Functions Of SOS1/EPS8/ABI1 In Metastasis-initiating Of Ovarian Cancer And Anti-metastatic Effects Of Its Short Peptide Inhibitor

Objective Epithelial-mesenehymal-transition (EMT) means the process that epithelial cell transforms to mesenchymal cell in morphology and acquires its metastasis capability, which is related to the metastasis of a variety of malignant tumors. As complex SOS1/EPS8/ABI1 plays a key role in metastasis capacity of ovarian cancer cells, it can be predicted that this complex may has a key function in the EMT of ovarian cancer cells. This study is to explore the functions of complex SOS1/EPS8/ABI1 in EMT of ovarian cancer and its start of metastasis, identify the sites of interactions among SOS1, EPS8 and ABI1 in the complex, design and synthesize targeted short peptide inhibitor of complex SOS1/EPS8/ABI1, and hope to provide new ideas for the pharmacotherapy of ovarian cancer.Methods Transwell migration and Western-Blot assays were performed to determine the correlations among the metastatic potentials of ovarian cancer cells, EMT and EMT related markers E-cadherin and vimentin. We then characterized the function of complex SOS1/EPS8/ABI1 in ovarian cancer cell migration and its EMT with genetic mutants and shRNAs. Finally, we identified the sites of interactions among SOS1, EPS8 and ABI1 in the complex, confirmed the effectiveness of short peptide inhibitor of complex SOS1/EPS8/ABI1 by Co-immunoprecipitation and GST-fusion protein pull-down assays.Results Invasive ovarian cancer cells displayed obvious LPA-stimulated cell migration (P<0.01), in the process of migration, there were Mesenehymal alteration, low expression of E-cadherin and high expression of vimentin. Non-invasive ovarian cancer cells displayed scarce LPA-stimulated cell migration (P>0.05), and there were high expression of E-cadherin, low expression of vimentin and no Mesenehymal alteration in the process of migration. Silencing any member of the complex SOS1/EPS8/ABI1 could block the metastatic potentials of invasive ovarian cancer cells (P>0.05), and there were high expression of E-cadherin, low expression of vimentin and no Mesenehymal alteration. Transferring the deletion genes could confer the corresponding non-invasive ovarian cancer cells with metastatic potentials (P<0.01). In the process of migration, there were Mesenehymal alteration, low expression of E-cadherin and high expression of vimentin. There was formation of complex SOS1/EPS8/ABI1 in LPA-stimulated ovarian cancer cell migration. SH3 domain of ABI1 could mediate its interaction towards SOS1, and poly-proline+PxxDY or PxxDY segment could mediate its interaction towards EPS8. At the same time, proline-rich segment of SOS1 could mediate its interaction towards ABI1 SH3 domain of EPS8 could mediate its interaction towards ABI1. Short peptide inhibitor p+p-8(pppppvdyedeeaavvqynd) could suppress the combination of ABI1 and EPS8, and short peptide inhibitor SH3-2 (fddfppppppppvdyedeeaavvqyndp) could suppress the combination of ABI1 and EPS 8. To in vitro cultured ovarian cancer cells, short peptide inhibitor TAT-p+p-8 could effectively suppress the combination of ABIl and EPS8, and gained the optimal effect within 24 hours. The inhibition of TAT-SH3-2 to ABI1and SOS1 is not significant.Conclusions EMT happened in the invasion process of ovarian cancer cell, and complex SOS1/EPS8/ABI1 takes E-cadherin as the target in the EMT of ovarian cancer cell. ABI1 combines to proline-rich domain of SOS 1 with its SH3 domain, and combines SH3 domain of EPS8 with its poly-proline+PxxDY segment. The synthesized short peptide inhibitor TAT-p+p-8 of complex SOS1/EPS8/ABI1 could effectively inhibit the combination of ABI1 and EPS8.It may provide a new approach with good application prospect for clinical treatment of the ovarian cancer patients.