P1-02-01: c-Jun N-Terminal Kinase 1 (JNK1) Inhibits Tumor Growth and Metastasis by Downregulating Epithelial to Mesenchymal Transition (EMT) and Stem Cell-Related Genes.
Authors: Ebelt ND , Van Den Berg CL 
Published: June 2012

Abstract
Murine and human mammary cancers often show dysregulation of important signaling pathways including the canonical Wingless-iNTegrated (Wnt) and avian ERythroBlastosis oncogene B (ErbB) pathways. Transgenic expression of Wnt ligands causes transformation of normal mammary cells in mice, and Wnt10b is frequently upregulated in human breast cancers. Overexpression of ErbB ligands and amplification of receptors have also been implicated in human breast cancer. JNK1 is a tumor suppressor in the skin and intestinal epithelium, and JNKs are known to integrate ErbB and Wnt pathways as well as others to control cell growth and differentiation. In a murine mammary cancer model where 4T1.2 cells were injected into the mammary gland, reducing JNK1 levels by expressing shRNA (shJNK1) resulted in increased tumor growth and lung metastasis compared to mice injected with control vector- (pSM2) expressing cells. A microarray analysis comparing gene expression between shJNK1 and control tumors revealed 2 and 2.5-fold increases in the ErbB pathway genes Nrg3 and ETV5, 2-fold increases in the Wnt genes Bcl-9 and Wnt10a, and a 1.6-fold increase in the EMT gene Twist1. RT-PCR analysis of in vitro grown 4T1.2 cell lines transfected with shJNK1 confirmed increased expression of Bcl-9 and ETV5, as well as the ETV5 target Cox-2. ErbB2 protein was also overexpressed. The shJNK1 cells showed upregulation of pERK in response to Heregulin1 (an Erbb2/3 ligand) and Fibroblast Growth Factor (FGF) 1, which further amplifies canonical Wnt signaling. In a p53-/− tumor model, ETV5, Bcl-9, and Cox-2 were still upregulated in jnk1-/−compared to wildtype tumor cells, indicating this effect is p53-independent. In the normal mammary gland, a 4-fold increase in ETV5 and a 5-fold increase in Twist1 were found in jnk1-/− mice compared to wildtype, further indicating that this effect is dependent on JNK1 alone. The ErbB and Wnt pathways are known to upregulate EMT and stem-cell related genes, however, the involvement of JNK1 in these effects is a novel hypothesis. Thus far, our data suggest that JNK1 deficiency targets these oncogenic pathways to contribute to a more aggressive tumor phenotype due to heightened EMT and “stem-cellness”. Further studies using cell sorting and differentiation assays will determine whether normal jnk1-/− glands contain a higher fraction of stem cells than wildtype glands. Inhibition of EMT genes and/or ETV5 in the p53-/−;jnk1-/− cancer cells will determine whether the tumor growth or metastasis phenotypes are dependent on these genes. EMT and stem-cell genes are frequently expressed in human breast cancer subtypes that exhibit low survival rates, and EMT is known to be linked to increased metastasis. Some of these sub-types, such as claudin-low tumors, currently have no molecularly-targeted treatments, therefore it is important to determine what proteins critically contribute to these phenotypes so that efficient and effective treatments can be developed.
Citation Information: Cancer Res 2011;71(24 Suppl):Abstract nr P1-02-01.