Abstract
We have previously reported that HET0016 [N-hydroxy-N′-(4-butyl-2 methylphenyl)formamidine], a selective inhibitor of CYP4A and thus 20-HETE (20-hydroxyeicosatetraenoic acid) synthesis, inhibits endothelial cell proliferation and decreases angiogenesis induced by human glioma cell U251. A stable 20-HETE agonist, WIT003 [20-hydroxyeicosa-5(Z),14(Z)-dienoic acid (1 μM)], increased U251 cell proliferation from 3.9- to 4.8-folds from T0 (time of the treatment). We examined the effects of HET0016 on the growth of U251. HET0016 inhibited U251 basal cell proliferation in a dose-dependent manner. 10 μM HET0016 suppressed 56% of U251 proliferation and significantly increased the proportions of the cells arrested in the G0/G1 phase of the cell cycle. Exposure to HET0016 (as early as 4 h) reduced protein tyrosine and p42/p44 MAPK (mitogen-activated protein kinase) phosphorylation. Furthermore, HET0016 significantly inhibited the U251 proliferation and phosphorylation of both the epidermal growth factor (EGF) receptor and p42/p44 MAPK induced by EGF. CYP4A mRNA and proteins were both present in U251. This suggests that HET0016 inhibited U251 proliferation by inhibiting 20-HETE synthesis. However, U251 did not synthesize 20-HETE in the presence of arachidonic acid. This implies that HET0016 suppresses U251 proliferation by mechanisms that are not yet clear but may involve activities other than inhibition of 20-HETE synthesis. We concluded that HET0016 may be the prototype of novel compounds that suppress human glioma cell proliferation.
Footnotes
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This work was supported by National Institutes of Health Grants EY014385 (to A.G.S.), GM31278 (to J.R.F.), and HL 036279 (to R.J.R.) and a grant from the Robert A. Welch Foundation (to J.R.F.). This study has been presented in part at the 2004 American Association for Cancer Research Annual Meeting (Mar 21–31), Orlando, FL.
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Article, publication date, and citation information can be found at http://jpet.aspetjournals.org.
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doi:10.1124/jpet.105.088567.
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ABBREVIATIONS: AA, arachidonic acid; P450, cytochrome P450; 20-HETE, 20-hydroxyeicosatetraenoic acid; HET0016, N-hydroxy-N′-(4-butyl-2-methylphenyl)formamidine; WIT003, 20-hydroxyeicosa-5(Z),14(Z)-dienoic acid; DDMS, dibromododecenyl methylsulfonimide; EEZE, 14,15-epoxyeicosa-5(Z)-enoic-methyl sulfonylimide; EET, epoxyeicosatrienoic acid; MAPK, mitogen-activated protein kinase; EGF, epidermal growth factor; EGFR, epidermal growth factor receptor; HUVEC, human umbilical vein endothelial cell(s); SAPK, stress-activated protein kinase; JNK, c-Jun NH2-terminal kinase; EtOH, ethanol; PBS, phosphate-buffered saline; PCR, polymerase chain reaction; TUNEL, terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling; GBM, glioblastoma multiforme.
- Received May 6, 2005.
- Accepted August 2, 2005.
- The American Society for Pharmacology and Experimental Therapeutics
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