Streptavidin in Antibody Pretargeting. 5. Chemical Modification of Recombinant Streptavidin for Labeling with the α-Particle-Emitting Radionuclides ²¹³Bi and ²¹¹At

Abstract

We are investigating the use of recombinant streptavidin (rSAv) as a carrier molecule for the shortlived α-particle emitting radionuclides 213 Bi (t 1/2 = 45.6 min) and 211 At (t 1/2 = 7.21 h) in cancer therapy. To utilize rSAv as a carrier, it must be modified in a manner that permits rapid chelation or bonding with these short-lived radionuclides, and also modified in a manner that diminishes its natural propensity for localization in kidney. Modification for labeling with 213 Bi was accomplished by conjugation of rSAv with the DTPA derivative p-isothiocyanato-benzyl-CHX-A″ (CHX-A″), 3a. Modification for direct labeling with 211 At was accomplished by conjugation of rSAv with an isothiocyanatophenyl derivative of a nido-carborane (nCB), 3b, or an isothiocyanatophenyldPEG ™ /decaborate(2-) derivative, 3c. After conjugation of the chelating or bonding moiety, rSAv was further modified by reaction with an excess (50-100 equivalents) of succinic anhydride. Succinylation of the lysine amines has previously been shown to greatly diminish kidney localization. rSAv modified by conjugation with 3a and succinylated radiolabeled rapidly with 213 Bi (< 5 min), providing a 72% isolated yield. 211 At labeling of modified rSAv was accomplished in aqueous solution using chloramine-T as the oxidant. Astatination of rSAv conjugated with 3b and succinylated occurred very rapidly (<1 min), providing a 50% isolated radiochemical yield. Astatination of rSAv conjugated with 3c and succinylated was also very rapid (<1 min) providing 66-71% isolated radiochemical yields. Astatination of succinylated rSAv, 2a, which did not have conjugated borane cage moieties, resulted in much lower radiolabeling yield (18%). The 213 Bi-or 211 At-labeled modified rSAv preparations were mixed with the corresponding 125 I-labeled rSAv, and dual-label in vivo distributions were obtained in athymic mice. The in vivo data show that 213 Bi-labeled succinylated rSAv [ 213 Bi]6a has tissue concentrations similar to 125 I-labeled modified rSAv [ 125 I] 6b, suggesting that 213 Bi is quite stable towards release from the chelate in vivo. In vivo data also indicate that the 211 At-labeled rSAv conjugated with 3b or 3c and succinylated are stable to in vivo deastatination, whereas succinylated rSAv lacking a boron cage moiety is subject to some deastatination. The modified rSAv conjugated with nido-carborane derivative 3b has a higher retention in many tissues than rSAv without the carborane conjugated. Interestingly, the rSAv conjugated with 3c, which also contains a m-dPEG 12 ™ moiety, has significantly decreased concentrations in blood and other tissues when compared with direct labeled rSAv, suggesting that it may be a good candidate for further study. In conclusion, rSAv that has been modified with CHX-A″ and succinylated (i.e. 5a) may be useful as a carrier of 213 Bi. The encouraging results obtained with the PEGylated decaborate(2-) derivative 3c and succinylated (i.e. 5c) suggests that its further study as a carrier of 211 At in pretargeting prot...

Radiobismuth for Therapy

“…RSAv was conjugated to CHX-A 00 followed by further modification with succinic anhydride since succinylation of the protein lysine amines diminishes kidney localization. The in vivo data showed that 213 Bi-labeled succinylated rSAv has tissue concentrations similar to those of 125 I-labeled modified rSAv which points at its stability towards 213 Bi release [64].…”

Biotinylated and Chelated Poly-L-Lysine as Effector for Pretargeting in Cancer Therapy and Imaging

“…That is, for molecules of the same size and shape, the more negatively charged ones are restricted from filtration to a greater extent [11]. Therefore, charge alterations by succinylation could be an additional tool for optimisation of tumor dose relative to kidney dose [24][25][26].…”

Astatine Radiopharmaceuticals: Prospects and Problems

“…As described in a previous section, to exploit the higher stability of boron-astatine bond strength, Wilbur's group has developed methods for labeling mAbs with 211 At using boron cage pendant groups [151]. Higher in vivo stability with respect to deastatination was obtained by using the so called Venus fly trap complexes; however, long blood residence times and high hepatic uptake was a problem [141].…”