title: Modular strategy for the construction of radiometalated antibodies for positron emission tomography based on inverse electron demand Diels-Alder click chemistry.
abstract: A modular system for the construction of radiometalated antibodies was developed based on the bioorthogonal cycloaddition reaction between 3-(4-benzylamino)-1,2,4,5-tetrazine and the strained dienophile norbornene. The well-characterized, HER2-specific antibody trastuzumab and the positron emitting radioisotopes (64)Cu and (89)Zr were employed as a model system. The antibody was first covalently coupled to norbornene, and this stock of norbornene-modified antibody was then reacted with tetrazines bearing the chelators 1,4,7,10-tetraazacyclo-dodecane-1,4,7,10-tetraacetic acid (DOTA) or desferrioxamine (DFO) and subsequently radiometalated with (64)Cu and (89)Zr, respectively. The modification strategy is simple and robust, and the resultant radiometalated constructs were obtained in high specific activity (2.7-5.3 mCi/mg). For a given initial stoichiometric ratio of norbornene to antibody, the (64)Cu-DOTA- and (89)Zr-DFO-based probes were shown to be nearly identical in terms of stability, the number of chelates per antibody, and immunoreactivity (>93% in all cases). In vivo PET imaging and acute biodistribution experiments revealed significant, specific uptake of the (64)Cu- and (89)Zr-trastuzumab bioconjugates in HER2-positive BT-474 xenografts, with little background uptake in HER2-negative MDA-MB-468 xenografts or other tissues. This modular system-one in which the divergent point is a single covalently modified antibody stock that can be reacted selectively with various chelators-will allow for both greater versatility and more facile cross-comparisons in the development of antibody-based radiopharmaceuticals.
authors.affiliation: Department of Radiology, Memorial Sloan-Kettering Cancer Center, New York, New York, United States.
authors.forename: Brian M
authors.initials: BM
authors.lastname: Zeglis
authors.forename: Priya
authors.initials: P
authors.lastname: Mohindra
authors.forename: Gabriel I
authors.initials: GI
authors.lastname: Weissmann
authors.forename: Vadim
authors.initials: V
authors.lastname: Divilov
authors.forename: Scott A
authors.initials: SA
authors.lastname: Hilderbrand
authors.forename: Ralph
authors.initials: R
authors.lastname: Weissleder
authors.forename: Jason S
authors.initials: JS
authors.lastname: Lewis
mesh_terms: D000818:Animals; D061067:Antibodies, Monoclonal, Humanized / Q000737:chemistry* / Q000493:pharmacokinetics; D045744:Cell Line, Tumor; D057930:Click Chemistry / Q000379:methods*; D005260:Female; D006573:Heterocyclic Compounds, 1-Ring / Q000737:chemistry* / Q000493:pharmacokinetics; D006801:Humans; D051379:Mice; D008819:Mice, Nude; D009369:Neoplasms / Q000000981:diagnostic imaging; D009636:Norbornanes / Q000737:chemistry* / Q000493:pharmacokinetics; D049268:Positron-Emission Tomography / Q000379:methods*; D019275:Radiopharmaceuticals / Q000737:chemistry* / Q000493:pharmacokinetics; D018719:Erb-b2 Receptor Tyrosine Kinases / Q000276:immunology; D000068878:Trastuzumab
chemical_list: D061067:Antibodies, Monoclonal, Humanized; D006573:Heterocyclic Compounds, 1-Ring; D009636:Norbornanes; D019275:Radiopharmaceuticals; D018719:Erb-b2 Receptor Tyrosine Kinases; D000068878:Trastuzumab; C534588:3-(4-benzylamino)-1,2,4,5-tetrazine; C071349:1,4,7,10-tetraazacyclododecane- 1,4,7,10-tetraacetic acid