Abstract 4113: Pre-metastatic niche formation in palmitic acid and Caco2 BBE secreted exosomes treated HepG2 cells by the CD98-integrin-FAK pathway

Abstract

Abstract Up to 60% of patients with colorectal cancer (CRC) will develop metastatic CRC. This study will use an in-vitro Non-Alcoholic Fatty Liver Disease (NAFLD) model in HepG2 cells to determine how CD98 and colon cancer secreted exosomes initiate a pre-metastatic niche. CD98 is a glycoprotein that mediates integrin signaling, amino acid transportation, inflammatory cytokines, and possibly endocytosis of exosomes. The pathway of CD98-integrin-FAK may mediate uptake of exosomes as well as the formation of pre-metastatic niche in the liver. Exosomes secreted from colon cancer and NAFLD have been implicated to initiate a pre-metastatic niche in the liver. This combinatorial study will mimic in-vivo conditions of metastatic CRC. HepG2 cells are treated with 0 μM or 7 μM of palmitic acid (PA) for 5 days as well as stimulated with and without 10 μg/mL LPS to mimic chronic inflammation in-vitro. After the 5 days of PA and LPS, then the cells are treated for additional 2 days with Caco2BBE secreted exosomes. The treatments were assessed for NAFLD and inflammatory markers FAS, ABCA1, PPARα, PPARϒ, IL-1β, IL-6, TNF-α, and CD98 by Oil-Red staining, Western blot, RT-qPCR, and immunocytochemistry (IMC). The same set experiments with a knock down of CD98 and creation of pre-metastatic niche were examined through CD98-integrin-FAK pathway. This model showed that low doses of PA formed lipid vacuoles and increased NAFLD markers FAS, PPARα, PPARγ, and ABCA1. The cytotoxicity test of the 0-7 uM of PA with and without LPS as well as exosomes indicated low toxicity. IMC of HepG2 cells showed a unique co-localization patterns with no treatment and PA causing association CD98 with integrin β1 and CD98 with caveolin-1, while the LPS and knock down of CD98 disrupted these interaction and allowed an interaction between CD98 and actin. The addition of Caco2BBE secreted exosomes with PA and LPS altered the inflammatory markers and cellular location of CD98. In conclusion, low doses of palmitic acid produced lipid vacuoles, little cytotoxicity, and increased NAFLD markers. The loss of CD98 led to cellular re-location of CD98 to the nucleus and could explain the decrease uptake of Caco2 BBE secreted exosomes and decrease in inflammatory markers. These data suggest that a pre-existing condition such as NAFLD and Caco2 BBE secreted exosomes play an integral role in the development of pre-metastatic niche by the CD98-integrin-FAK pathway. The identifying the nuances of CD98-integrin-FAK pathway will elucidate future nano-particle therapeutics for metastatic CRC. Citation Format: Jenniffer L M Stetler, Brandon S. Canup, Hamed Laroui. Pre-metastatic niche formation in palmitic acid and Caco2 BBE secreted exosomes treated HepG2 cells by the CD98-integrin-FAK pathway. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 4113.