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Research has been conducted on the human epidermal growth factor receptor. The crystal structure of the receptor's ectodomain bound to transforming growth factor alpha and the ability of the receptor to bind a family of variable ligands have been investigated and are discussed.

The development of HER2-directed monoclonal antibodies and tyrosine kinase inhibitors have provided benefits to cancer patients, as well as produced many insights into the biology of the ErbB receptor family. Current therapies based on ErbB family members have resulted in improved overall survival with associated improvements in quality of life for the cancer patients that respond to treatment. Compared to monotherapy using either two antibodies to block the HER2 receptor blockade or combinatorial approaches with HER2 antibodies and standard therapies has provided additional benefits. Despite the therapeutic success of existing HER2 therapies, personalising treatment and overcoming resistance to these therapies remains a significant challenge. The heterogeneous intra-tumoural HER2 expression and lack of fully predictive and prognostic biomarkers remain significant barriers to improving the use of HER2 antibodies. Imaging modalities using radiolabelled pertuzumab and trastuzumab allow quantitative assessment of intra-tumoural HER2 expression, HER2 antibody saturation and the success of different drug delivery systems to be assessed. Molecular imaging with HER2 antibodies has the potential to be a non-invasive, predictive and prognostic technique capable of influencing therapeutic decisions, predicting response and failure of treatments as well as providing insights into receptor recycling and signalling. Similarly, conjugating HER2 antibodies with novel toxic payloads or combining HER2 antibodies with cellular immunotherapy provide exciting new opportunities for the management of tumours overexpressing HER2. Future research will lead to higher therapeutic responses, lower toxicities and providing insight into the mechanisms of resistance to HER2-targeted treatments. [ABSTRACT FROM AUTHOR]

The epidermal growth factor receptor (EGFR) has at least two fundamental conformations: an inactive tethered conformation and an active untethered, ligand-bound "back-to- back" dimer, which may be part of an oligomeric complex. Monoclonal antibody (mAb) 806 is an EGFR-specific anti- body that only binds a transitional form of the receptor after it untethers but before forming the back-to-back, ligated, active oligomer. We have shown that AG1478, a tyrosine kinase inhibitor of the EGFR, synergistically inhibits the growth of tumors overexpressing EGFR when used in combination with mAb 806 but the mechanism for this was not elucidated (Johns, T. G., Luwor, R. B., Murone, C., Walker, F., Weinstock, J., Vitali, A. A., Perera, R. M., Jungbluth, A. A., Stockert, E., Old, L. J., Nice, E. C., Burgess, A. W., and Scott, A. M. (2003) Proc. Natl. Acad. Sci. U. S. A. 100, 15871-15876). We now show that AG 1478 increases binding of mAb 806 to the cell surface through two distinct mechanisms: an immediate effect on the conformation of EGFR and a longer term increase in cell surface under-glycosylated EGFR, an event known to increase mAb 806 reactivity. Cross-linking studies demonstrated the presence of spontaneously occurring mAb 806-reactive dimers on the surface of cells overexpressing EGFR, which are rapidly increased by AG1478. Because they react with mAb 806, these dimers must exist in a conformation distinct from the ligated back-to-back dimer. Indeed, we detected similar dimers in 293T cells expressing the EGFR lacking the small dimerization/activation arm essential to the formation of the back-to-back dimer. Thus, some of the EGFR on the cell surface of cancer cells must exist as an untethered dimer that adopts a previously unreported conformation that is inactive. This information was used to optimize the therapeutic synergy between mAb 806 and AG1478 in a xenograft model. [ABSTRACT FROM AUTHOR]

