Chemotherapy has long been applied for the treatment of cancer. However, chemotherapy can cause various side effects due to their non-targetability and toxicity. Recently, cell-penetrating peptide (CPPs) have attracted attention as a safe non-viral vector. CPPs are composed of naturally occurring amino acids and can deliver a variety of substances because of its ability to penetrate cells. However, CPPs possess some limitations which are unstability and low delivery efficacy of high-molecular-weight substances because of its linear structure and short sequences. To overcome these drawbacks, it is necessary to develop the new types of CPPs.
In chapter 2, we designed a modified nona-arginine (mR9, Cys-R9-Cys-R9-Cys) and synthesized a branched-mR9 (B-mR9) using disulfide bonds. Redox-cleavability of B-mR9 was confirmed by treating reducing agents, which indicated that selective intracellular gene release was achieved. This novel CPP having branched structure demonstrated higher pDNA transfection efficacy and cell viability than conventional CPPs and PEI 25k. In the siRNA experiments, the B-mR9/VEGF siRNA (siVEGF) polyplex exhibited an incremental serum resistance and silencing effects in various cancer cell lines. To demonstrate the therapeutic efficacy in vivo, B-mR9/siVEGF was administered intravenously into NCI-H460 bearing mice. The mice treated with B-mR9 showed significant tumor inhibition compared to the other experimental groups without toxicity. The branched CPP will be probably used as the novel gene delivery platform.
In chapter 3, we synthesized a poly(L-methionine-block-L-lysine)-PLGLAG-PEG (MLMP) composed of a ROS-responsive methionine, a cell permeable lysine chain, and a protease-cleavable linker. The micelles formed by the methionine and lysine chain loads DOX, and the cell-permeability is masked by PEGylation. When the DOX-loaded MLMPs reach a tumor tissue, the linkers are cleaved by matrix metalloproteinase (MMP). After cleavages, the PEG chains are released and micelles can penetrate into cells by revealing cell-permeability. The phase transition of the methionine chain occurs due to abundant ROS in the cytosol of cancer cells, resulting in the collapse of the micelle structure and the release of DOX. We confirm ROS- and enzyme-responsiveness, and outstanding DOX delivery efficacy of the MLMP. In an in vivo study, its long retention with tumor targetability is confirmed by a bio-distribution analysis, and MLMP (DOX) exhibits significant tumor inhibition compared to free DOX. The ROS- and protease-responsive MLMP can likely be used as a novel drug delivery platform.