Novel Strategy Utilizing Extracellular Cysteine-Rich Domain of Membrane Receptor for Constructing D-Peptide Mediated Targeted Drug Delivery Systems: A Case Study on Fn14

The development of proteolysis-resistant D-peptide ligands for targeted drug/gene delivery has been greatly limited, due to the challenge that lies in the chemical synthesis of membrane receptors without altering their structures. In the present research, a novel strategy utilizing self-stabilized extracellular CRD of the membrane receptor was developed to construct D-peptide ligands and their mediated targeted drug delivery systems. Fn14, a cell surface receptor overexpressed in many cancers including pancreatic and triple negative breast cancers, was selected as the model receptor. Fn14 CRD was synthesized and folded, and used to screen Fn14 binding peptides using phage display (L-peptide) and mirror-image phage display (D-peptide) techniques, respectively. The D-peptide ligand successfully mediated targeted drug delivery to Fn14 positive tumor cells. In addition, the D-peptide possessed better target-binding affinity, stromal barrier permeability, and tumor targeting ability in vivo when conjugated with liposomes. More importantly, D-peptide mediated liposomal paclitaxel delivery significantly inhibited pancreatic tumor growth in a subcutaneous xenograft model and drastically prolonged survival in a lung metastasis of breast cancer mouse model. This study demonstrated that mirror-image phage display based on the CRD of membrane receptor can be a promising strategy to advance active targeted drug delivery via biostable D-peptides.