Abstract 1898: Potential molecular mechanisms of intrinsic resistance to EGFR-targeting monoclonal antibodies in HNSCC
Authors: Boeckx Carolien , Limame Ridha , Beeck Ken Op de , Wouters An , Deschoolmeester Vanessa , Specenier Pol , Vermorken Jan B. , Peeters Marc , Camp Guy Van , Baay Marc , Lardon Filip 
Published: August 2012

Background: Head and Neck Squamous Cell Carcinoma (HNSCC) comprises 90% of all head and neck cancers and is still a major cause of morbidity and mortality worldwide. The epidermal growth factor receptor (EGFR) is overexpressed in 80-95% of all HNSCC tumors. Since EGFR initiates important signal transduction pathways in HNSCC carcinogenesis, anti-EGFR therapeutics were developed in order to prevent its activation. However, many tumors remain non-responsive, since the single-agent response rate of the monoclonal antibody cetuximab is less than 15%. Therefore, biomarkers predicting sensitivity to EGFR-targeted therapy are needed. Methods: First, the IC50 (drug dose reducing cell growth to 50%) of cetuximab and panitumumab was determined on seven HNSCC cell lines by the sulforhodamine B assay. In addition, these cell lines were screened for mutations in K-Ras by High Resolution Melting Analysis, EGFRvIII mutation and HPV infection by PCR. Cell growth was monitored in real-time using an xCELLigence system (Roche Applied Science) in order to determine the optimal incubation period at which a clear effect of cetuximab treatment on cell growth could be detected. Next, “sensitive” and “resistant” cell lines were treated with 15nM cetuximab for the predefined incubation period and RNA was isolated using the Trizol method. A genome-wide gene expression analysis was performed by comparison of expression profiles of “sensitive” and “resistant” HNSCC cells. Results: Neither K-Ras nor EGFRvIII mutation and no HPV DNA could be detected. Based on the IC50 values for cetuximab and panitumumab, cell lines were either classified as “sensitive” (n=3), “resistant” (n=2) or “intermediate” (n=2). The IC50 values of the “sensitive” cell lines ranged between 0.05 - 0.43nM for cetuximab and 0.01 - 0.45nM for panitumumab. Discussion: The IC50 values of cetuximab and panitumumab were very similar, as expected since both drugs belong to the same family of EGFR-targeting therapeutics. The cell lines used in our study did not show K-Ras or EGFRvIII mutations and were HPV negative. The xCELLigence results indicate that cetuximab (15nM) starts showing a clear effect on cell growth after 13h, implying that the signalling pathways elicited by cetuximab have been activated at this time point. Preliminary results from microarray analysis indicate that intrinsic cetuximab-resistance in HNSCC can be mediated cell line-dependently by different signaling pathways. In addition, putative characteristics of epithelial-to-mesenchymal transition were seen in “resistant” compared to “sensitive” cell lines.

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 1898. doi:1538-7445.AM2012-1898