title: Cellular uptake and in vivo distribution of polyhistidine peptides.
abstract: Cell-penetrating peptides (CPPs) are arginine/lysine-rich sequences, and they are effectively internalized into cells. In this process, positive charge is crucial. In the present study, we found polyhistidine peptides (PHPs), as the novel CPP, which are efficiently internalized into cells in a positive charge-independent manner. Interestingly, cellular uptake of the PHPs increased as the chain length increased, reaching a maximum uptake at H16 (HHHHHHHHHHHHHHHH-NH2). This H16 peptide showed up to 14.6-fold higher cell-penetrating capacity against HT1080 human fibrosarcoma cells relative to a major CPP, the octa-arginine (RRRRRRRR-NH2) peptide. Cellular uptake of the H16 peptide is mainly due to macropinocytosis and most of the H16 peptide localizes in the lysosome and Golgi apparatus. However, a cytoplasmic pro-apoptotic domain (KLAKLAKKLAKLAK-NH2) conjugated to the H16 peptide showed cytotoxic effects. This indicates that a proportion of the H16 peptide escapes from the macropinosome to the cytoplasm. In a protein transduction study, green fluorescence protein fused to the H16 peptide (GFP-H16) was purified by Ni-NTA chromatography, detected using an anti-His-tag antibody and internalized into cells. This serial process reveals that H16 functions as a His-tag and protein transduction domain. Furthermore, in vivo distribution analysis showed that the H16 peptide accumulates immediately in tumor tissue and is retained up to 132h following injection into the tumor (HT1080 human fibrosarcoma)-bearing mice. This is the first observation of a His-polymer being internalize into cells efficiently. The findings suggest that PHPs are novel CPPs. In particular, the H16 peptide represents a promising drug delivery carrier candidate in medical and biotechnological fields.
authors.affiliation: Department of Bioresources Science, Graduate School of Agricultural Sciences, Tottori University, Tottori 680-8553, Japan. Electronic address: itaka@muses.tottori-u.ac.jp.
authors.forename: Takashi
authors.initials: T
authors.lastname: Iwasaki
authors.affiliation: Department of Bioresources Science, Graduate School of Agricultural Sciences, Tottori University, Tottori 680-8553, Japan.
authors.forename: Yoshihisa
authors.initials: Y
authors.lastname: Tokuda
authors.affiliation: Department of Bioresources Science, Graduate School of Agricultural Sciences, Tottori University, Tottori 680-8553, Japan.
authors.forename: Ayaka
authors.initials: A
authors.lastname: Kotake
authors.affiliation: Faculty of Exploratory Technology, Asubio Pharma Co., Ltd., Kobe 650-0047, Japan.
authors.forename: Hiroyuki
authors.initials: H
authors.lastname: Okada
authors.affiliation: Faculty of Exploratory Technology, Asubio Pharma Co., Ltd., Kobe 650-0047, Japan.
authors.forename: Shuji
authors.initials: S
authors.lastname: Takeda
authors.affiliation: Department of Bioresources Science, Graduate School of Agricultural Sciences, Tottori University, Tottori 680-8553, Japan.
authors.forename: Tsuyoshi
authors.initials: T
authors.lastname: Kawano
authors.affiliation: Division of Functional Genomics, Research Center for Bioscience and Technology, Tottori University, Yonago 683-8503, Japan.
authors.forename: Yuji
authors.initials: Y
authors.lastname: Nakayama
mesh_terms: D000818:Animals; D002460:Cell Line; D045744:Cell Line, Tumor; D002470:Cell Survival / Q000187:drug effects; D057846:Cell-Penetrating Peptides / Q000493:pharmacokinetics* / Q000494:pharmacology; D016503:Drug Delivery Systems; D004705:Endocytosis; D049452:Green Fluorescent Proteins / Q000235:genetics; D006639:Histidine / Q000493:pharmacokinetics* / Q000494:pharmacology; D006801:Humans; D008819:Mice, Nude; D009369:Neoplasms / Q000378:metabolism; D014018:Tissue Distribution
chemical_list: D057846:Cell-Penetrating Peptides; D049452:Green Fluorescent Proteins; C033223:polyhistidine; D006639:Histidine
keywords: Cell-penetrating peptide (CPP); Endocytosis; Histidine; Human fibrosarcoma; Macropinocytosis