleic Acid; Linoleic Acids; Linoleic Acids, Conjugated; Lipoxygenase; Mice; Mice, Inbred BALB C; Signal Transduction