Abstract 4155: Interstitial fluid flow-induced hepatocellular carcinoma cell invasion requires MEK/ERK signaling

Abstract

Abstract Background: Worldwide hepatocellular carcinoma (HCC) is the third leading cancer-related cause of death. Within the United States, HCC maintains a rising incidence rate of 3% each year since 1992 with a 5 year survival rate of less than 15%. The American Cancer Society has estimated 33,190 new cases off HCC and 23,000 HCC related deaths in the year 2014 within the United States. Our work aims to determine the role of biomechanical forces such as interstitial fluid flow (IFF) in HCC cell invasion. We hypothesized that HCC cells utilize the hypothesized ‘autologous chemotaxis’ mechanism to invade via secretion of the chemokine CXCL12 and expression of its receptor CXCR4. Autologous chemotaxis occurs when a biologically significant chemokine gradient is established by IFF, driving downstream invasion. In an effort to elucidate signaling pathways involved in IFF-induced HCC invasion, we have identified a potential role for MEK/ERK signaling. Materials and Methods: HCC cells or primary rat hepatocytes (PRH) were seeded into in a 3-D matrix composed of Matrigel and type I rat tail collagen, then exposed to static or interstitial flow conditions for 24 hours. The functional role of CXCR4 was tested using the CXCR4-antagonist AMD3100. MEK and ERK were inhibited using U0126 (25 μM) and FR180294 (10 μM), respectively. Phospho-MEK/ERK was assessed using western blot. Results and Discussion: The 3D invasion assay demonstrated that the HCC cells, Huh7 and Hep3B, invaded significantly more in response to IFF (5.5 fold increase in invasion compared to their respective static condition), while PRHs showed no response. AMD3100 was used to block the CXCR4 receptor which resulted in reduced invasion in Huh7 and Hep3B cells. Furthermore when U0126 and FR180204 were incorporated into the 3D invasion assay a significant reduction in flow-induced invasion was observed in Huh7 cells. However, interstitial flow or CXCR4 inhibition did not affect pMEK/ERK, suggesting that MEK/ERK are not modulated directly by IFF and are not downstream of CXCR4. Conclusion: Our results demonstrate that interstitial fluid flow stimulates HCC cell invasion. CXCR4/CXCL12 signaling and MEK/ERK signaling both are required for invasion, but appear to operate via separate mechanisms. Further investigation into these two separate mechanisms would provide better insight into flow-induced HCC invasion. This information could potentially enhance current cancer therapies and provide patients with better treatment options. Citation Format: Arpit D. Shah, Michael Bouchard, Adrian C. Shieh. Interstitial fluid flow-induced hepatocellular carcinoma cell invasion requires MEK/ERK signaling. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 4155. doi:10.1158/1538-7445.AM2015-4155