Abstract B8: Targeting CXCR4/CXCL12 axis by AMD3100 inhibits ECTC-associated metastasis in hepatocellular carcinoma.

Abstract Background: Hepatocellular carcinoma (HCC) is characterized by high incidence of blood-borne metastasis. We have recently found that endothelium-coated tumor cell cluster (ECTC), a distinct vascular morphology, is significantly correlated with micrometastatic emboli and might represent a novel metastatic pathway of HCC. The aim of this study is to uncover the mechanisms of ECTC formation and the therapeutic potential of ECTC inhibition. Methods: ECTC was evaluated by immunohistochemical staining (IHC) of CD34. CXCR4 and CXCL12 expression in tumor vessels were examined by immunofluorescence staining. Human umbilical venous endothelial cells (HUVEC) were treated with CXCL12 and cell branches were assessed. HUVEC migration towards CXCL12 was tested by transwell assay. Orthotopic HCC was induced by intrahepatic injection of Hepa1-6 in male C57/BL6 mice. To block CXCR4/CXCL12 interaction, AMD3100 was injected i.p. every other day one week after the implantation. Tumor volumes and intrahepatic metastases were evaluated at the 3rd week after the injection. Endothelial cell branching and ECTC formation were measured by IHC. Results: ECTC-associated endothelial cells specifically expressed CXCR4 while endothelial cells from non-ECTC samples did not. Furthermore, CXCL12 (the unique CXCR4 ligand) was more abundant in ECTC vessels in comparison to non-ECTC vessels, suggesting a possible involvement of CXCR4/CXCL12 axis in ECTC formation. In vitro experiments revealed that CXCL12 promoted the branching and chemotactic migration of HUVEC. Mouse orthotopic HCC model showed that the intrahepatic metastasis incidence was inhibited by the specific CXCR4 antagonist AMD3100 (25% vs. 100%), accompanied with smaller tumor volumes. Moreover, endothelial cell branching (34% reduction, P<0.05) and ECTC formation (62% reduction, P<0.01) were significantly reduced by AMD3100 administration. Conclusion: CXCR4/CXCL12 interaction may promote ECTC formation by facilitating endothelial cell branching and directional migration. Disrupting CXCR4-CXCL12 interaction with AMD3100 reduced tumor metastasis by decreasing ECTC formation. Therefore, remodeling tumor vascular morphology by targeting CXCR4/CXCL12 axis may serve as a novel therapy for HCC treatment. Citation Information: Mol Cancer Ther 2013;12(11 Suppl):B8. Citation Format: Jing Yan, Jing Liang, Jing Xu, Limin Zheng. Targeting CXCR4/CXCL12 axis by AMD3100 inhibits ECTC-associated metastasis in hepatocellular carcinoma. [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2013 Oct 19-23; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2013;12(11 Suppl):Abstract nr B8.