Oncogenic mutations at the EGFR ectodomain structurally converge to remove a steric hindrance on a kinase-coupled cryptic epitope

• Published in: Proceedings of the National Academy of Sciences - PNAS 2019-05, Vol.116 (20), p.10009-10018
• Main Authors: Orellana, Laura, Thorne, Amy H., Lema, Rafael, Gustavsson, Johan, Parisian, Alison D., Hospital, Adam, Cordeiro, Tiago N., Bernadó, Pau, Scott, Andrew M., Brun-Heath, Isabelle, Lindahl, Erik, Cavenee, Webster K., Furnari, Frank B., Orozco, Modesto
• Format: Article
• Language: English
• Subjects: Biological Sciences; cancer; Clonal deletion; Computer simulation; Convergence; Coupling (molecular); cryptoepitope; Dimerization; Epidermal growth factor; Epidermal growth factor receptors; Epitopes; Flow cytometry; Gene deletion; Genes, erbB-1; Glioblastoma - genetics; Growth factors; HEK293 Cells; Humans; intermediate; Kinases; Missense mutation; Molecular dynamics; Molecular modelling; Mutation; Mutation, Missense; mutational heterogeneity; Physical Sciences; PNAS Plus; Small angle X ray scattering; Steric hindrance; structural convergence; X-ray scattering
• ISSN: 0027-8424; 1091-6490; 1091-6490
• DOI: 10.1073/pnas.1821442116

Epidermal growth factor receptor (EGFR) signaling is initiated by a large ligand-favored conformational change of the extracellular domain (ECD) from a closed, self-inhibited tethered monomer, to an open untethered state, which exposes a loop required for strong dimerization and activation. In glioblastomas (GBMs), structurally heterogeneous missense and deletion mutations concentrate at the ECD for unclear reasons. We explore the conformational impact of GBM missense mutations, combining elastic network models (ENMs) with multiple molecular dynamics (MD) trajectories. Our simulations reveal that the main missense class, located at the I-II interface away from the self-inhibitory tether, can unexpectedly favor spontaneous untethering to a compact intermediate state, here validated by smallangle X-ray scattering (SAXS). Significantly, such intermediate is characterized by the rotation of a large ECD fragment (N-TR1), deleted in the most common GBM mutation, EGFRvIII, and that makes accessible a cryptic epitope characteristic of cancer cells. This observation suggested potential structural equivalence of missense and deletion ECD changes in GBMs. Corroborating this hypothesis, our FACS, in vitro, and in vivo data demonstrate that entirely different ECD variants all converge to remove N-TR1 steric hindrance from the 806-epitope, which we show is allosterically coupled to an intermediate kinase and hallmarks increased oncogenicity. Finally, the detected extraintracellular coupling allows for synergistic cotargeting of the intermediate with mAb806 and inhibitors, which is proved herein.