Enhanced Intracellular Protein Transduction by Sequence Defined Tetra-Oleoyl Oligoaminoamides Targeted for Cancer Therapy

Intracellular protein delivery presents a novel promising prospect for cell biology research and cancer therapy. However, inefficient cellular uptake and lysosomal sequestration hinder productive protein delivery into the cytosol. Here, a library of 16 preselected sequence-defined oligoaminoamide oligomers is evaluated for intracellular protein delivery. All oligomers, containing polyethylene glycol (PEG) for shielding and optionally folic acid as targeting ligand, manifest cellular internalization of disulfide-conjugated enhanced green fluorescent protein (EGFP). However, only a PEGylated folate-receptor targeted two-arm oligomer (729) containing both arms terminally modified with two oleic acids shows persistent intracellular protein survival and nuclear import of nlsEGFP (which contains a nuclear localization sequence) in folate-receptor-positive KB carcinoma cells, validating both effective endolysosomal escape and following subcellular transport. Furthermore, using ribonuclease A as a therapeutic cargo protein, among the tested oligomers, the oleic acid modified targeted two-arm oligomers exert the most significant tumor cell killing of KB tumor cells. An investigation of structure-activity relationship elucidates that the incorporated oleic acids play a vital role in the enhanced intracellular protein delivery, by promoting stable formation of 25-35 nm lipo-oligomer protein nanoparticles and by membrane-active characteristics facilitating intracellular cytosolic delivery.