title: Labeling internalizing anti-epidermal growth factor receptor variant III monoclonal antibody with (177)Lu: in vitro comparison of acyclic and macrocyclic ligands.
abstract: BACKGROUND
  The monoclonal antibody (mAb) L8A4, reactive with the epidermal growth factor receptor variant III (EGFRvIII), internalizes rapidly in glioma cells after receptor binding. Combining this tumor-specific mAb with the low-energy beta-emitter (177)Lu would be an attractive approach for brain tumor radioimmunotherapy, provided that trapping of the radionuclide in tumor cells after mAb intracellular processing could be maximized.
  
  
  METHODS
  L8A4 mAb was labeled with (177)Lu using the acyclic ligands [(R)-2-amino-3-(4-isothiocyanatophenyl)propyl]-trans-(S,S)-cyclohexane-1,2-diamine-pentaacetic acid (CHX-A''-DTPA), 2-(4-isothiocyanatobenzyl)-diethylenetriaminepenta-acetic acid (pSCN-Bz-DTPA) and 2-(4-isothiocyanatobenzyl)-6-methyldiethylenetriaminepentaacetic acid (1B4M-DTPA), and the macrocyclic ligands S-2-(4-isothiocyanatobenzyl)-1,4,7,10-tetraazacyclododecane-tetraacetic acid (C-DOTA) and alpha-(5-isothiocyanato-2-methoxyphenyl)-1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid (MeO-DOTA). Paired-label internalization and cellular processing assays were performed on EGFRvIII-expressing U87.DeltaEGFR glioma cells over 24 h to directly compare (177)Lu-labeled L8A4 to L8A4 labeled with (125)I using either iodogen or N-succinimidyl 4-guanidinomethyl-3-[(125)I]iodobenzoate ([(125)I]SGMIB). In order to facilitate comparison of labeling methods, the primary parameter evaluated was the ratio of (177)Lu to (125)I activity retained in U87.DeltaEGFR cells.
  
  
  RESULTS
  All chelates demonstrated higher retention of internalized activity compared with mAb labeled using iodogen, with (177)Lu/(125)I ratios of >20 observed for the three DTPA chelates at 24 h. When compared to L8A4 labeled using SGMIB, except for MeO-DOTA, internalized activity for (125)I was higher than (177)Lu from 1-8 h with the opposite behavior observed thereafter. At 24 h, (177)Lu/(125)I ratios were between 1.5 and 3, with higher values observed for the three DTPA chelates.
  
  
  CONCLUSIONS
  The nature of the chelate used to label this internalizing mAb with (177)Lu influenced intracellular retention in vitro, although at early time points, only MeO-DOTA provided more favorable results than radioiodination of the mAb via SGMIB.
authors.affiliation: Department of Radiology, Duke University Medical Center, Durham, NC 27710, USA.
authors.forename: Marc
authors.initials: M
authors.lastname: Hens
authors.forename: Ganesan
authors.initials: G
authors.lastname: Vaidyanathan
authors.forename: Phil
authors.initials: P
authors.lastname: Welsh
authors.forename: Michael R
authors.initials: MR
authors.lastname: Zalutsky
mesh_terms: D000818:Animals; D000911:Antibodies, Monoclonal / Q000737:chemistry* / Q000493:pharmacokinetics / Q000627:therapeutic use; D001932:Brain Neoplasms / Q000532:radiotherapy; D045744:Cell Line, Tumor; D002614:Chelating Agents / Q000737:chemistry*; D002851:Chromatography, High Pressure Liquid; D066246:ErbB Receptors / Q000276:immunology*; D005910:Glioma / Q000532:radiotherapy; D007553:Isotope Labeling / Q000379:methods*; D008024:Ligands; D008187:Lutetium / Q000493:pharmacokinetics; D047028:Macrocyclic Compounds / Q000737:chemistry*; D051379:Mice; D016499:Radioimmunotherapy; D011868:Radioisotopes / Q000737:chemistry* / Q000493:pharmacokinetics / Q000627:therapeutic use; D019275:Radiopharmaceuticals / Q000737:chemistry* / Q000493:pharmacokinetics / Q000627:therapeutic use
chemical_list: D000911:Antibodies, Monoclonal; D002614:Chelating Agents; D008024:Ligands; D047028:Macrocyclic Compounds; D011868:Radioisotopes; D019275:Radiopharmaceuticals; D008187:Lutetium; D066246:ErbB Receptors