title: Development of a multifunctional luciferase reporters system for assessing endoplasmic reticulum-targeting photosensitive compounds.
abstract: Photodynamic therapy (PDT) is a recently developed antitumor modality utilizing the generation of reactive oxygen species (ROS), through light irradiation of photosensitizers (PSs) localized in tumor. Interference with proper functioning of endoplasmic reticulum (ER) by ER-targeting PDT is a newly proposed strategy to achieve tumor cell death. The aim of this study is to establish a multifunctional model to screen and assess ER-targeting PSs based on luciferase reporters system. Upregulation of GRP78 is a biomarker for the onset of ER stress. CHOP is a key initiating player in ER stress-induced cell death. Here, the most sensitive fragments of GRP78 and CHOP promoters responding to ER-targeting PDT were mapped and cloned into pGL3-basic vector, forming -702/GRP78-Luc and -443/CHOP-Luc construct, respectively. We demonstrated that -702/GRP78-Luc expression can be used to indicate the ER-targeting of PSs, meanwhile estimate the ROS level induced by low-dose ER-targeting PDT. Moreover, the luciferase signaling of -443/CHOP-Luc showed highly consistence with apoptosis rate caused by ER-targeting PDT, suggesting that -443/CHOP-Luc can evaluate the antitumor properties of PSs. Hypericin, Foscan® and methylene blue were applied to verify the sensitivity and reliability of our model. These results proved that GRP78-CHOP model may be suitable to screen ER-targeting photosensitive compounds with lower cost and higher sensitivity than traditional ways.
authors.affiliation: Department of Molecular & Cellular Pharmacology, Biomedical Nanotechnology Center, State Key Laboratory of Bioreactor Engineering & Shanghai Key Laboratory of New Drug Design, School of Pharmacy, East China University of Science and Technology, #268, 130 Meilong Road, Shanghai, 200237, People's Republic of China.
authors.forename: Shengchao
authors.initials: S
authors.lastname: Lin
authors.forename: Lingling
authors.initials: L
authors.lastname: Zhang
authors.forename: Kecheng
authors.initials: K
authors.lastname: Lei
authors.forename: Anle
authors.initials: A
authors.lastname: Zhang
authors.forename: Ping
authors.initials: P
authors.lastname: Liu
authors.forename: Jianwen
authors.initials: J
authors.lastname: Liu
mesh_terms: D000873:Anthracenes; D017209:Apoptosis / Q000187:drug effects; D004305:Dose-Response Relationship, Drug; D004721:Endoplasmic Reticulum / Q000187:drug effects* / Q000378:metabolism / Q000473:pathology; D000091342:Endoplasmic Reticulum Chaperone BiP; D017930:Genes, Reporter; D006360:Heat-Shock Proteins / Q000235:genetics*; D057166:High-Throughput Screening Assays / Q000379:methods*; D006801:Humans; D008156:Luciferases / Q000096:biosynthesis* / Q000235:genetics; D061986:MCF-7 Cells; D008652:Mesoporphyrins / Q000494:pharmacology; D008751:Methylene Blue / Q000494:pharmacology; D018384:Oxidative Stress / Q000187:drug effects; D010569:Perylene / Q000031:analogs & derivatives / Q000494:pharmacology; D010778:Photochemotherapy; D017319:Photosensitizing Agents / Q000494:pharmacology*; D011401:Promoter Regions, Genetic / Q000187:drug effects*; D017382:Reactive Oxygen Species / Q000378:metabolism; D051742:Transcription Factor CHOP / Q000235:genetics*; D014162:Transfection
chemical_list: D000873:Anthracenes; C497249:DDIT3 protein, human; D000091342:Endoplasmic Reticulum Chaperone BiP; C000717247:HSPA5 protein, human; D006360:Heat-Shock Proteins; D008652:Mesoporphyrins; D017319:Photosensitizing Agents; D017382:Reactive Oxygen Species; D051742:Transcription Factor CHOP; D010569:Perylene; C004965:hypericin; D008156:Luciferases; C072269:temoporfin; D008751:Methylene Blue