title: Role of polymeric endosomolytic agents in gene transfection: a comparative study of poly(L-lysine) grafted with monomeric L-histidine analogue and poly(L-histidine).
abstract: Endosomal entrapment is one of the main barriers that must be overcome for efficient gene expression along with cell internalization, DNA release, and nuclear import. Introducing pH-sensitive ionizable groups into the polycationic polymers to increase gene transfer efficiency has proven to be a useful method; however, a comparative study of introducing equal numbers of ionizable groups in both polymer and monomer forms, has not been reported. In this study, we prepared two types of histidine-grafted poly(L-lysine) (PLL), a stacking form of poly(L-histidine) (PLL-g-PHis) and a mono-L-histidine (PLL-g-mHis) with the same number of imidazole groups. These two types of histidine-grafted PLL, PLL-g-PHis and PLL-g-mHis, showed profound differences in hemolytic activity, cellular uptake, internalization, and transfection efficiency. Cy3-labeled PLL-g-PHis showed strong fluorescence in the nucleus after internalization, and high hemolytic activity upon pH changes was also observed from PLL-g-PHis. The arrangement of imidazole groups from PHis also provided higher gene expression than mHis due to its ability to escape the endosome. mHis or PHis grafting reduced the cytotoxicity of PLL and changed the rate of cellular uptake by changing the quantity of free ε-amines available for gene condensation. The subcellular localization of PLL-g-PHis/pDNA measured by YOYO1-pDNA intensity was highest inside the nucleus, while the lysotracker, which stains the acidic compartments was lowest among these polymers. Thus, the polymeric histidine arrangement demonstrate the ability to escape the endosome and trigger rapid release of polyplexes into the cytosol, resulting in a greater amount of pDNA available for translocation to the nucleus and enhanced gene expression.
authors.affiliation: Department of Pharmaceutics and Pharmaceutical Chemistry, The University of Utah , Skaggs Research Building, Rm 2760, 30S, 2000E, Salt Lake City, Utah 84112, United States.
authors.forename: Hee Sook
authors.initials: HS
authors.lastname: Hwang
authors.forename: Jun
authors.initials: J
authors.lastname: Hu
authors.forename: Kun
authors.initials: K
authors.lastname: Na
authors.forename: You Han
authors.initials: YH
authors.lastname: Bae
mesh_terms: D045744:Cell Line, Tumor; D002470:Cell Survival / Q000187:drug effects; D004247:DNA / Q000008:administration & dosage / Q000737:chemistry; D011992:Endosomes / Q000187:drug effects*; D015870:Gene Expression / Q000187:drug effects; D006367:HeLa Cells; D006639:Histidine / Q000008:administration & dosage* / Q000737:chemistry*; D006801:Humans; D061986:MCF-7 Cells; D010957:Plasmids / Q000737:chemistry; D011107:Polylysine / Q000008:administration & dosage* / Q000737:chemistry*; D011108:Polymers / Q000008:administration & dosage* / Q000737:chemistry*; D014162:Transfection / Q000379:methods
chemical_list: D011108:Polymers; D011107:Polylysine; C033223:polyhistidine; D006639:Histidine; D004247:DNA