title: Nepsilon-(3-[*I]Iodobenzoyl)-Lys5-Nalpha-maleimido-Gly1-GEEEK ([*I]IB-Mal-D-GEEEK): a radioiodinated prosthetic group containing negatively charged D-glutamates for labeling internalizing monoclonal antibodies.
abstract: Novel methods are needed for the radiohalogenation of cell-internalizing proteins and peptides because rapid loss of label occurs after lysosomal processing when these molecules are labeled using conventional radioiodination methodologies. We have developed a radiolabeled prosthetic group that contains multiple negatively charged D-amino acids to facilitate trapping of the radioactivity in the cell after proteolysis of the labeled protein. N(epsilon)-(3-[(125)I]iodobenzoyl)-Lys(5)-N(alpha)-maleimido-Gly(1)-GEEEK ([(125)I]IB-Mal-D-GEEEK) was synthesized via iododestannylation in 90.3 +/- 3.9% radiochemical yields. This radioiodinated agent was conjugated to iminothiolane-treated L8A4, an anti-epidermal growth factor receptor variant III (EGFRvIII) specific monoclonal antibody (mAb) in 54.3 +/- 17.7% conjugation yields. In vitro assays with the EGFRvIII-expressing U87MGDeltaEGFR glioma cell line demonstrated that the internalized radioactivity for the [(125)I]IB-Mal-D-GEEEK-L8A4 conjugate increased from 14.1% at 1 h to 44.7% at 24 h and was about 15-fold higher than that of directly radioiodinated L8A4 at 24 h. A commensurately increased tumor uptake in vivo in athymic mice bearing subcutaneous U87MGDeltaEGFR xenografts (52.6 +/- 14.3% injected dose per gram versus 17.4 +/- 3.5% ID/g at 72 h) also was observed. These results suggest that [(125)I]IB-Mal-d-GEEEK is a promising reagent for the radioiodination of internalizing mAbs.
authors.affiliation: Department of Radiology, Duke University Medical Center, Durham, North Carolina 27710, USA. ganesan.v@duke.edu
authors.forename: Ganesan
authors.initials: G
authors.lastname: Vaidyanathan
authors.forename: Kevin L
authors.initials: KL
authors.lastname: Alston
authors.forename: Darrel D
authors.initials: DD
authors.lastname: Bigner
authors.forename: Michael R
authors.initials: MR
authors.lastname: Zalutsky
mesh_terms: D000818:Animals; D000911:Antibodies, Monoclonal / Q000737:chemistry*; D001932:Brain Neoplasms / Q000737:chemistry; D045744:Cell Line, Tumor; D005910:Glioma / Q000737:chemistry; D018698:Glutamic Acid / Q000737:chemistry*; D007457:Iodine Radioisotopes / Q000737:chemistry*; D007462:Iodobenzenes / Q000737:chemistry* / Q000493:pharmacokinetics; D051379:Mice; D009842:Oligopeptides / Q000737:chemistry* / Q000493:pharmacokinetics; D014018:Tissue Distribution
chemical_list: D000911:Antibodies, Monoclonal; D007457:Iodine Radioisotopes; D007462:Iodobenzenes; C545084:N(epsilon)-(3-iodobenzoyl)-Lys(5)-Nalpha-maleimido-Gly(1)-glycyl-glutamyl-glutamyl-gluamyl-lysine; D009842:Oligopeptides; D018698:Glutamic Acid