Outsmarting refractory glioblastoma usingin silicoguided, phage display derived peptides targeting oncogenic EGFRvIII

ABSTRACT EGFRvIII mutation contributes to tumor aggressiveness and poor prognosis in multiple cancers. Despite being tumor-specific, EGFRvIII are not readily detected since they coexist with EGFR. Here, we developed novel, aqueous-dispersible peptides (1-15 kDa) that can detect and actively target EGFRvIII expressing refractory tumors, while sparing wild-type EGFR and normal astrocytes. Based on in silico molecular modeling, docking and biologics-based approaches, four EGFRvIII-targeting peptides were identified using phage display technology employing a) antibody fragments (AF), and b) random peptide library (RPL). Among RPL-based peptides, RB5 showed appreciable interactions with EGFRvIII receptor, as indicated by binding energy of -10.6 kcal/mol, in silico. In vitro binding studies using ELISA and immunocytochemistry further confirmed that RB5 could specifically target EGFRvIII+ glioblastoma cell, U87MG.ΔEGFR. Interestingly, AF-based peptide, H13, also showed considerable specificity towards U87MG.ΔEGFR. Importantly, these peptides exerted no cytotoxicity (MTS assay), or affected the downstream phosphorylation (western blot) in EGFRvIII cells. Thus, our study demonstrated that rationally designed peptide molecules can precisely target EGFRvIII tumors with minimal or no intracellular signaling interference. Owing to simple molecular architectures, these non-toxic, aqueous-dispersible peptides hold strong potential for conjugation with antitumor drugs and therapeutic nanoparticles for treating EGFRvIII positive refractory cancers.