Epidermal growth factor (EGF) receptor (EGFR) signalling is implicated in tumour invasion and metastasis. However, whether there are EGFR signalling pathways specifically used for tumour invasion still remains elusive. Overexpression of Arf6 and its effector, AMAP1, correlates with and is crucial for the invasive phenotypes of different breast cancer cells. Here we identify the mechanism by which Arf6 is activated to induce tumour invasion. We found that GEP100/BRAG2, a guanine nucleotide exchanging factor (GEF) for Arf6, is responsible for the invasive activity of MDA-MB-231 breast cancer cells, whereas the other ArfGEFs are not. GEP100, through its pleckstrin homology domain, bound directly to Tyr1068/1086-phosphorylated EGFR to activate Arf6. Overexpression of GEP100, together with Arf6, caused non-invasive MCF7 cells to become invasive, which was dependent on EGF stimulation. Moreover, GEP100 knockdown blocked tumour metastasis. GEP100 was expressed in 70% of primary breast ductal carcinomas, and was preferentially co-expressed with EGFR in the malignant cases. Our results indicate that GEP100 links EGFR signalling to Arf6 activation to induce invasive activities of some breast cancer cells, and hence may contribute to their metastasis and malignancy.

Objective: The biological activities of interleukin-17 (IL-17), a newly cloned cytokine, have not been fully elucidated. The present study was designed to assess the in vitro and in vivo effect of transfecting the IL-17 gene into tumor cells. Methods: A complementary DNA (cDNA) encoding human IL-17 (hIL-17) was obtained by polymerase chain reaction amplification from the human CD4+ T cell cDNA library and inserted into the plasmid pRc/cytomegalovirus to construct an expression vector for the hIL-17 gene. Murine Meth-A fibrosarcoma cells were transfected with the hIL-17 gene using the lipofectin method. The hIL-17 gene-expressing clone (Meth-A/IL-17) was selected and analyzed for cytokine expression by Northern blot. Results: There was no significant difference in the in vitro proliferation rate among parent Meth-A, cells transfected with vector alone and Meth-A/IL-17 cells. When the tumor cells were transplanted subcutaneously into BALB/c nude (nu+/nu+) mice, there was no difference in in vivo growth rates among the three cell lines. Challenge with tumor cells in conventional BALB/c mice, however, resulted in the rejection of Meth-A/IL-17 cells, but the other two lines did grow. After immunization with Meth-A/IL-17 cells, the mice were rechallenged by parent Meth-A or syngeneic MOPC-104E plasmacytoma cells; the immunized mice rejected the Meth-A cells, but not the MOPC-104E cells. Injecting the anti-thy 1,2 (CD90), anti-CD4 or anti-CD8 monoclonal antibody into conventional BALB/c mice resulted in the resumption of in vivo growth of Meth-A/IL-17 cells, but injecting the anti-asialo GM1 antibody did not. Furthermore, flow cytometric analysis demonstrated a significant increase in the expression of major histocompatibility complex (MHC) class I and class II antigens and lymphocyte function-associated antigen-1 on Meth-A/IL-17 cells. Conclusion: Meth-A cells transfected with the hIL-17 gene can induce tumor-specific antitumor immunity by augmenting the expression of MHC class I and II antigens, and both CD4+ and CD8+ T cells may play important roles in inducing antitumor immunity, suggesting the possibility of developing a tumor vaccine incorporating IL-17-transfected tumor cells.

No abstract

No abstract