title: Tempo-spatial Activation of Sequential Quadruple Stimuli for High Gene Expression of Polymeric Gene Nanocomplexes.
abstract: The clinical application of intracellular gene delivery via nanosized carriers is hindered by intracellular multistep barriers that limit high levels of gene expression. To solve these issues, four different intracellular or external stimuli that can efficiently activate a gene carrier, a gene, or a photosensitizer (pheophorbide A [PhA]) were assessed in this study. The designed nanosized polymeric gene complexes were composed of PhA-loaded thiol-degradable polycation (PhA@RPC) and cytomegalovirus (CMV) promoter-equipped pDNA. After cellular internalization of the resulting PhA@RPC/pDNA complexes, the complexes escaped endosomal sequestration, owing to the endosomal pH-induced endosomolytic activity of RPC in PhA@RPC. Subsequently, intracellular thiol-mediated polycation degradation triggered the release of PhA and pDNA from the complexes. Late exposure to light (for example, 12 h post-treatment) activated the released PhA and resulted in the production of reactive oxygen species (ROS). Intracellular ROS successively activated NF-κB, which then reactivated the CMV promoter in the pDNA. These sequential, stimuli-responsive chemical and biological reactions resulted in high gene expression. In particular, the time-point of light exposure was very significant to tune efficient gene expression as well as negligible cytotoxicity: early light treatment induced photochemical internalization but high cytotoxicity, whereas late light treatment influenced the reactivation of silent pDNA via PhA-generated ROS and activation of NF-κB. In conclusion, the quadruple triggers, such as pH, thiol, light, and ROS, successively influenced a gene carrier (RPC), a photosensitizer, and a genetic therapeutic, and the tempo-spatial activation of the designed quadruple stimuli-activatable nanosized gene complexes could be potential in gene delivery applications.
authors.affiliation: Department of Pharmacy and Integrated Research Institute of Pharmaceutical Sciences, College of Pharmacy, The Catholic University of Korea , 43 Jibong-ro, Bucheon-si, Gyeonggi-do 14662, Republic of Korea.
authors.forename: Hana
authors.initials: H
authors.lastname: Cho
authors.affiliation: Next-generation Pharmaceutical Research Center, Korea Institute of Toxicology , 141 Gajeong-ro, Yuseong-gu, Daejeon, 34114, Republic of Korea.
authors.forename: Young-Woo
authors.initials: YW
authors.lastname: Cho
authors.affiliation: Next-generation Pharmaceutical Research Center, Korea Institute of Toxicology , 141 Gajeong-ro, Yuseong-gu, Daejeon, 34114, Republic of Korea.
authors.forename: Sun-Woong
authors.initials: SW
authors.lastname: Kang
authors.affiliation: Department of Pharmacy and Integrated Research Institute of Pharmaceutical Sciences, College of Pharmacy, The Catholic University of Korea , 43 Jibong-ro, Bucheon-si, Gyeonggi-do 14662, Republic of Korea.
authors.forename: Mi-Kyoung
authors.initials: MK
authors.lastname: Kwak
authors.affiliation: Department of Polymer Science and Engineering, Chungnam National University , 99 Daehak-ro, Yuseong-gu, Daejeon 34134, Republic of Korea.
authors.forename: Kang Moo
authors.identifier: 0000-0002-2406-6659
authors.initials: KM
authors.lastname: Huh
authors.affiliation: Department of Pharmaceutics and Pharmaceutical Chemistry, The University of Utah , 30 S 2000 E, Rm 2972, Salt Lake City, Utah 84112, United States.|Utah-Inha Drug Delivery Systems (DDS) and Advanced Therapeutics Research Center , 9 Songdomirae-ro, Yeonsu-gu, Incheon 21988, Republic of Korea.
authors.forename: You Han
authors.initials: YH
authors.lastname: Bae
authors.affiliation: Department of Pharmacy and Integrated Research Institute of Pharmaceutical Sciences, College of Pharmacy, The Catholic University of Korea , 43 Jibong-ro, Bucheon-si, Gyeonggi-do 14662, Republic of Korea.
authors.forename: Han Chang
authors.identifier: 0000-0003-0696-1155
authors.initials: HC
authors.lastname: Kang
mesh_terms: D045744:Cell Line, Tumor; D002734:Chlorophyll / Q000008:administration & dosage / Q000031:analogs & derivatives; D004247:DNA / Q000378:metabolism*; D011992:Endosomes / Q000187:drug effects / Q000378:metabolism; D015870:Gene Expression / Q000187:drug effects*; D018014:Gene Transfer Techniques; D006367:HeLa Cells; D006801:Humans; D006863:Hydrogen-Ion Concentration; D016328:NF-kappa B / Q000378:metabolism; D053758:Nanoparticles / Q000008:administration & dosage*; D017319:Photosensitizing Agents / Q000008:administration & dosage; D010957:Plasmids / Q000235:genetics; D011073:Polyamines / Q000008:administration & dosage; D000071228:Polyelectrolytes; D011108:Polymers / Q000008:administration & dosage*; D017382:Reactive Oxygen Species / Q000378:metabolism; D014162:Transfection / Q000379:methods
chemical_list: D002734:Chlorophyll; D004247:DNA; D016328:NF-kappa B; D017319:Photosensitizing Agents; D011073:Polyamines; D000071228:Polyelectrolytes; D011108:Polymers; D017382:Reactive Oxygen Species; C032623:pheophorbide a
keywords: CMV promoter; photosensitizer; polymeric gene delivery; reactive oxygen species; stimuli-responsive gene delivery