Abstract LB-229: PTEN modulates EGFRvIII-mediated SHP-2 activation and translocation to affect glioblastoma sensitivity to tyrosine kinase inhibitors (TKI)

Abstract

Abstract EGFRvIII, a constitutively active truncated mutant of epidermal growth factor receptor (EGFR), has been associated with increase neoplastic transformation and poor overall survival (OS). An earlier report showed that expression of EGFRvIII in gliomas provides resistance to chemotherapy. Co-expression of EGFRvIII and PTEN in a small subset of recurrent GBMs has been shown to increase sensitivity to tyrosine kinase inhibitors (TKI), including Erlotinib (tarceva) and Geftinib. However, the exact mechanism of EGFRvIII and PTEN interaction in response to TKI is still unresolved. Our recent reports showed that SHP-2 PTPase and its phosphatase activity are required for the EGFRvIII-mediated transformation. The aim of the present work was to investigate the molecular interaction between EGFRvIII/SHP-2 activation complex and PTEN in response to tarceva treatment and its effect on glioblastoma transformation. Our recent report demonstrated an increased SHP-2 phosphorylation in U87MG. EGFRvIII (PTEN-deficient) when compared to PTEN-intact LN229.EGFRvIII cells. In the present study, we show that tarceva treatment abolished EGFRvIII, EGFR, SHP-2 and Erk1/2, but Src family kinase (SFK), phosphorylation in LN229.EGFRvIII cells at all time intervals ranging from 30 minutes to 6 hours. On the contrary, phosphorylation of EGFRvIII, Src and Erk1/2 in U87MG. EGFRvIII cells was inhibited in early time points, but was restored in 2 to 6 hours. Surprisingly, phosphorylation of SHP-2 at Tyr542 residue was unaffected by tarceva treatment, but EGFR phosphorylation was inhibited in a time-dependent manner. PTEN deficiency resulted in resistance to tarceva treatment as assessed by MTT proliferation and soft agar transformation assays. Immunofluorescent labeling of U87MG. EGFRvIII cells with anti-phospho-SHP-2 (Tyr542) antibody showed perinuclear localization of SHP-2, whereas LN229.EGFRvIII cells exhibited membrane staining of phosphorylated SHP-2 (Tyr542). Our results showed that tarceva increased perinuclear SHP-2 Tyr542 in U87MG. EGFRvIII cells. Interestingly, stable expression of PTEN in U87MG. EGFRvIII cells conferred relocalization of phospho-SHP-2 to the membrane, a phenotype resembling LN229.EGFRvIII cells. Moreover, PTEN-expressing U87MG. EGFRvIII clones grew slower and showed a partially untransformed phenotype. These results suggest that SHP-2 is downstream effectors of PTEN and that PTEN deficiency may lead to SHP-2 activation and perinuclear localization by EGFRvIII, which may results in increased resistance to TKIs. Therefore, targeting SHP-2 may provide a new and effective therapeutic approach for treatment of glioblastoma. 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 LB-229.