SIB-DOTA: A trifunctional prosthetic group potentially amenable for multi-modal labeling that enhances tumor uptake of internalizing monoclonal antibodies

Abstract

A major drawback of internalizing monoclonal antibodies (mAbs) radioiodinated with direct electrophilic approaches is that tumor retention of radioactivity is compromised by the rapid washout of iodo-tyrosine, the primary labeled catabolite for mAbs labeled via this strategy. In our continuing efforts to develop more versatile residualizing labels that could overcome this problem, we have designed SIB-DOTA, a prosthetic labeling template that combines the features of the prototypical, dehalogenation resistant N-succinimidyl 3-iodobenzoate (SIB) with DOTA, a useful macrocyclic chelator for labeling with radiometals. Herein we describe the synthesis of the unlabeled standard of this prosthetic moiety, its protected tin precursor, and radioiodinated SIB-DOTA. An anti-EGFRvIII-reactive mAb, L8A4 was radiolabeled with [131I]SIB-DOTA in 27.1 ± 6.2% (n = 2) conjugation yields and its targeting properties to the same mAb labeled with [125I]SGMIB both in vitro and in vivo using U87MG·ΔEGFR cells and xenografts were compared. In vitro paired-label internalization assays showed that the intracellular radioactivity from [131I]SIB-DOTA-L8A4 was 21.4 ± 0.5% and 26.2 ± 1.1% of initially bound radioactivity at 16 and 24 h, respectively. In comparison, these values for [125I]SGMIB-L8A4 were 16.7 ± 0.5% and 14.9 ± 1.1%. Similarly, the SIB-DOTA prosthetic group provided better tumor targeting in vivo than SGMIB over 8 d period. These results suggest that SIB-DOTA warrants further evaluation as a residualizing agent for labeling internalizing mAbs including those targeted to EGFRvIII.

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

“…Due to the solubility of tetrabutyl ammonium salts in organic solvents, acidification can be avoided and the salt can be used directly for re-esterification with N -hydroxysuccinimide. Dimethyl 5-iodoisophthalate was hydrolyzed as reported [27], and the resultant 5-iodoisophthalic acid was converted to its mono trimethylsilylethyl ester 12 . Formation of the diester was unavoidable resulting in only modest yields for the desired monoester (25%).…”

“…The design hypothesis has been to utilize acylation agents that will result in the generation of labeled catabolites that would be charged at lysosomal pH and therefore will not be able to cross the lysosomal and cell membranes [10-14]. The most successful residualizing agent developed to date based on this premise is the guanidine-moiety-containing acylation agent, N -succinimidyl 4-guanidinomethyl-3-[ 131 I]iodobenzoate ([ 131 I]SGMIB) [15, 16], which likely reflects the fact that guanidine has a pKa of ∼13, and should remain exclusively positively charged at lysosomal pH.…”

D-amino acid peptide residualizing agents bearing N-hydroxysuccinimido- and maleimido-functional groups and their application for trastuzumab radioiodination

“…A Mal- d -GEEEK-IB standard and the corresponding tin precursor N ∊ -(3-(tri- n -butylstannyl)benzoyl)-Lys 5 - N α -maleimido-Gly- d -GEEEK (Mal- d -GEEEK-TB) were synthesized as reported before [15]. Bis- N -hydroxysuccinimidyl 5-iodo-isophthalate and bis- N -hydroxysuccinimidyl 5-(tri- n -butylstannyl)isophthalate were prepared as reported [17]. Fmoc-Gly-Wang resin and Fmoc- d -Glu-(O t Bu)-OH were obtained from Novabiochem (West Chester, PA).…”

Radiolabeling Strategies for Tumor-Targeting Proteinaceous Drugs

“…This chelating agent allows for the radiolabeling of a halogen and a metal within the same molecule. [131] The potential of delivering a mixture of radiolabels, each with their own unique properties, may be of great benefit in the future of RIT.…”

“…The direct radioiodination of a protein is a key method for the synthesis of tumor-targeting radiopharmaceuticals, like 131 I-labeled rituximab [50] (Table 3). Generally there are two basic approaches of protein radioiodination.…”