title: Preclinical evaluation of anti-HER2 Affibody molecules site-specifically labeled with 111In using a maleimido derivative of NODAGA.
abstract: BACKGROUND
  Affibody molecules have demonstrated potential for radionuclide molecular imaging. The aim of this study was to synthesize and evaluate a maleimido derivative of the 1,4,7-triazacyclononane-1-glutaric acid-4,7-diacetic acid (NODAGA) for site-specific labeling of anti-HER2 Affibody molecule.
  
  
  METHODS
  The maleimidoethylmonoamide NODAGA (MMA-NODAGA) was synthesized and conjugated to Z(HER2:2395) Affibody molecule having a C-terminal cysteine. Labeling efficiency, binding specificity to and cell internalization by HER2-expressing cells of [(111)In-MMA-NODAGA-Cys(61)]-Z(HER2:2395) were studied. Biodistribution of [(111)In-MMA-NODAGA-Cys(61)]-Z(HER2:2395) and [(111)In-MMA-DOTA-Cys(61)]-Z(HER2:2395) was compared in mice.
  
  
  RESULTS
  The affinity of [MMA-NODAGA-Cys(61)]-Z(HER2:2395) binding to HER2 was 67 pM. The (111)In-labeling yield was 99.6%±0.5% after 30 min at 60°C. [(111)In-MMA-NODAGA-Cys(61)]-Z(HER2:2395) bound specifically to HER2-expressing cells in vitro and in vivo. Tumor uptake of [(111)In-MMA-NODAGA-Cys(61)]-Z(HER2:2395) in mice bearing DU-145 xenografts (4.7%±0.8% ID/g) was lower than uptake of [(111)In-MMA-DOTA-Cys(61)]-Z(HER2:2395) (7.5%±1.6% ID/g). However, tumor-to-organ ratios were higher for [(111)In-MMA-NODAGA-Cys(61)]-Z(HER2:2395) due to higher clearance rate from normal tissues.
  
  
  CONCLUSIONS
  MMA-NODAGA is a promising chelator for site-specific labeling of targeting proteins containing unpaired cysteine. Appreciable influence of chelators on targeting properties of Affibody molecules was demonstrated.
authors.affiliation: Unit of Biomedical Radiation Sciences, Rudbeck Laboratory, Uppsala University, Uppsala, Sweden.
authors.forename: Mohamed
authors.initials: M
authors.lastname: Altai
authors.forename: Anna
authors.initials: A
authors.lastname: Perols
authors.forename: Amelie Eriksson
authors.initials: AE
authors.lastname: Karlström
authors.forename: Mattias
authors.initials: M
authors.lastname: Sandström
authors.forename: Frederic
authors.initials: F
authors.lastname: Boschetti
authors.forename: Anna
authors.initials: A
authors.lastname: Orlova
authors.forename: Vladimir
authors.initials: V
authors.lastname: Tolmachev
mesh_terms: D000085:Acetates / Q000737:chemistry*; D000818:Animals; D000911:Antibodies, Monoclonal / Q000737:chemistry* / Q000276:immunology / Q000493:pharmacokinetics; D001665:Binding Sites; D045744:Cell Line, Tumor; D002614:Chelating Agents / Q000737:chemistry; D006573:Heterocyclic Compounds, 1-Ring / Q000737:chemistry*; D006801:Humans; D007205:Indium Radioisotopes / Q000737:chemistry*; D007553:Isotope Labeling / Q000379:methods*; D008297:Male; D008301:Maleimides / Q000737:chemistry*; D051379:Mice; D018719:Erb-b2 Receptor Tyrosine Kinases / Q000276:immunology*; D011993:Recombinant Fusion Proteins / Q000737:chemistry* / Q000276:immunology / Q000493:pharmacokinetics; D013379:Substrate Specificity; D013438:Sulfhydryl Compounds / Q000737:chemistry
chemical_list: D000085:Acetates; D000911:Antibodies, Monoclonal; D002614:Chelating Agents; D006573:Heterocyclic Compounds, 1-Ring; D007205:Indium Radioisotopes; D008301:Maleimides; D018719:Erb-b2 Receptor Tyrosine Kinases; D011993:Recombinant Fusion Proteins; D013438:Sulfhydryl Compounds; C572723:1-(1,3-carboxypropyl)-4,7-carboxymethyl-1,4,7-triazacyclononane; C043592:maleimide