Abstract 4124: Activation of AMP-activated protein kinase as a novel mechanism of resistance to EGFR-targeted therapy

Abstract

Abstract Epidermal growth factor receptor (EGFR)-mediated cell signaling is aberrantly regulated in many types of human cancers. Targeting EGFR with receptor-blocking antibodies or small molecule tyrosine kinase inhibitors has shown clinical activity against several types of cancers; however, most patients have had only moderate or no response. Our laboratory recently found that one of the antitumor mechanisms of cetuximab, which is an FDA-approved EGFR-blocking antibody, is to disrupt the biosynthetic and biogenetic metabolisms in cancer cells through inhibition of hypoxia-inducible factor 1 alpha (HIF-1α)-regulated glycolysis. In current study, we tested a novel hypothesis that AMP-activated protein kinase (AMPK) functions as a metabolic checkpoint activated in response to the decline in ATP levels by EGFR-targeted therapy. We selected three human head and neck squamous cell carcinoma cell lines, HN5 and FaDu (cetuximab-sensitive) and UMSCC1 (cetuximab-insensitive at low range <20nM) in this study. We found cetuximab significantly reduced cellular ATP levels and glucose consumptions in cultured cells through inhibition of HIF-1α activity in the cetuximab-sensitive HN5 and FaDu cells, but moderately reduced cellular ATP level and glucose consumption in UMSCC1 cells. By measuring the level of T-172 activation-specific phosphorylation of AMPK and the level of phosphorylation of acetyl CoA carboxylase (ACC) at S-79 (the site known to be phosphorylated by AMPK), we found that AMPK was strongly activated after cetuximab treatment in HN5 and FaDu cells, compared to untreated cells, but AMPK was only moderately elevated in UMSCC1 cells that contain a higher basal level of activated AMPK. Knockdown of LKB1 expression by specific siRNA prevented cetuximab-induced activation of AMPK, indicating that cetuximab activates AMPK through the canonical ATP-LKB1-AMPK signaling pathway. Inhibition of AMPK by AMPK siRNA or an AMPK small molecule inhibitor compound C (C.C.) markedly potentiated cetuximab-induced inhibition of cellular ATP level and cell proliferation in HN5 and UMSCC1 cells. Furthermore, we found that inhibition of AMPK by C.C. markedly improved response of HN5-R and DiFi5 cells, both of which are cetuximab-resistant sublines derived from their parental cetuximab-sensitive cells. These findings indicate that activation of AMPK is an important mechanism of resistance to cetuximab, attempting to restore cellular energy level after cetuximab treatment. Taken together, our findings suggest a novel mechanism in which AMPK is activated in response to the decline in ATP level by cetuximab-induced downregulation of HIF-1α. Novel strategies to block this AMPK-mediated cell energy restoring mechanism may sensitize cancer cells to cetuximab-mediated EGFR inhibition. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 4124. doi:1538-7445.AM2012-4124