Abstract 1582: Rho GDP dissociation inhibitor 2 (RhoGDI2) plays a functionally important role on EGFR signaling pathway in adenocarcinoma cells of lung cancer
Authors: Shin Jung-Young , Zhang Xiang-Hua , Yoon Seong-Ae , Chae Hiun Suk , Kang Jin-Hyoung 
Published: February 2011

It has been known that Rho GDP dissociation inhibitor 2 (RhoGDI2) is a regulator of Rho family GTPase. We identified a RhoGDI2 protein through the protein expression profile and then investigated its functional role on EGFR downstream signal in NSCLC (non-small cell lung cancer).

Three adenocarcinoma cell lines were selected according to the EGFR and K-RAS mutation status; PC9 (E476-A750 del in exon 19, wild type K-RAS), H1975 (T790M in exon 20 and L858R in exon 21, wild type K-RAS), A549 (wild type EGFR, K-RAS mutation in codon 12). Among 342 protein spots detected on the 2D gel, 134 spots were initially selected as differentially expressed proteins using proteome analysis. It was noteworthy that the ubiquitination-related proteins were highly expressed in EGFR TKIs-sensitive PC9 cells. On the other hand, the EGFR TKIs-resistant H1975 cells harboring both T790M and L858R mutations expressed the signal transduction-related proteins such as Zinc finger protein, G protein-coupled receptor and Ras oncogene family protein. We observed that p-EGFR (Y1047), p-AKT, and p-ERK were highly activated in PC9 cells in contrast to no or weak activation of these signaling proteins in A549 and H1975 cells.

In PC9 cell, the expression of RhoGDI2 was significantly reduced after gefitinib treatment. On the contrary, its expression was enhanced by gefitinib in H1975 cells. Accordingly, RhoGDI2 of the final six protein spots was selected for further study.

In six NSCLC cell lines, RhoGDI2 mRNA and protein expressions were evaluated using RT-PCR and Western blot analysis, respectively. The highest expression level was observed in PC-9 cells. When RhoGDI2 siRNA was transfected in PC9 cells, it was shown that RhoGDI2 protein expression was markedly reduced. Additionally, in this transfected PC-9 cells, no changes in the expression of p-ERK and truncated caspase-3 were detected, but total EGFR and p-AKT expression were increased. When gefitinib was added in the PC9 cells transfected with RhoGDI siRNA, the expression level of total EGFR was less decreased compared to that of PC-9 cells transfected with GAPDH (positive control). The RhoGDI2 siRNA transfected PC-9 cells treated with gefitinib showed more increased expression level of actin compared to that of positive control. We observed that the metastatic potential of the RhoGDI2 siRNA transfected PC-9 cells with lack of its expression was significantly increased in transwell migration assay. Taken together, it is suggested that RhoGDI2 protein is an important down-stream molecule of EGFR signaling pathway in PC9 cells harboring EGFR sensitive mutation, and plays a functional role in cell migration and 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 1582.