Experimental System Design and Modelling of EGFR Extracellular Domain and Its Mutant Binding to Antibody Interacting Partner

TL;DR

This study models how a specific EGFR mutation affects antibody binding and drug resistance in cancer, offering a method to detect the mutation for personalized treatment.

Contribution

A novel experimental and computational approach to model EGFR mutation effects on antibody binding and develop specific detection probes.

Findings

The R497K mutation in EGFR reduces Cetuximab binding affinity by altering domain III structure.

Cetuximab also binds to domain IV of MEGFR, suggesting alternative interaction sites.

Designed oligonucleotide probes can specifically detect the R497K mutation in transfected cells.

Abstract

The EGFR pathway is activated by ligand binding, and EGFR overexpression is linked to malignancies like colorectal and head and neck cancer. This pathway is targeted by monoclonal antibodies such as Cetuximab; however, drug resistance can arise, frequently because of EGFR gene alterations like mutation, particularly in domain III, which inhibits Cetuximab binding. EGFR and MEGFR (R497K mutated EGFR) plasmids were transfected into Chinese hamster ovary (CHO) cells, which do not express EGFR. Real-time PCR was performed using probes that were specifically developed for the R497K mutation. Furthermore, Cetuximab binding to EGFR and MEGFR was examined using molecular modeling. According to molecular modeling, the R497K mutation modifies the domain III structure, which lowers the binding affinity of Cetuximab. Curiously, Cetuximab also showed binding to MEGFR’s domain IV. Real-time PCR showed that the probes specifically identified MEGFR in transfected CHO cells. The R497K mutation may result in treatment resistance by decreasing Cetuximab binding or increasing competitive ligand binding. Therefore, for individualized treatment, it is essential to find EGFR mutations in patient tumor samples. The R497K mutation may be successfully detected by the designed oligonucleotide probes, allowing for the early identification of potential resistance and directing the development of suitable treatment strategies.

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