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Cell-penetrating mechanism of intracellular targeting albumin: Contribution of macropinocytosis induction and endosomal escape

Author:
Ichimizu, Shota, Watanabe, Hiroshi, Maeda, Hitoshi, Hamasaki, Keisuke, Ikegami, Komei, Chuang, Victor Tuan Giam, Kinoshita, Ryo, Nishida, Kento, Shimizu, Taro, Ishima, Yu, Ishida, Tatsuhiro, Seki, Takahiro, Katsuki, Hiroshi, Futaki, Shiroh, Otagiri, Masaki, Maruyama, Toru
Source:
Journal of controlled release 2019 v.304 pp. 156-163
ISSN:
0168-3659
Subject:
CXCR4 receptor, Western blotting, cytosol, dextran, drug carriers, electrostatic interactions, endosomes, fluorescent antibody technique, heparan sulfate, heparin, human serum albumin, lysosomes, proteoglycans, signal transduction, staining
Abstract:
We recently developed a cell-penetrating drug carrier composed of albumin (HSA) combined with palmitoyl-cyclic-(D-Arg)12. While it is possible that the palmitoyl-cyclic-(D-Arg)12/HSA enters the cell mainly via macropinocytosis, the mechanism responsible for the induction of macropinocytosis and endosomal escape remain unknown. We report herein that palmitoyl-cyclic-(D-Arg)12/HSA might interact with heparan sulfate proteoglycan and the chemokine receptor CXCR4 followed by multiple activations of the PKC/PI3K/JNK/mTOR signaling pathways to induce macropinocytosis. This result was further confirmed by a co-treatment with 70 kDa dextran, a macropinocytosis marker. Using liposomes that mimic endosomes, the leakage of 5,6-carboxyfluorescein from liposome was observed in the presence of palmitoyl-cyclic-(D-Arg)12/HSA only in the case of the anionic late endosome-like liposomes but not the neutral early endosome-like liposomes. Heparin largely inhibited this leakage, suggesting the importance of electrostatic interactions between palmitoyl-cyclic-(D-Arg)12/HSA and the late-endosomal membrane. Immunofluorescence staining and Western blotting data indicated that the intact HSA could be transferred from endosomes to the cytosol. These collective data suggest that the palmitoyl-cyclic-(D-Arg)12/HSA is internalized via macropinocytosis and intact HSA is released from the late endosomes to the cytoplasm before the endosomes fuse with lysosomes. Palmitoyl-cyclic-(D-Arg)12/HSA not only functions as an intracellular drug delivery carrier but also as an inducer of macropinocytosis.
Agid:
6441784