Probing EGFR, HER2 and c-Met protein-protein interactions using an antibody array.

The receptor tyrosine kinase (RTK) family comprises important disease drivers and validated drug targets. Cooperation between certain RTKs is often achieved through physical interactions and is thought be a contributing factor to some cases of tumor resistance to targeted therapies. Epidermal Growth Factor Receptor (EGFR), HER2/ErbB2, and c-Met are prominent examples of disease driver RTKs that cooperate and form heterodimers. Current methods to detect RTK protein-protein interactions lack proper throughput, are not quantitative, or rely on highly engineered systems and cumbersome workflows. We have developed a planar surface antibody array-based sandwich assay that enables the detection and relative quantification of interactions between EGFR, HER2, and c-Met. The antibody-based assay revealed co-existing combinatorial receptor interactions that were unique to each cell type tested. The assay also pointed to an existence of distinct topological configurations of EGFR-c-Met and EGFR-HER2 heterodimers, unveiling previously not described nuances of these interactions. Lastly, the array-based assay allowed monitoring the disruption of receptor complexes using small molecule inhibitors and revealed differential sensitivity of EGFR-HER2 and EGFR-c-Met complexes to a small molecule EGFR kinase inhibitor, gefitinib. The antibody array sandwich assay described here allows utilizing larger panel of affinity reagents and can be applied to broader range of cases with various topological requirements, thus offering an advantage over technologies that have strict proximity requirement. This novel protein interaction array enables parallel detection of RTK protein-protein interactions, allowing a more systematic monitoring of response to activators or inhibitors. Citation Information: Mol Cancer Ther 2013;12(11 Suppl):C202. Citation Format: Jacob Knowles, Stephen Capodilupo, Richard Noring, Roberto Campos-Gonzalez, Julio Herrera, Zev Gechtman. Probing EGFR, HER2 and c-Met protein-protein interactions using an antibody array. [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2013 Oct 19-23; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2013;12(11 Suppl):Abstract nr C202.