EGFR-Targeting of a Dual Peptide-siRNA Complex for Oral Cancer Treatment

Objectives: The objective of this study was to examine the feasibility of utilizing two peptides, one with cancer cell-targeting specificity and the second with endosome-disruptive properties, to co-deliver bioactive siRNAs into oral cancer cells overexpressing the epidermal growth factor receptor (EGFR) and induce silencing of the targeted oncoprotein, CIP2A.
Methods: To ascertain whether the cellular uptake of the dual peptide-siRNA complex into high-EGFR expressing oral cancer cells was mediated by direct interaction with EGFR, we performed a set of blocking experiments using an anti-EGFR antibody and analyzed the effects by fluorescence microscopy. Subsequently, a xenograft oral cancer tumor mouse model was used to perform in/ex vivo bioimaging to analyze the ability of the dual peptide to mediate delivery of siRNAs designed to target CIP2A (siCIP2A) to tumor tissues. Real-time PCR analysis was used to determine the extent of CIP2A silencing mediated by the different treatment groups in vivo.
Results: Fluorescence microscopy analyses demonstrated that preincubation of EGFR-overexpressing oral cancer cells with an anti-EGFR antibody specifically reduced the cellular uptake of siCIP2As complexed to the dual peptide compared to an IgG isotype control non-specific antibody, suggesting that the specific uptake of siRNAs into cells by the dual peptide carrier was dependent on the availability of EGFR on the cell surface. Furthermore, in/ex vivo imaging and real-time PCR analyses demonstrated the successful delivery and accumulation of bioactive siCIP2As into tumor tissues 48 hours-post tail vein injection of the dual peptide-siCIP2A complex.
Conclusions: Together, these data demonstrate the clinical potential of a dual peptide strategy for siRNA-based therapeutics by mediating the effective targeting and delivery of bioactive siRNAs into EGFR-overexpressing oral cancer cells.