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.