P120 Catenin Mediated Epithelial-To-Mesenchymal Plasticity Determines The Metastatic Potential Of Pancreatic Ductal Adenocarcinoma

Introduction: The majority of pancreatic ductal adenocarcinoma (PDAC) patients present with metastases and nearly all will succumb to disease within 6-12 months of clinical presentation (Hidalgo, 2010). Recently, p120catenin (p120) was identified as a “cancer candidate gene” in sleeping beauty-induced PDAC and p120 loss is associated with poor patient survival (Mann et al, 2012). P120 is critical in stabilizing E-cadherin (E-cad) at the adherens junctions. In this study, we wished to unravel new roles for p120 in the context of PDAC initiation and progression (epithelial-mesenchymal transition or EMT), as well as metastasis. Results: We have introduced a floxed p120 allele into the Pdx1cre;LSL-KrasG12D;R26YFP (KCY) PDAC mouse model. Mice with homozygous p120 deletion in the context of a LSL-KrasG12D allele (KCYp120fl/fl) are not viable. However, mice with heterozygous p120 loss (KCYp120fl/wt) are born according to Mendelian ratios. At 20 weeks of age, KCYp120fl/wt mice show a remarkable acceleration of the Kras-driven phenotype. KCYp120fl/wt mice (n=21) harbor the entire spectrum of precursor lesions, including PanINs (1-3), mucinous cystic neoplasms (MCN) and intraductal papillary mucinous neoplasms (IPMN) and metastatic PDAC. The phenotype in control KCYp120wt/wt mice (n=12) was restricted to early PanINs. We next asked whether the second allele of p120 is lost during metastatic dissemination. We demonstrated that the remaining p120 allele is expressed in liver metastases, indicating that p120 loss-of-heterozygosity did not occur. Of note, in liver metastases, p120 co-localizes with E-cad, thereby indicating that a single p120 allele is sufficient to stabilize E-cad. This raised the question of whether one p120 allele is required to establish epithelial integrity at the metastatic site. In order to address this mechanistically, and since KCYp120fl/fl are not viable, we performed three-dimensional (3D) culture experiments with pancreatic cells isolated from non-recombined LSL-KrasG12D/+;p120wt/wt;R26YFP (KYp120wt/wt), KYp120fl/wt and KYp120fl/fl mice. Cells were Cre-recombined in vitro. In 3D culture, KYp120wt/wt cells form round multicellular cysts. Monoallelic p120 loss only disrupts the symmetry of cysts while biallelic loss completely prevents cells from forming organized, cyctic structures. We next injected these cell lines orthotopically into the pancreata of immunodeficient mice. KYp120wt/wt cells form PanIN-like structures. Interestingly, KYp120fl/wt establish large cysts reminiscent of IPMN/MCN lesions. In the cysts, p120 is localized at the plasma membrane with E-cad. However, the invasive fronts of tumors show significantly less p120 expression. Finally, KYp120fl/fl cells form poorly differentiated tumors invading into the surrounding stroma. Given the fact that p120 stabilizes E-cad and that E-cad is critical in EMT and MET, we injected the KYp120wt/wt, KYp120fl/wt and KYp120fl/fl cell lines directly into the portal veins of these mice. KYp120fl/wt cells colonize the liver and retain the remaining p120 allele and display membranous E-cad localization. By contrast, although KYp120fl/fl cells are able to generate invasive primary pancreatic tumors when being injected orthotopically, these cells fail to colonize the liver. Conclusions: Our findings demonstrate that monoallelic loss of p120 accelerates Kras-driven PDAC formation. However, one allele of p120 is required to establish epithelial integrity and metastases. Taken together, these results for the first time underscore the importance of p120-mediated EMT plasticity in order to complete the metastatic cascade in vivo. Furthermore, therapeutic strategies to prevent EMT alone may not be sufficient in light of our results but rather require novel approaches to target EMT plasticity so as to enhance survival in metastatic PDAC. This abstract is also presented as Poster A63. Citation Format: Maximilian Reichert, Basil S. Bakir, Christopher M. Hahn, Gregory P. Botta, Andrew D. Rhim, Robert H. Vonderheide, Albert B. Reynolds, Yingtao Bi, Ramana Davuluri, Burcu Saka, N. Volkan Adsay, Anil K. Rustgi. p120 catenin mediated epithelial-to-mesenchymal plasticity determines the metastatic potential of pancreatic ductal adenocarcinoma. [abstract]. In: Proceedings of the AACR Special Conference on Pancreatic Cancer: Innovations in Research and Treatment; May 18-21, 2014; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2015;75(13 Suppl):Abstract nr PR05.