title: An activity-based near-infrared glucuronide trapping probe for imaging β-glucuronidase expression in deep tissues.
abstract: β-glucuronidase is an attractive reporter and prodrug-converting enzyme. The development of near-IR (NIR) probes for imaging of β-glucuronidase activity would be ideal to allow estimation of reporter expression and for personalized glucuronide prodrug cancer therapy in preclinical studies. However, NIR glucuronide probes are not yet available. In this work, we developed two fluorescent probes for detection of β-glucuronidase activity, one for the NIR range (containing IR-820 dye) and the other for the visible range [containing fluorescein isothiocyanate (FITC)], by utilizing a difluoromethylphenol-glucuronide moiety (TrapG) to trap the fluorochromes in the vicinity of the active enzyme. β-glucuronidase-mediated hydrolysis of the glucuronyl bond of TrapG generates a highly reactive alkylating group that facilitates the attachment of the fluorochrome to nucleophilic moieties located near β-glucuronidase-expressing sites. FITC-TrapG was selectively trapped on purified β-glucuronidase or β-glucuronidase-expressing CT26 cells (CT26/mβG) but not on bovine serum albumin or non-β-glucuronidase-expressing CT26 cells used as controls. β-glucuronidase-activated FITC-TrapG did not interfere with β-glucuronidase activity and could label bystander proteins near β-glucuronidase. Both FITC-TrapG and NIR-TrapG specifically imaged subcutaneous CT26/mβG tumors, but only NIR-TrapG could image CT26/mβG tumors transplanted deep in the liver. Thus NIR-TrapG may provide a valuable tool for visualizing β-glucuronidase activity in vivo.
authors.affiliation: Graduate Institute of Medicine, Kaohsiung Medical University, Kaohsiung, Taiwan.
authors.forename: Ta-Chun
authors.initials: TC
authors.lastname: Cheng
authors.forename: Steve R
authors.initials: SR
authors.lastname: Roffler
authors.forename: Shey-Cherng
authors.initials: SC
authors.lastname: Tzou
authors.forename: Kuo-Hsiang
authors.initials: KH
authors.lastname: Chuang
authors.forename: Yu-Cheng
authors.initials: YC
authors.lastname: Su
authors.forename: Chih-Hung
authors.initials: CH
authors.lastname: Chuang
authors.forename: Chien-Han
authors.initials: CH
authors.lastname: Kao
authors.forename: Chien-Shu
authors.initials: CS
authors.lastname: Chen
authors.forename: I-Hong
authors.initials: IH
authors.lastname: Harn
authors.forename: Kuan-Yi
authors.initials: KY
authors.lastname: Liu
authors.forename: Tian-Lu
authors.initials: TL
authors.lastname: Cheng
authors.forename: Yu-Ling
authors.initials: YL
authors.lastname: Leu
mesh_terms: D000818:Animals; D002417:Cattle; D045744:Cell Line, Tumor; D003432:Cross-Linking Reagents / Q000737:chemistry; D004354:Drug Screening Assays, Antitumor / Q000379:methods; D005456:Fluorescent Dyes / Q000737:chemistry; D005966:Glucuronidase / Q000096:biosynthesis* / Q000737:chemistry*; D020719:Glucuronides / Q000737:chemistry*; D006801:Humans; D008099:Liver / Q000473:pathology; D008247:Lysosomes / Q000378:metabolism; D051379:Mice; D008807:Mice, Inbred BALB C; D009368:Neoplasm Transplantation; D009369:Neoplasms / Q000473:pathology; D011355:Prodrugs / Q000737:chemistry; D012710:Serum Albumin, Bovine / Q000378:metabolism; D013055:Spectrophotometry, Infrared / Q000379:methods; D019265:Spectroscopy, Near-Infrared / Q000379:methods
chemical_list: D003432:Cross-Linking Reagents; D005456:Fluorescent Dyes; D020719:Glucuronides; D011355:Prodrugs; D012710:Serum Albumin, Bovine; D005966:Glucuronidase