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Bottone Jr FG, Moon Y, Kim JS, Alston-Mills B, Ishibashi M, Eling TE.. The anti-invasive activity of cyclooxygenase inhibitors is regulated by the transcription factor ATF3 (activating transcription factor 3). Mol Cancer Ther 4: 693-703

Laboratory of Molecular Carcinogenesis, National Institute of Environmental Health Sciences, NIH, P.O. Box 12233, 111 T.W. Alexander Drive, Research Triangle Park, NC 27709, USA.
Molecular Cancer Therapeutics (Impact Factor: 6.11). 06/2005; 4(5):693-703. DOI: 10.1158/1535-7163.MCT-04-0337
Source: PubMed

ABSTRACT We previously showed that nonsteroidal anti-inflammatory drugs (NSAID) such as sulindac sulfide, which has chemopreventive activity, modulate the expression of several genes detected by microarray analysis. Activating transcription factor 3 (ATF3) was selected for further study because it is a transcription factor involved in cell proliferation, apoptosis, and invasion, and its expression is repressed in human colorectal tumors as compared with normal adjacent tissue. In this report, we show that ATF3 mRNA and protein expression are up-regulated in HCT-116 human colorectal cancer cells following treatment with NSAIDs, troglitazone, diallyl disulfide, and resveratrol. To ascertain the biological significance of ATF3, we overexpressed full-length ATF3 protein in the sense and antisense orientations. Overexpression of ATF3 in the sense orientation decreased focus formation in vitro and reduced the size of mouse tumor xenografts by 54% in vivo. Conversely, overexpression of antisense ATF3 was protumorigenic in vitro, however, not in vivo. ATF3 in the sense orientation did not modulate apoptosis, indicating another mechanism is involved. With microarray analysis, several genes relating to invasion and metastasis were identified by ATF3 overexpression and were confirmed by real-time reverse transcription-PCR, and several of these genes were modulated by sulindac sulfide, which inhibited invasion in these cells. Furthermore, overexpression of ATF3 inhibited invasion to a similar degree as sulindac sulfide treatment, whereas antisense ATF3 increased invasion. In conclusion, ATF3 represents a novel mechanism in which NSAIDs exert their anti-invasive activity, thereby linking ATF3 and its gene regulatory activity to the biological activity of these compounds.

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    • "The following protocol was used for each PCR: 94°C for 1 min (1 cycle), followed by 9°C for 1 min, 57°C for 1 min, and 72°C for 2 min (26 cycles), and a final extension phase at 72°C for 5 min. Primer sequences for human ATF-3 were 5′-CGGTTTCCGTCTGGGCTTCT-3′ (forward) and 5′-GCACCTCAAAGCTGTTCCGTCCC-3′ (reverse) [16]; for human p53 were 5′-CGGTTTCCGTCTGGGCTTCT-3′ (forward) and 5′-GCACCTCAAAGCTGTTCCGTCCC-3′ (reverse); for human GAPDH mRNA were 5′- TGAAGGTCGGTGTGAACGGATTTGG-3′ (forward) and 5′-ACGACATACTCAGCACCGGCCTCAC-3′ (reverse). PCR products were run on a 2% agarose gel containing 0.5 g/mL ethidium bromide. "
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    ABSTRACT: Background and Purpose Regulation of the homeostasis of vascular endothelium is critical for the processes of vascular remodeling and angiogenesis under physiological and pathological conditions. Urotensin II (U-II), a potent vasoactive peptide, participates in vascular and myocardial remodeling after injury. We investigated the protective effect of U-II on doxorubicin (DOX)-induced apoptosis in cultured human umbilical vein endothelial cells (HUVECs) and the potential mechanisms involved in this process. Experimental Approach Cultured HUVECs were treated with vehicle, DOX (1 µM), U-II, or U-II plus DOX. Apoptosis was evaluated by DNA strand break level with TdT-mediated dUTP nick-end labeling (TUNEL) staining. Western blot analysis was employed to determine the related protein expression and flow cytometry assay was used to determine the TUNEL positive cells. Key Results U-II reduced the quantity of cleaved caspase-3 and cytosol cytochrome c and increased Bcl-2 expression, which results in protecting HUVECs from DOX-induced apoptosis. U-II induced Activating transcription factor 3 (ATF3) at both mRNA and protein levels in U-II-treated cells. Knockdown of ATF3 with ATF3 siRNA significantly reduced ATF3 protein levels and U-II protective effect under DOX-treated condition. U-II downregulated p53 expression in DOX-induced HUVECs apoptosis, and it rapidly activated extracellular signal-regulated protein kinase (ERK) and Akt. The DOX induced change of p53 was not affected by U-II antagonist (urantide) under ATF-3 knockdown. The inhibitory effect of U-II on DOX-increased apoptosis was attenuated by inhibitors of ERK (U0126) and PI3K/Akt (LY294002). Conclusion and Implications Our observations provide evidence that U-II protects HUVECs from DOX-induced apoptosis. ERK-Akt phosphorylation, ATF3 activation, and p53 downregulation may play a signal-transduction role in this process.
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    • "In this context, we are aware of the complexity of sulindac actions and of the Wnt/β-catenin signaling pathway. For instance, the anti-invasive activity of sulindac was also regulated by ATF3 in colon cancer cells [47]. Therefore, we tested the effect of sulindac with respect to the inhibition of S100A4 expression and cell migration in additional colon cancer cell lines. "
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    • "Colon cancer cells Reduced tumorigenicity Bottone et al. (2005) Phenotypes indicate the consequences of ATF3 deficiency in the corresponding stress models or the consequences of ectopically expressing ATF3 (transgenic or injection models). a Gain-of-function approach. "
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