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Molecular and Cellular Biology, April 2009, p. 2243-2253, Vol. 29, No. 8
0270-7306/09/$08.00+0 doi:10.1128/MCB.00959-08
Copyright © 2009, American Society for Microbiology. All Rights Reserved.
Small-Molecule Activation of p53 Blocks Hypoxia-Inducible Factor 1
and Vascular Endothelial Growth Factor Expression In Vivo and Leads to Tumor Cell Apoptosis in Normoxia and Hypoxia
Jun Yang,1,
Afshan Ahmed,2,
Evon Poon,2
Nina Perusinghe,1
Alexis de Haven Brandon,3
Gary Box,3
Melanie Valenti,3
Suzanne Eccles,3
Kasper Rouschop,4
Brad Wouters,5 and
Margaret Ashcroft1,2*
Cell Growth Regulation and Angiogenesis Team, Cancer Research UK Centre for Cancer Therapeutics, the Institute of Cancer Research, 15 Cotswold Rd., SaHon, Surrey SM2 5NG, United Kingdom,1 Hypoxia Signalling and Angiogenesis Laboratory, Centre for Cell Signalling and Molecular Genetics, Department of Metabolism and Experimental Therapeutics, Division of Medicine, Univerisity College London, Rayne Building, 5 University Street, London WC1E 6JJ, United Kingdom,2 Tumor Biology and Metastasis Team, Cancer Research UK Centre for Cancer Therapeutics, the Institute of Cancer Research, 15 Cotswold Rd., Sutton, Surrey SM2 5NG, United Kingdom,3 Maastricht Radiation Oncology (Maastro), University of Maastricht, 6200 Maastricht, The Netherlands,4 Ontario Cancer Institute, Princess Margaret Hospital, Toronto, Ontario, Canada M5G OA35
Received 17 June 2008/ Returned for modification 3 September 2008/ Accepted 30 January 2009
The p53 tumor suppressor protein negatively regulates hypoxia-inducible factor 1
(HIF-1). Here, we show that induction of p53 by the small-molecule RITA (reactivation of p53 and induction of tumor cell apoptosis) [2,5-bis(5-hydroxymethyl-2-thienyl) furan] (NSC-652287) inhibits HIF-1 and vascular endothelial growth factor expression in vivo and induces significant tumor cell apoptosis in normoxia and hypoxia in p53-positive cells. RITA has been proposed to stabilize p53 by inhibiting the p53-HDM2 interaction. However, induction of p53 alone was insufficient to block HIF-1 induced in hypoxia and has previously been shown to require additional stimuli, such as DNA damage. Here, we identify a new mechanism of action for RITA: RITA activates a DNA damage response, resulting in phosphorylation of p53 and 
H2AX in vivo. Unlike other DNA damage response-inducing agents, RITA treatment of cells induced a p53-dependent increase in phosphorylation of the subunit of eukaryotic initiation factor 2, requiring PKR-like endoplasmic reticulum kinase activity, and led to the subsequent downregulation of HIF-1 and p53 target proteins, including HDM2 and p21. Through the identification of a new mechanism of action for RITA, our study uncovers a novel link between the DNA damage response-p53 pathway and the protein translational machinery.
* Corresponding author. Mailing address: Centre for Cell Signalling and Molecular Genetics, Division of Medicine, University College London, Rayne Building, 5 University Street, London WC1E 6JJ, United Kingdom. Phone: 44 (0) 207 679 0799. Fax: 44 (0) 207 679 6211. E-mail: m.ashcroft{at}ucl.ac.uk
Published ahead of print on 17 February 2009.
These authors contributed equally to this work.
Molecular and Cellular Biology, April 2009, p. 2243-2253, Vol. 29, No. 8
0270-7306/09/$08.00+0 doi:10.1128/MCB.00959-08
Copyright © 2009, American Society for Microbiology. All Rights Reserved.
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