Epidermal growth factor receptor trafficking in squamous cell carcinoma cell lines

Abstract

The cancer therapy cetuximab targets epidermal growth factor receptor (EGFR) in head and neck squamous cell carcinoma (HNSCC), but 80% of patients don’t respond. EGFR can internalise into cells through at least two modes of endocytosis involving dynamin: clathrin-mediated endocytosis (CME) or fast endophilin-mediated endocytosis (FEME) which is clathrin-independent and endocytoses receptors at a much faster rate. A combination therapy of cetuximab with dynamin inhibition to block EFGR endocytosis increases cancer cell death in cell lines and mouse models, but which mode is responsible for this? Characterisation of five SCC cell lines showed that EGFR localisation in two of them, cetuximab-resistant KJD cells and cetuximab-sensitive A431 cells, mimicked the phenotypes of cetuximab-resistant and -sensitive tumour cells from patients. These two lines were selected for focussed studies. Higher amounts of dynamin and endophilin and a faster initial rate of EGFR uptake in KJD cells suggested hallmarks of EGFR trafficking through FEME. EGFR also colocalised with endophilin and early endosomes, and endophilin depletion reduced EGFR internalisation. However, some EGFR was also found to colocalise with clathrin, where clathrin and dynamin inhibition reduced EGFR uptake in KJD cells. These findings proposed that more than one mode of endocytosis may be trafficking EGFR and in turn, be implicated in cetuximab resistance. Comparison of the proteomes of SCC cells revealed decreased processes and pathways involving clathrin in KJD cells. This thesis has demonstrated that EGFR trafficking differs between a cetuximab-sensitive and -resistant SCC cell line where both CME and FEME are likely to be involved in cetuximab resistance. Enriching our knowledge of endocytosis modes and establishing those mediating EGFR function will help to identify specific proteins to target in the future for combination therapeutics with cetuximab in the hope of improving patient outcomes.