Abstract 3633: eEF2 in acquired lapatinib resistance in HER2 positive breast cancer cells

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Abstract Approximately 25% of breast cancers overexpress HER2. Although HER2 targeted therapies, trastuzumab and more recently lapatinib, have improved prognosis for HER2 breast cancer patients not all HER2 positive tumors respond to HER2 targeted therapies. Using phosphoproteomic profiling, we examined mechanisms of resistance in an in vitro model of acquired lapatinib resistance. SKBR3 cells were conditioned in lapatinib containing media (200 nM) for six months, after which time the resulting SKBR3-L cells exhibited significantly reduced growth inhibition (27.7 ± 0.1 %) when treated with lapatinib (1 µM) treatment compared to parental SKBR3 (90.7 ± 0.2 %) (p<0.01). Using phosphoproteomic profiling, eukaryotic elongation factor 2 (eEF2) was identified as a phosphoprotein which was significantly altered in the SKBR3-L cells compared to the parental SKBR3 cells. One form of phospho-eEF2 was up-regulated and 6 forms were down-regulated in SKBR3-L compared to SKBR3. eEF2 is a phosphoprotein with an essential role in protein synthesis. Phosphorylation of eEF2 at Thr56 and Thr58 by eEF2 kinase inhibits eEF2 activity. eEF2 kinase activity is regulated by the mTOR pathway, via phosphorylation by p70S6 kinase. To examine the role of eEF2 in lapatinib resistance, levels of phospho-eEF2, eEF2, phospho-p70S6K and p70S6K, were examined in a panel of HER2 positive breast cancer cell lines following 24 hour treatment with lapatinib (1 µM). Treatment with lapatinib resulted in a decrease in phospho-p70S6K and an increase in phospho-eEF2, with no change in the levels of total protein, in lapatinib sensitive cell lines. In order to further examine the role the mTOR pathway and eEF2, plays in acquired resistance to lapatinib, the effect of rapamycin, alone and in combination with lapatinib, was investigated in SKBR3 par and SKBR3-L cells. Combined treatment with lapatinib (200 nM) and rapamycin (2 nM) results in similar inhibition of growth (64.5 ± 3.3 %) in SKBR3-L cells as lapatinib alone in SKBR3 parental cells (62.5 ± 0.6 %). In conclusion, response to lapatinib is associated with decreased eEF2 activity in HER2 positive cell lines. Furthermore, inhibition of mTOR signaling, which causes inhibition of eEF2 activity, appears to restore sensitivity to lapatinib in our model of acquired lapatinib resistance. 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 3633.