Differential display of mRNA analysis identified Pdcd4 (Cmarik et al., PNAS 1999) as a novel suppressor of transformation. Antisense expression of the novel pdcd4 gene converts transformation resistant (P-) to sensitive (P+) cells and pdcd4 sense expression (Yang et al Oncogene 2001) converts P+ to P- cells, thus establishing a causal relationship to prevention of tumor promoter induced transformation. Furthermore pdcd4 expression suppresses tumor phenotype in transformed mouse JB6 cells (Yang et al Oncogene 2003). A surprising discovery is that Pdcd4 expression in human cancer cell lines is predictive for sensitivity to tamoxifen and geldanamycin. Moreover, expression of Pdcd4 actually confers sensitivity to these drugs (Jansen et al Molec Cancer Ther 2004). Examination of the possible inhibitory effect of Pdcd4 on molecular events known to be required for tumor promotion revealed that Pdcd4 over-expression inhibited the activation of transcription factor AP-1 but not of NFkappa B or of ornithine decarboxylase(Yang et al Oncogene 2001). The AP-1 inhibiting activity of Pdcd4 appears to be attributable to blocking the transactivation of cJun and cFos (Yang et al Oncogene 2003). Although expression of Pdcd4 protein blocks AP-1 activation, Pdcd4 does not interact directly with Jun or Fos proteins. Analysis of Pdcd4 binding partners by a yeast two-hybrid assay and co-immunoprecipitation identified the translation initiation factors RNA helicase eIF4A and scaffold eIF4G as major binding partners (Yang et al Molec Cell Biol 2003, MCB 2004). Binding of Pdcd4 to eIF4A is required for Pdcd4 to inhibit the RNA helicase activity of eIF4A, to inhibit translation initiation, and to inhibit the activation of AP-1 dependent transcription required for neoplastic transformation. Mutational analysis defines two helical MA-3 domains as required for binding to eIF4A and for inhibiting translation (Yang et al Molec Cell Biol 2004). Residues on eIF4A required for binding Pdcd4 have also been characterized (Zakowicz RNA 2005). In collaboration with the laboratory of Alexander Wlodawer, the crystal structure of the C-terminal MA3 domain has been solved (LaRonde-LeBlanc, Santhanam et al MCB 2007). Analysis of the crystal structure predicts a mechanism by which Pdcd4 acts by competing with eIF4A binding partner eIF4G to inhibit translation initiation, a prediction that has been experimentally confirmed. Pdcd4 expression is downregulated in a number of human cancers, is diagnostic for human colon cancer staging and prognostic for survival (Muduluru et al Cancer 2007). A gene therapy approach to intervention in mouse lung carcinogenesis has revealed that lung inhalation administration of Pdcd4 induces apoptosis and inhibits AP-1 in mouse lung (Hwang et al Gene Ther 2007 and Jin et al Mol Ca Ther 2006)as well as inhibiting xenograft tumor growth (Kim YK, Kwon JT, Choi JY, Jiang HL, Arote R, Jere D, Je YH, Cho MH, Cho CS, Cancer Gene Ther. 2010). Pdcd4 overexpression inhibits invasion by human cancer cells. The mechanism involves targeting expression of a kinase upstream of cJun N-terminal kinase to consequently inhibit AP-1 dependent transcription (Yang et al MCB 2006). Recent investigation of the mechanism by which tumor suppressor Pdcd4 is down regulated during carcinogenesis revealed tumor promoter induced destabilization of the protein by a mechanism involving signaling through Akt, S6kinase and MEK/ERK (Schmid, Jansen et al, Cancer Res 2008). In collaboration with Michele Pagano at NYU, Pdcd4 has emerged as a target for degradation by the ubiquitin ligase betaTRCP (Dorrello et al Science 2006). Moreover, in collaboration with Heike Allgayer (Mannheim) and Yong Li (Louisville) we found that Pdcd4 is a functionally significant target of microRNA miR-21 (Asangani et al Oncogene 2008, Lu et al Oncogene 2008). Current research is focused on identifying specific transformation relevant mRNAs whose translation is inhibited by Pdcd4 expression andmay mediate its tumor suppressor activity. A recent inquiry into the structural features that