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EGFR Glycosylation in Cancer
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Cancer is a leading cause of mortality around the world. Head and neck (HNC) and esophageal cancers (EC) are the sixth and eighth most common cancers worldwide, respectively. A common feature in both diseases is the overexpression of epidermal growth factor receptor (EGFR).
EGFR a transmembrane glycoprotein and cell-surface receptor is mainly involved in regulation of cell proliferation, survival, differentiation and its deregulation is associated with cancer. EGFR is highly glycosylated, and it is known that aberrant glycosylation interferes with its functions.
Several EGFR-targeted therapies have been already developed, though showing some undesirable secondary effects, such as skin toxicity, as normal keratinocytes also express EGFR. Identification of a specific target for tumor cells would be essential to minimize those effects. Since EGFR is glycosylated and aberrant glycosylation is a common feature in cancer, identifying a distinct EGFR glycosylation pattern between cancer and normal cells is a promising approach to improve specificity of targeted therapies.
In this work, primary human keratinocytes, cell lines and patient-derived tissues representative of HNC and EC were used to implement an optimized strategy to obtain EGFR enriched protein fractions from which EGFR glycosylation pattern could be defined. This thesis reports the results attained for the optimization steps on the different methodologies explored for protein extraction both from cells and tissues, EGFR detection and quantification, EGFR enrichment approaches and glycoprofiling methods (lectin ELISA and LC-MS analysis). Although promising strategies are proposed and validated to extract EGFR from different biological sources in adequate conditions for further mass spectrometry (MS) analysis, some difficulties were found in glycans detection by MS. Further work needs to be done to surpass this method sensitivity problem, as EGFR amounts recovered from enrichment protocols are extremely low, adding to the fact that when dealing with patient-derived samples, specimens are limited in size and availability.
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Head and neck cancer Esophageal cancer Epidermal growth factor receptor (EGFR) N-Glycosylation Protein enrichment methodologies Mass spectrometry analysis