N-Succinimidyl guanidinomethyl iodobenzoate protein radiohalogenation agents: Influence of isomeric substitution on radiolabeling and target cell residualization

Introduction: N-succinimidyl 4-guanidinomethyl-3-[*I]iodobenzoate ([*I]SGMIB) has shown promise for the radioiodination of monoclonal antibodies (mAbs) and other proteins that undergo extensive internalization after receptor binding, enhancing tumor targeting compared to direct electrophilic radioiodination. However, radiochemical yields for [I-131]SGMIB synthesis are low, which we hypothesize is due to steric hindrance from the Boc-protected guanidinomethyl group ortho to the tin moiety. To overcome this, we developed the isomeric compound, N-succinimidyl 3-guanidinomethyl-5-[I-131]iodobenzoate (iso-[I-131]SGMIB) wherein this bulky group was moved from ortho to meta position. Methods: Boc(2)-iso-SGMIB standard and its tin precursor, N-succinimidyl 3-((1,2-bis(tert-butoxycarbonyl) guanidino)methyl)-5-(trimethylstannyl)benzoate (Boc(2)-iso-SGMTB), were synthesized using two disparate routes, and iso-[*I]SGMIB synthesized from the tin precursor. Two HER2-targeted vectors - trastuzumab (Tras) and a nanobody 5 F7 (Nb) - were labeled using iso-[*I]SGMIB and [*I]SGMIB. Paired-label internalization assays in vitro with both proteins, and biodistribution in vivo with trastuzumab, labeled using the two isomeric prosthetic agents were performed. Results: When the reactions were performed under identical conditions, radioiodination yields for the synthesis of Boc(2)-iso-[I-131]SGMIB were significantly higher than those for Boc(2)-[I-131]SGMIB (70.7 +/- 2.0% vs 56.5 +/- 5.5%). With both Nb and trastuzumab, conjugation efficiency also was higher with iso-[I-131]SGMIB than with [I-131]SGMIB (Nb, 33.1 +/- 7.1% vs 28.9 +/- 13.0%; Tras, 45.1 +/- 4.5% vs 34.8 +/- 10.3%); however, the differences were not statistically significant. Internalization assays performed on BT474 cells with 5 F7 Nb indicated similar residualizing capacity over 6 h; however, at 24 h, radioactivity retained intracellularly for iso-[I-131]SGMIB-Nb was lower than for [I-125]SGMIB-Nb (46.4 +/- 1.3% vs 56.5 +/- 2.5%); similar results were obtained using Tras. Likewise, a paired-label biodistribution of Tras labeled using iso-[I-125]SGMIB and [I-131] SGMIB indicated an up to 22% tumor uptake advantage at later time points for [I-131]GMIB-Tras. Conclusion: Given the higher labeling efficiency obtained with iso-SGMIB, this residualizing agent might be of value for use with shorter half-life radiohalogens. (c) 2014 Elsevier Inc. All rights reserved.