Sa1804 Dysregulation of WNT5A/ROR2 Signaling Pathway Characterizes the Progression of Barrett's-Associated Esophageal Adenocarcinoma

Abstract

BACKGROUND. Colorectal cancers (CRC) account for nearly 10% of cancer deaths in industrialized countries. Recent evidence points to a central role for the nuclear receptor Liver Receptor Homolog-1 (LRH-1) in intestinal tumorigenesis. Interaction of LRH-1 with the Wnt/β-catenin pathway, highly active in a critical subpopulation of CRC cancer cells, underscores the importance of elucidating LRH-1's role in this disease. Reduction of LRH1 diminishes tumor burden in murine models of CRC. The central role of receptor LRH-1 in CRC pathogenesis and a wealth of structural data at atomic resolution make LRH-1 an attractive target. Methods: To evaluate the contributions of LRH-1 to CRC growth, we constructed stably transduced CRC cell lines expressing Tet-inducible shRNA directed against LRH-1. Cell growth of the LRH-1 knockdown lines was compared with growth of cells expressing a non-silencing control RNA. To explore alterations in gene expression due to LRH-1 suppression, we performed a microarray analysis of our knockdown cell lines. Additionally, we sought to develop and characterize specific, non-cytotoxic inhibitors of LRH-1 function for use as both potential therapeutic agents and more broadly as research tools to study organogenesis and tumorigenesis. We applied differential scanning fluorimetry (DSF), used to measure qualitative binding, to screen over 200 compounds for association with the LRH-1 ligand binding domain. Results: LRH-1mRNA knockdown results in impaired cell proliferation. Cluster analysis of microarray gene expression demonstrates significant alterations in signal transduction, bile acid and cholesterol metabolism, and control of apoptosis. Moreover, we discovered 9 unique compounds that interact with LRH-1, but not the closely related receptor SF-1. We have further demonstrated that four of these compounds have antiproliferative effects with IC50 values ranging from 8.6 to 30 μM. Conclusions: Silencing of LRH-1 expression in CRC cell lines leads to impaired cell growth, demonstrating that LRH-1 is a viable target in CRC. LRH-1 may exert its effects via multiple signaling networks, with inhibition leading to cell cycle arrest or apoptosis. General features for binding shared between all compounds include a linear array of hydrophobic groups with polar atoms likely allowing for registration within the hormone pocket. Interestingly, two compounds showing antiproliferative activity share a cholesterol motif which may be related to LRH-1's function in bile acid and cholesterol homeostasis. Support: K12 HD07222, F32 CA163092, T32 DK007762