Abstract 5126: Actopaxin: A novel regulator of cell migration and invasion in human hepatocellular carcinoma (2010)

Abstract Background: The mortality rate of hepatocellular carcinoma (HCC) is high due to tumor recurrence and development of metastasis. The elementary steps leading to the development of metastasis is a complex process involving cell spreading, lamellipodia formation and cell migration. Formation of focal adhesions is a key regulator of cell motility and cell invasion. Actopaxin, a focal adhesion and cytoskeleton-associated protein, is required for such processes. The aim of this study was to examine the role of Actopaxin in HCC cell migration, invasion and development of metastasis. Methods: The expression of Actopaxin in primary and metastatic liver cancer cell-lines was examined by western blot and RT-PCR. The functional and molecular effects of Actopaxin expression were studied by enforced expression or down-regulation in HCC cell-lines. Immunofluorescence staining for actin cytoskeleton and focal adhesion was performed to examine the effect of Actopaxin expression on cell shape, stress fiber organization, and focal adhesion. The clinicopathological significance of Actopaxin expression in tumorous tissues of 39 HCC patients was also investigated. Results: High protein level of Actopaxin was found in the metastatic HCC cell lines H2M, MHCC-97L and MHCC-97H as compared with other non metastatic HCC cell lines. Expression of a shorter form of Actopaxin was also detected in some of these cell lines, and subsequent RT-PCR and DNA sequencing identified two forms of Actopaxin coding sequences. The longer form (LF-Actopaxin) is identical to that published in NCBI Genbank database (Accession # NP_060692), while the unreported short form (SF-Actopaxin) lacks a fragment in the C-terminal, resulting in an incomplete 2nd CH domain which consists binding sites for its downstream activation targets. Enforced expression of LF-Actopaxin in primary cell line PLC readily enhanced cell migration in wound healing assays, as well as enhanced stress fiber formation, filopodia and lamellipodia (cell protrusions) on the cell surfaces. Such effects were not observed in SF-Actopaxin overexpressed PLC cells. Ecotpic expression of LF-Actopaxin, but not its corresponding short form resulted in elevated protein levels of ILK, PINCH and phospho-paxillin, which are involved in focal adhesion complex formation and integrin signaling pathway. Study in clinical samples showed that LF-actopaxin expression was positively correlated with tumor size (p&amp;lt;0.05) while SF-actopaxin expression was lower in HCC tumor compared with corresponding nontumorous liver (p&amp;lt;0.001) and lower in HCC with metastasis compared with HCC without metastasis (p&amp;lt;0.05). Conclusion: This study demonstrated for the first time the pro-migratory effects of Actopaxin in human HCC, and the existence of a short form which lacks a complete CH domain that is critical for cell migration, re-organization of cytoskeletal events and turnover of focal adhesions. 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 5126.