Abstract 3434: Lipopoylsaccharide promotes human esophageal cancer cell metastatic potential via the selectin/sialyl Lewis X axis

Abstract

Abstract Background: Emerging data suggest that post-operative infections may increase esophageal cancer recurrence, possibly accounting for the poor prognosis associated with this malignancy. We have previously shown that lipopolysaccharide (LPS), a gram-negative bacterial antigen, injected into mice increases in vivo cancer cell adhesion to the hepatic sinusoids. However, the influence of LPS directly on the cancer cell is unknown. We thus sought to elucidate the direct impact of LPS and its sole trans-membrane receptor, Toll-Like Receptor 4 (TLR4) on human esophageal cancer cell metastatic potential. Methods: C57BL/6 mice were prepared for hepatic intravital microscopy and received intra-arterial inoculations with highly metastatic human esophageal squamous cells (HKESC-2; 1.5×106 cells/100μl) pre-incubated in LPS (0.1 μg/ml x 48hrs) or control media. HKESC-2-sinusoidal endothelial cell interactions were quantified with direct in vivo visualization of the hepatic sinusoids. In vitro static adhesion assay was use to measure cancer cell adhesion to fibronectin, collagen I and IV. SB203580, a p38 antagonist, was use to block LPS signaling. Two important adhesion molecules we have previously shown to be involved in the adhesion of these cancer cells to hepatic sinusoids, selectins and sialyl Lewis X (sLeX), were blocked using murine i.v. injection of fucoidin and incubation of cancer cells with blocking antibodies respectively. Results are presented as mean +/− SEM and MWU-test determined significance (* p<0.05). Results: HKESC-2 cells express TLR4 by flow cytometry and RT-PCR respectively. Immunoblots of LPS incubated HKESC-2 cells revealed increased phosphorylation of p38 MAP Kinase, an intracellular TLR4 downstream signaling molecule. Thus treated cells had a 2-fold increase in vitro adhesion to fibronectin but not to Collagen 1 or 4, an effect that was reversed by inhibition of p38 phosphorylation. Murine hepatic intravital microscopy demonstrated that LPS treated esophageal cancer cells had a 2.3 fold increase in adhesion to sinusoidal endothelium as compared to control (cells/field of view: 33.2+/−3.6 vs. 14.2 +/−0.5)*. This LPS-facilitated in vivo adhesion was attenuated by blockade of selectins and sLeX (7.7 +/− 0.4*; 14.37 +/− 0.8*). Conclusion: Using a physiologically relevant in vivo model of the early steps of cancer metastasis we have demonstrated that LPS-TLR4 mediated inflammation increased the metastatic potential of human esophageal cancer cells, at least partially through increased selectin-sialyl Lewis X binding. These findings identify LPS-TLR4 binding as a potential therapeutic target for the treatment and prevention of metastasis. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3434.