Abstract 973: MEKK3 sustains EMT and stemness in pancreatic cancer by regulating YAP and TAZ transcriptional activity

Abstract

Abstract Background: Pancreatic cancer (PC) is one of the most threatening and poorly understood human malignancies. MEKK3 (MAP3K3) is an intracellular serine/threonine kinase activated by different signaling pathways, including IL1, TNFα and TLR8, which regulates in turn NF-κB, JNK and p38 activation.The transcriptional regulators Yes Associated Protein (YAP) and transcriptional coactivator with a PDZ binding domain (TAZ) are central determinants of malignancy and have been identified as a critical oncogenic effectors of KRAS in pancreatic cancer. We hypothesized that MEKK3 could sustain pancreatic cancer by inducing YAP/TAZ oncogenic activities. Methods: MEKK3 was knocked-out (KO) in AsPC1, Panc1, and MDA-Panc28 PC cell lines by CRISPR/Cas9. The expression of MEKK3, E-cadherin, YAP/TAZ and their target genes was measured by both Western blot and qPCR. Cell migration was assessed by transwell and wound healing assays. Stemness was measured by spheroid formation and percentage of CD44+/CD24+/EpCAM+ cancer stem cells (CSC). Effects of MEKK3 KO was evaluated in vivo in an orthotopic nude mouse model with GFP-expressing Panc1 PC cells. Expression of YAP/TAZ and their target genes in tumor samples was measured by IHC. Results: MEKK3 KO resulted in the reversion of both Epithelial-to-Mesenchymal Transition (EMT) and cell migration, as well as in the impairment of cell proliferation. More interestingly, the size of 3D colonies arising from MEKK3 KO cell lines was smaller than the ones formed by their respective controls, due to a reduced percentage of CD44+/CD24+/EpCAM+ CSC. In MEKK3 KO cells, the recruitment of YAP/TAZ onto the promoters of their target genes was significantly impaired and this paired with a reduction of the expression of the same genes. In nude mice, MEKK3 KO significantly reduced tumor growth and prolonged overall survival. Conclusions: Our study is the first to demonstrate that genetic silencing of MEKK3 is a valid approach to revert in vivo the aggressiveness of PC by modulating YAP and TAZ transcriptional activities. Inhibition of MEKK3 could represent an exciting therapeutic strategy for the treatment of PC, which should be warranted further development. Citation Format: Geny Piro, Raffaela Santoro, Marco Zanotto, Carmine Carbone, Giampaolo Tortora, Davide Melisi. MEKK3 sustains EMT and stemness in pancreatic cancer by regulating YAP and TAZ transcriptional activity [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 973.