Epidermal growth factor receptor-targeted antibody therapy - Mechanisms of action and modulators of therapeutic efficacy

Abstract

Cancer is an increasing disease in the world population, and in recent years there has been substantial interest in the development of novel therapeutic agents specifically targeting growth factor receptors on tumor cells. The epidermal growth factor receptor (EGFR) represents a tyrosine kinase cell surface receptor involved in important signaling pathways. Aberrant EGFR activation drives tumorigenesis, and EGFR over-expression is observed in many human cancers. At present, EGFR inhibition represents a validated cancer therapeutic strategy. A number of EGFR inhibitors are currently approved for clinical use, and can be divided into two classes: tyrosine kinase inhibitors (TKI) and monoclonal antibodies (mAb). The aim of this thesis was to characterize the mechanisms of action of EGFR-targeted antibodies, and to identify patient, tumor and drug-related factors that modulate their therapeutic efficacy. Special focus was given to zalutumumab, a novel fully human EGFR-targeted antibody. EGFR mAb act by two mechanisms of action. First, blockade of EGFR signaling, which was most effective in vivo at complete receptor saturation, and required relatively high antibody doses. EGFR mAb compete with ligands for the ligand-binding site and prevent receptor activation by limiting EGFR molecular flexibility. At low receptor occupancy, anti-tumor effects were likely based on engagement of the immune effector mechanism antibody-dependent cell-mediated cytotoxicity (ADCC). Further, patient and tumor related characteristics were found to impact the therapeutic efficacy of EGFR mAb. EGFR expression in normal and tumor tissues was observed to accelerate EGFR mAb plasma clearance in cynomolgus monkeys at low doses but not at high doses, resulting in nonlinear pharmacokinetics. Consequently, antibody saturation of EGFR may be affected, thereby affecting the local mechanisms of action of therapeutic EGFR mAb. In parallel studies, we examined whether zalutumumab Fc-glycosylation impacted antibody effector functions. Glycosylation of antibody Fc is a highly consistent property of antibody derived from a single cell line. Variations in glycosylation by antibodies derived from distinct cell lines, however, significantly impact antibody activity. Fab-mediated effector functions (e.g. EGFR signaling blockade) were not affected by differences in antibody glycosylation, in contrast to Fc-mediated effector functions. Low fucosylated zalutumumab appeared more efficient in FcgammaRIII-mediated ADCC by mononuclear cells, than fully fucosylated zalutumumab. Surprisingly, however, granulocyte-mediated ADCC, was found to be triggered via FcgammaRIIa and was initiated more effectively with high fucosylated EGFR antibodies. Fc-fucosylation may, thus, prove critical for therapeutic antibodies predominantly recruiting MNC for ADCC, while fucosylation levels are likely less relevant when PMN play a significant role in therapeutic effects initiated by EGFR mAb. Finally, the impact of tumor-derived EGFR mutations (L858R, G719S, delE746-A750, T790M) on the anti-tumor activity of EGFR antibodies was investigated. These mutations were correlated with clinical responses to TKI therapy, and described to alter EGFR kinase activity. In vitro analyses demonstrated mAb mediated anti-tumor effects not to be affected by any of these mutations. Somatic tumor-associated EGFR mutations are, therefore, not expected to impact the activity of therapeutic antibodies. In conclusion, multiple elements may modulate therapeutic efficacy during cancer treatment. Prognostic assays to stratify patients responsive to EGFR therapy will facilitate the future clinical development of such agents. A thorough understanding of the various factors modulating the efficacy of antibody therapeutics will contribute to further improvements in the prospects for cancer patients.