Abstract
The epidermal growth factor (EGF) receptor is a classical receptor tyrosine kinase with an extracellular ligand-binding domain and an intracellular kinase domain. Mutations in the EGF receptor have been shown to drive uncontrolled cell growth and are associated with a number of different tumors. Two different types of ATP-competitive EGF receptor tyrosine kinase inhibitors have been identified that bind to either the active (type I) or inactive (type II) conformation of the kinase domain. Despite the fact that both types of inhibitors block tyrosine kinase activity, they exhibit differential efficacies in different tumor types. Here, we show that in addition to inhibiting kinase activity, these inhibitors allosterically modulate ligand binding. Our data suggest that the conformations of the EGF receptor extracellular domain and intracellular kinase domain are coupled and that these conformations exist in equilibrium. Allosteric regulators, such as the small-molecule tyrosine kinase inhibitors, as well as mutations in the EGF receptor itself, shift the conformational equilibrium among the active and inactive species, leading to changes in EGF receptor-binding affinity. Our studies also reveal unexpected positive cooperativity between EGF receptor subunits in dimers formed in the presence of type II inhibitors. These findings indicate that there is strong functional coupling between the intracellular and extracellular domains of this receptor. Such coupling may impact the therapeutic synergy between small-molecule tyrosine kinase inhibitors and monoclonal antibodies in vivo.
Highlights
The epidermal growth factor (EGF)2 receptor is a transmembrane receptor tyrosine kinase that is the founding member of the ErbB receptor family [1, 2]
Treatment of CHO cells expressing the EGF receptor with the tyrosine kinase inhibitor, erlotinib, was found to significantly increase the affinity of the receptor for EGF (Fig. 1A) from an EC50 of ϳ3 nM in untreated cells to an EC50 of ϳ0.3 nM after treatment with erlotinib. To determine whether this effect was due to the inhibition of receptor kinase activity, we assessed the effect on EGF binding of lapatinib, another smallmolecule inhibitor of the EGF receptor kinase
Erlotinib is a type I inhibitor that binds to the active conformation of the EGF receptor kinase [17], whereas lapatinib is a type II inhibitor that binds to the inactive conformation of the kinase [18]
Summary
The epidermal growth factor (EGF) receptor is a classical receptor tyrosine kinase with an extracellular ligand-binding domain and an intracellular kinase domain. Our studies reveal unexpected positive cooperativity between EGF receptor subunits in dimers formed in the presence of type II inhibitors These findings indicate that there is strong functional coupling between the intracellular and extracellular domains of this receptor. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health. Despite the fact that both inhibitors block EGF receptor tyrosine kinase activity, they show differential efficacy in different tumor types (19 –21) This suggests that they possess other properties, besides kinase inhibition, that distinguish their effects on tumorigenic forms of the EGF receptor. Our data suggest that the conformations of the EGF receptor extracellular domain and intracellular kinase domain are coupled and that these high affinity and low affinity conformations exist in an equilibrium that can be shifted by small-molecule tyrosine kinase inhibitors.
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