title: A fluorescent imaging method for analyzing the biodistribution of therapeutic monoclonal antibodies that can distinguish intact antibodies from their breakdown products.
abstract: Many monoclonal antibodies have been developed for therapy over the last 2 decades. In the development of therapeutic antibodies, the preclinical assessment of an antibody's biodistribution is important for the prediction of the antibody's efficacy and safety. For imaging analyses of such biodistributions, radioisotope (RI) labeling and fluorescence labeling methods are typically used, but the resulting data are limited because these methods cannot distinguish breakdown products from intact antibodies. To resolve this problem, we developed a novel method using fluorescent resonance energy transfer (FRET)-type labeling and a spectral unmixing tool. With FRET-type labeling (labeling with 2 species of fluorophore), different fluorescence properties of labeled intact antibodies and their breakdown products (the hydrolyzed/digested type of breakdown products) are made visible. With the spectral unmixing tool, the fluorescence of a solution containing the intact antibody and its breakdown products could be unmixed in proportion to their contents. Moreover, when labeled antibodies that targeted either human epidermal growth factor receptor-2 or epidermal growth factor receptor were injected into nude mice implanted subcutaneously with tumor cells, the accumulation of the injected labeled antibodies and their breakdown products in the tumor could be separately analyzed by both whole-mouse imaging and a tumor homogenate analysis. These results suggest that our method using FRET-type labeling and a spectral unmixing tool could be useful in distinguishing breakdown products from intact antibodies.
authors.affiliation: a National Institute of Health Sciences ; Tokyo , Japan.
authors.forename: Takuo
authors.initials: T
authors.lastname: Suzuki
authors.forename: Chihiro
authors.initials: C
authors.lastname: Miyazaki
authors.forename: Akiko
authors.initials: A
authors.lastname: Ishii-Watabe
authors.forename: Minoru
authors.initials: M
authors.lastname: Tada
authors.forename: Kumiko
authors.initials: K
authors.lastname: Sakai-Kato
authors.forename: Toru
authors.initials: T
authors.lastname: Kawanishi
authors.forename: Nana
authors.initials: N
authors.lastname: Kawasaki
mesh_terms: D000818:Animals; D000970:Antineoplastic Agents / Q000737:chemistry / Q000493:pharmacokinetics / Q000494:pharmacology; D045744:Cell Line, Tumor; D031541:Fluorescence Resonance Energy Transfer; D005456:Fluorescent Dyes / Q000737:chemistry / Q000493:pharmacokinetics / Q000494:pharmacology; D006801:Humans; D051379:Mice; D008819:Mice, Nude; D009374:Neoplasms, Experimental / Q000188:drug therapy / Q000378:metabolism / Q000473:pathology; D018719:Erb-b2 Receptor Tyrosine Kinases / Q000037:antagonists & inhibitors*; D000068878:Trastuzumab / Q000737:chemistry / Q000494:pharmacology; D023041:Xenograft Model Antitumor Assays
chemical_list: D000970:Antineoplastic Agents; D005456:Fluorescent Dyes; D018719:Erb-b2 Receptor Tyrosine Kinases; D000068878:Trastuzumab; C508053:ERBB2 protein, human
keywords: antibody; biodistribution; breakdown; fluorescence imaging; fluorescent resonance energy transfer (FRET)