Transfer of Dyes and Drugs into Cells Using EGFR-Targeted Nanosyringes.

Selective targeting of drug loaded nanovectors to specific epitopes highly expressed on the surface of cancer cells is a goal for nanotechnologists. We have modified our previously described PEGylated-hydrophilic carbon clusters (PEG-HCCs) so that the epidermal growth factor receptor (EGFR) binding peptide, GE11, is attached using click chemistry at the end of each PEG. The resulting nanosyringe, PepEGFR-PEG-HCC, can be loaded with a wide range of hydrophobic drugs and dyes. We show that, both in vitro and in vivo, this payload can be delivered to cancer cells expressing EGFR. We can observe the activation of EGFR and track the normal physiological internalization and recycling/signaling pathways of this tyrosine kinase following binding of PepEGFR-PEG-HCC. We also demonstrate the competitive binding of the nanosyringe to EGFR with its normal activator, EGF, as well as observing the colocalization of the nanosyringe with clathrin, the coated pit integral protein. The internalization of the drug/dye loaded nanosyringe can be inhibited by using anti-EGFR antibodies, the drug erlotinib, or Pitstop-1, the clathrin coated pit formation specific inhibitor. To further demonstrate the specificity of the drug loaded nanovectors, we demonstrated that, in both flank and intracranial xenograft mouse models, dye delivery is highly specific to tumors and no other tissues. Finally, using nanosyringes loaded with esterase sensitive fluorescein diacetate, we demonstrated that the drug payloads can be in vivo delivered to the cytosol of cancer cells within the mouse brain.