The epidermal growth factor receptor (EGFR) is a member of the erbB tyrosine kinase family of receptors. For many years it has been believed that receptor activation occurs via a monomer-dimer transition that is associated with a conformational change to activate the kinase. However, little is known about the quaternary state of the receptor at normal levels of expression («105 receptors/cell). We employed multidimensional microscopy techniques to gain insight into the state of association of the human EGFR, in the absence and presence of ligand, on the surface of intact BaF/3 cells (50,000 receptors/cell). Image correlation microscopy of an EGFR-enhanced green fluorescent protein chimera was used to establish an average degree of aggregation on the submicron scale of 2.2 receptors/cluster in the absence of ligand increasing to 3.7 receptors/cluster in the presence of ligand. Energy transfer measurements between mixtures of fluorescein isothiocyanate-EGF and Alexa 555-EGF were performed using fluorescence life. time imaging microscopy as a function of the donor: acceptor labeling ratio to gain insight into the spatial disposition of EGFR ligand binding sites on the nanometer scale. In the context of a two-state Förster resonance energy transfer (FRET)/non-RET model, the data are consistent with a minimum transfer efficiency of 75% in the FRET population. The microscopy data are related to biophysical data on the EGFR in the A431 cell line and the three-dimensional structure of the ligated EGFR extracellular domain. In the context of a monomer-dimer-oligomer model, the biophysical data are consistent with a significant fraction of ligated EGFR tetramers comprising two dimers juxtaposed in a side-by-side (or slightly staggered) arrangement. Our data are consistent with a specific higher order association of the ligand-bound EGFR on the nanometer scale and indicate the existence of distinct signaling entities beyond the level of the EGFR dimer which could play an important role in receptor transactivation. [ABSTRACT FROM AUTHOR]

The epidermal growth factor receptor (EGFR) is overexpressed in many epithelial cancers, an observation often correlated with poor clinical outcome. Overexpression of the EGFR is commonly caused by EGFR gene amplification and is sometimes associated with expression of a variant EGFR (de2-7 EGFR or EGFRvIII) bearlng an internal deletion in its extracellular domain. Monoclonal antibody (mAb) 806 is a novel EGFR antibody with significant antitumor activity that recognizes both the de2-7 EGFR and a subset of the wild type (wt) EGFR when overexpressed but does not bind the wt EGFR expressed in normal tissues. Despite only binding to a low proportion of the wt EGFR expressed in A431 tumor cells (∼10%), mAb 806 displays robust antitumor activity against A431 xenografts grown in nude mice. To elucidate the mechanism leading to its unique specificity and mode of antitumor activity, we have determined the EGFR binding epitope of mAb 806. Analysis of mAb 806 binding to EGFR fragments expressed either on the surface of yeast or in an immunoblot format identified a disulfide-bonded loop (amino acids 287-302) that contains the mAb 806 epitope. Indeed, mAb 806 binds with apparent high affinity (∼30 nM) to a synthetic EGFR peptide corresponding to these amino acids. Analysis of EGFR structures indicates that the epitope is fully exposed only in the transitional form of the receptor that occurs because EGFR changes from the inactive tethered conformation to a ligand-bound active form. It would seem that mAb 806 binds this small proportion of transient receptors, preventing their activation, which in turn generates a strong antitumor effect. Finally, our observations suggest that the generation of antibodies to transitional forms of growth factor receptors may represent a novel way of reducing normal tissue targetlng yet retaining antitumor activity. [ABSTRACT FROM AUTHOR]

Recent crystallographic data on the isolated extracellular domain of the epidermal growth factor receptor (EGFR) have suggested a model for its activation by ligand. We have tested this model in the context of the full-length EGFR displayed at the cell surface, by introducing mutations in two regions (CR1 and CR2) of the extracellular domain thought to be critical for regulation of receptor activation. Mutations in the CR1 and CR2 domains have opposing effects on ligand binding affinity, receptor dimerization, tyrosine kinase activation, and signaling competence. Tyr246 is a critical residue in the CR1 loop, which is implicated in the positioning and stabilization of the receptor dimer interface after ligand binding; mutations of Tyr246 impair or abolish receptor function. Mutations in CR2, which weaken the interaction that restricts the receptor to the tethered (inactive) state, enhance responsiveness to EGF by increasing affinity for the ligand. However, weakening of the CR1/CR2 interaction does not result in spontaneous activation of the receptors' kinase. We have used an antibody (mAb 806), which recognizes a transition state of the EGF receptor between the negatively constrained, tethered state and the fully active back-to-back dimer conformation, to follow conformational changes in the wild-type and mutant EGF receptors after ligand binding. Our results suggest that EGFR on the cell surface can be untethered, but this form is inactive; thus, untethering of the receptor is not sufficient for activation, and ligand binding is essential for the correct positioning of the two receptor subunits to achieve kinase activation. [ABSTRACT FROM AUTHOR]