title: Evaluation of a maleimido derivative of NOTA for site-specific labeling of affibody molecules.
abstract: Radionuclide molecular imaging has the potential to improve cancer treatment by selection of patients for targeted therapy. Affibody molecules are a class of small (7 kDa) high-affinity targeting proteins with appreciable potential as molecular imaging probes. The NOTA chelator forms stable complexes with a number of radionuclides suitable for SPECT or PET imaging. A maleimidoethylmonoamide NOTA (MMA-NOTA) has been prepared for site-specific labeling of Affibody molecules having a unique C-terminal cysteine. Coupling of the MMA-NOTA to the anti-HER2 Affibody molecule Z(HER2:2395) resulted in a conjugate with an affinity (dissociation constant) to HER2 of 72 pM. Labeling of [MMA-NOTA-Cys(61)]-Z(HER2:2395) with (111)In gave a yield of >95% after 20 min at 60 °C. In vitro cell tests demonstrated specific binding of [(111)In-MMA-NOTA-Cys(61)]-Z(HER2:2395) to HER2-expressing cell lines. In mice bearing prostate cancer DU-145 xenografts, the tumor uptake of [(111)In-MMA-NOTA-Cys(61)]-Z(HER2:2395) was 8.2 ± 0.9% IA/g and the tumor-to-blood ratio was 31 ± 1 (4 h postinjection). DU-145 xenografts were clearly visualized by a gamma camera. Direct in vivo comparison of [(111)In-MMA-NOTA-Cys(61)]-Z(HER2:2395) and [(111)In-MMA-DOTA-Cys(61)]-Z(HER2:2395) demonstrated that both conjugates provided equal radioactivity uptake in tumors, but the tumor-to-organ ratios were better for [(111)In-MMA-NOTA-Cys(61)]-Z(HER2:2395) due to more efficient clearance from normal tissues. In conclusion, coupling of MMA-NOTA to a cysteine-containing Affibody molecule resulted in a site-specifically labeled conjugate, which retains high affinity, can be efficiently labeled, and allows for high-contrast imaging.
authors: Unit of Biomedical Radiation Sciences, Rudbeck Laboratory, Uppsala University, Uppsala, Sweden. Vladimir.tolmachev@bms.uu.se
authors: Vladimir
authors: V
authors: Tolmachev
authors: Mohamed
authors: M
authors: Altai
authors: Mattias
authors: M
authors: Sandström
authors: Anna
authors: A
authors: Perols
authors: Amelie Eriksson
authors: AE
authors: Karlström
authors: Frederic
authors: F
authors: Boschetti
authors: Anna
authors: A
authors: Orlova
mesh_terms: D000818:Animals; D006571:Heterocyclic Compounds / Q000737:chemistry / Q000493:pharmacokinetics; D006573:Heterocyclic Compounds, 1-Ring; D008297:Male; D008301:Maleimides / Q000737:chemistry / Q000493:pharmacokinetics; D051379:Mice; D008819:Mice, Nude; D057054:Molecular Imaging; D015394:Molecular Structure; D019275:Radiopharmaceuticals / Q000737:chemistry / Q000493:pharmacokinetics; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000493:pharmacokinetics; D013194:Staining and Labeling; D014018:Tissue Distribution; D023041:Xenograft Model Antitumor Assays
chemical_list: D006571:Heterocyclic Compounds; D006573:Heterocyclic Compounds, 1-Ring; D008301:Maleimides; D019275:Radiopharmaceuticals; D011993:Recombinant Fusion Proteins; C048993:1,4,7-triazacyclononane-N,N',N''-triacetic acid