Methods for Identifying Subcellular Targeting Ligands and Selected Applications

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2015-09-09

Authors

Umlauf, Benjamin J.

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Abstract

Subcellular localization plays an essential role in targeting drug therapies as generally the pro-drug or linker relies on physical conditions of a particular subcellular compartment to function. We developed two methods that allow for selecting targeting ligands that both internalize specifically in cancer cells as well as accumulate in a defined subcellular location. The first method utilizes endocytic selection pressure to identify targeting peptides from a phage displayed peptide library that internalize specifically in cancer cells via a defined mechanism of endocytosis while the second couples together a traditional biopanning selection protocol with a secondary immunofluorescent screen to directly identify functional targeting peptides. In addition we also present a method for reducing amplification bias during phage display experiments. Finally, we demonstrate the utility of a novel peptide, termed H1299.3 selected via the endocytic pressure method, to serve as targeting ligand for a novel immunotherapy. H1299.3 targeted liposomes facilitate delivery of antigenic peptide specifically in MHC class 1 molecules of cancer cells resulting in activation of secondary immune response against the cancer cells. Thus, we present two novel methods for identify targeting ligands with selective internalization that accumulate in defined subcellular localizations as well as a novel application for a newly identified targeting ligand.

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