HMLEs (HMLE-SNAIL and Kras-HMLE, Kras-HMLE-SNAIL pairs) serve as excellent model system to interrogate the effect of SNAIL targeted agents that reverse epithelial-to-mesenchymal transition (EMT). We had earlier developed a SNAIL-p53 interaction inhibitor (GN-25) that was shown to suppress SNAIL function. In this report, using systems biology and pathway network analysis, we show that GN-25 could cause reversal of EMT leading to mesenchymal-to-epithelial transition (MET) in a well-recognized HMLE-SNAIL and Kras-HMLE-SNAIL models.
GN-25 induced MET was found to be consistent with growth inhibition, suppression of spheroid forming capacity and induction of apoptosis. Pathway network analysis of mRNA expression using microarrays from GN-25 treated Kras-HMLE-SNAIL cells showed an orchestrated global re-organization of EMT network genes. The expression signatures were validated at the protein level (down-regulation of mesenchymal markers such as TWIST1 and TWIST2 that was concurrent with up-regulation of epithelial marker E-Cadherin), and RNAi studies validated SNAIL dependent mechanism of action of the drug. Most importantly, GN-25 modulated many major transcription factors (TFs) such as inhibition of oncogenic TFs Myc, TBX2, NR3C1 and led to enhancement in the expression of tumor suppressor TFs such as SMAD7, DD1T3, CEBPA, HOXA5, TFEB, IRF1, IRF7 and XBP1, resulting in MET as well as cell death.
Our systems and network investigations provide convincing pre-clinical evidence in support of the clinical application of GN-25 for the reversal of EMT and thereby reducing cancer cell aggressiveness.
Oncology | Pathology
Azmi et al.: Systems analysis reveals a transcriptional reversal of the mesenchymal phenotype induced by SNAIL-inhibitor GN-25. BMC Systems Biology 2013 7:85. References 1. 2. 3. 5. Klymkowsky MW, Savagner P: Epithelial-mesenchymal transition: a cancer researcher's conceptual friend and foe. Am J Pathol 2009, 174:1588â€“1593. Singh A, Settleman J: EMT, cancer stem cells and drug resistance: an emerging axis of evil in the war on cancer. Oncogene 2010, 29:4741â€“4751. Dave B, Mittal V, Tan NM, Chang JC: Epithelial-mesenchymal transition, cancer stem cells and treatment resistance. Breast Cancer Res 2012, 14:202. 4. Medici D, Hay ED, Olsen BR: Snail and Slug promote epithelial- 6. mesenchymal transition through beta-catenin-T-cell factor-4-dependent expression of transforming growth factor-beta3. Mol Biol Cell 2008, 19:4875â€“4887. Becker KF, Rosivatz E, Blechschmidt K, Kremmer E, Sarbia M, Hofler H: Analysis of the E-cadherin repressor Snail in primary human cancers. Cells Tissues Organs 2007, 185:204â€“212. Harney AS, Lee J, Manus LM, Wang P, Ballweg DM, LaBonne C, et al: Targeted inhibition of Snail family zinc finger transcription factors by oligonucleotide-Co(III) Schiff base conjugate. Proc Natl Acad Sci USA 2009, 106:13667â€“13672. Casas E, Kim J, Bendesky A, Ohno-Machado L, Wolfe CJ, Yang J: Snail2 is an essential mediator of Twist1-induced epithelial mesenchymal transition and metastasis. Cancer Res 2011, 71:245â€“254. Peinado H, Quintanilla M, Cano A: Transforming growth factor beta-1 induces snail transcription factor in epithelial cell lines: mechanisms for epithelial mesenchymal transitions. J Biol Chem 2003, 278:21113â€“21123. Baritaki S, Chapman A, Yeung K, Spandidos DA, Palladino M, Bonavida B: Inhibition of epithelial to mesenchymal transition in metastatic prostate cancer cells by the novel proteasome inhibitor, NPI-0052: pivotal roles of Snail repression and RKIP induction. Oncogene 2009, 28:3573â€“3585. 10. Mani SA, Guo W, Liao MJ, Eaton EN, Ayyanan A, Zhou AY, et al: The 9. 7. 8. 11. 12. 13. epithelial-mesenchymal transition generates cells with properties of stem cells. Cell 2008, 133:704â€“715. Lee SH, Lee SJ, Jung YS, Xu Y, Kang HS, Ha NC, et al: Blocking of p53-Snail binding, promoted by oncogenic K-Ras, recovers p53 expression and function. Neoplasia 2009, 11:22â€“31. Lee SH, Park BJ: p53 activation by blocking Snail: a novel pharmacological strategy for cancer. Curr Pharm Des 2011, 17:610â€“617. Lee SH, Shen GN, Jung YS, Lee SJ, Chung JY, Kim HS, et al: Antitumor effect of novel small chemical inhibitors of Snail-p53 binding in K-Ras-mutated cancer cells. Oncogene 2010, 29:4576â€“4587. 14. Rhodes DR, Yu J, Shanker K, Deshpande N, Varambally R, Ghosh D, et al: ONCOMINE: a cancer microarray database and integrated data-mining platform. Neoplasia 2004, 6:1â€“6. 15. Craene BD, Berx G: Regulatory networks defining EMT during cancer 16. initiation and progression. Nat Rev Cancer 2012, 13:97â€“110. Sanchez-Tillo E, Liu Y, Cuatrecasas M, Fanlo L, Siles L, de BO, et al: EMT- activating transcription factors in cancer: beyond EMT and tumor invasiveness. Cell Mol Life Sci 2012, 69:3429â€“3456. 17. Venkov C, Plieth D, Ni T, Karmaker A, Bian A, George AL Jr, et al: Transcriptional networks in epithelial-mesenchymal transition. PLoS One 2011, 6:e25354. 18. Morel AP, Hinkal GW, Thomas C, Fauvet F, Courtois-Cox S, Wierinckx A, et al: EMT inducers catalyze malignant transformation of mammary epithelial cells and drive tumorigenesis towards claudin-low tumors in transgenic mice. PLoS Genet 2012, 8:e1002723. 19. Bao B, Ali S, Banerjee S, Wang Z, Logna F, Azmi AS, et al: Curcumin analogue CDF inhibits pancreatic tumor growth by switching on suppressor microRNAs and attenuating EZH2 expression. Cancer Res 2012, 72:335â€“345. 20. Azmi AS, Philip PA, Beck FW, Wang Z, Banerjee S, Wang S, et al: MI-219-zinc combination: a new paradigm in MDM2 inhibitor-based therapy. Oncogene 2011, 30:117â€“126. 21. Azmi AS, Aboukameel A, Bao B, Sarkar FH, Philip PA, Kauffman M, et al: Selective inhibitors of nuclear export block pancreatic cancer cell proliferation and reduce tumor growth in mice. Gastroenterology 2013, 144:447â€“456.