distinguish mRNAs that are sensitive to translation inhibition revealed the surprising finding that the 3'UTR and microRNA binding sites are important (Santhanam et al PLoS ONE 2009). Tools for drug discovery targeting translation initiation have been generated in collaboration with Tobias Schmid (Goethe Univ, Frankfurt), Bruce Shapiro, Nahum Sonenberg (McGill Univ) and Curt Henrich(Blees et al, J Biomol Screen 2010). A high throughput screen for stabilizers of the novel tumor suppressor Pdcd4 has yielded several hits (Blees et al J Biomol Screen 2010, Zhao et al J Nat Prod 2011, and Grkovic et al J Nat Prod 2011, Blees PLoS ONR 2012). Polysome profiling has identified mRNAs that shift into or out of the actively translating fraction when Pdcd4 expression is down- or up-regulated in human breast cancer cells (Yoshikawa et al submitted 2012). We have recently discovered a new (indirect) transcriptional target of Pdcd4, namely the enzyme lysyl oxidase (Santhanam et al Oncogene 2010). Lysyl oxidase is important in mediating hypoxic response and breast cancer cell invasion. Recent studies (Gaur et al Neuro-Oncology 2011) have extended the observation of Pdcd4 as a tumor suppressor to glioblastoma (GBM)cells. Pdcd4 is a major target of oncogenic micro-RNA miR21 in GBM cells which show almost undetectable Pdcd4. Stable expression of Pdcd4 in GBM cells suppresses xenograft growth. NFkB/p65 has emerged as a new protein-protein interaction target of Pdcd4 that is functionally significant in GBM progression (Hwang et al 2013, submitted).

Agency
National Institute of Health (NIH)
Institute
National Cancer Institute (NCI)
Type
Investigator-Initiated Intramural Research Projects (ZIA)
Project #
1ZIABC010026-18
Application #
8763054
Study Section
Project Start
Project End
Budget Start
Budget End
Support Year
18
Fiscal Year
2013
Total Cost
$383,850
Indirect Cost
Name
National Cancer Institute Division of Basic Sciences
Department
Type
DUNS #
City
State
Country
Zip Code
Hwang, Soon-Kyung; Baker, Alyson R; Young, Matthew R et al. (2014) Tumor suppressor PDCD4 inhibits NF-κB-dependent transcription in human glioblastoma cells by direct interaction with p65. Carcinogenesis 35:1469-80
Narasimhan, Madhusudhanan; Rathinam, Marylatha; Riar, Amanjot et al. (2013) Programmed cell death 4 (PDCD4): a novel player in ethanol-mediated suppression of protein translation in primary cortical neurons and developing cerebral cortex. Alcohol Clin Exp Res 37:96-109
Blees, Johanna S; Bokesch, Heidi R; Rübsamen, Daniela et al. (2012) Erioflorin stabilizes the tumor suppressor Pdcd4 by inhibiting its interaction with the E3-ligase β-TrCP1. PLoS One 7:e46567
Gaur, Arti B; Holbeck, Susan L; Colburn, Nancy H et al. (2011) Downregulation of Pdcd4 by mir-21 facilitates glioblastoma proliferation in vivo. Neuro Oncol 13:580-90
Grkovic, Tanja; Blees, Johanna S; Colburn, Nancy H et al. (2011) Cryptocaryols A-H, α-pyrone-containing 1,3-polyols from Cryptocarya sp. implicated in stabilizing the tumor suppressor Pdcd4. J Nat Prod 74:1015-20
Zhao, Li-Xing; Huang, Sheng-Xiong; Tang, Shu-Kun et al. (2011) Actinopolysporins A-C and tubercidin as a Pdcd4 stabilizer from the halophilic actinomycete Actinopolyspora erythraea YIM 90600. J Nat Prod 74:1990-5
Schmid, Tobias; Bajer, Magdalena M; Blees, Johanna S et al. (2011) Inflammation-induced loss of Pdcd4 is mediated by phosphorylation-dependent degradation. Carcinogenesis 32:1427-33
Young, Matthew R; Santhanam, Arti N; Yoshikawa, Noriko et al. (2010) Have tumor suppressor PDCD4 and its counteragent oncogenic miR-21 gone rogue? Mol Interv 10:76-9
Blees, Johanna S; Schmid, Tobias; Thomas, Cheryl L et al. (2010) Development of a high-throughput cell-based reporter assay to identify stabilizers of tumor suppressor Pdcd4. J Biomol Screen 15:21-9
Yasuda, Michiko; Schmid, Tobias; Rübsamen, Daniela et al. (2010) Downregulation of programmed cell death 4 by inflammatory conditions contributes to the generation of the tumor promoting microenvironment. Mol Carcinog 49:837-48

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