Reagents and Methods for PET Using Bispecific Antibody Pretargeting and 68Ga-Radiolabeled Bivalent Hapten-Peptide-Chelate Conjugates

Journal of Nuclear Medicine

Chien-Hsing Chang¹  William J. McBride¹  Ivan D. Horak¹  Hans J. Hansen¹  David M. Goldenberg¹  German R. Tejada¹  Agatha Sheerin¹  Edmund A. Rossi¹  Habibe Karacay¹  Gary L. Griffiths¹  Robert M. Sharkey¹ 

关键词: ⁶⁸Ga;    ⁶⁷Ga;    PET;    bispecific antibodies;    2-step pretargeting;   

DOI  :  

学科分类：医学（综合）

来源: Society of Nuclear Medicine

【 摘 要 】

The aim of this work was to develop reagents and methods potentially useful in PET, using 68Ga in a 2-step pretargeting protocol. Methods: We prepared bispecific antibodies (bsAbs) for disease-specific targeting of carcinoembryonic antigen-positive cells and recognition of later-administered bivalent hapten-peptide conjugates. The secondary antibody arm (antibody 679) recognizes a histaminyl-succinyl-glycine (HSG) structural subunit. The bsAbs were prepared as Fab′ × Fab′ conjugates using chemical cross-linking methods and as bispecific diabodies using recombinant DNA technologies. A HSG-bivalent hapten conjugate bearing the macrocyclic ring chelating agent 1,4,7,10-tetraazacyclododecane-N,N′,N″,N‴-tetraacetic acid (DOTA) was designed to be readily radiolabeled with 68Ga taken directly from a 68Ge/68Ga generator system. Reagents were tested in vitro and, then, for their targeting properties in a preclinical animal model of human cancer. Results: A chemically cross-linked hMN-14 × 679 F(ab′)2 and a fully humanized bispecific diabody construct (BS1.5H), expressed in Escherichia coli, were prepared for this work. We synthesized the bivalent peptide termed IMP 241 [DOTA-Phe-Lys(HSG)-d-Tyr-Lys(HSG)-NH2] and labeled it with 68Ga and 67Ga at temperatures from 45°C to 100°C, over times of 15 min to 1 h, establishing 15 min at 95°C as a useful condition for 68Ga labeling. When we formulated the IMP 241 bivalent hapten-peptide with ammonium acetate buffer at pH 4–5 and eluted the 68Ga from the generator directly into the peptide solution, we achieved an almost quantitative incorporation of the 68Ga into IMP 241, as analyzed by size-exclusion high-performance liquid chromatography, after mixing the complex with the 679 antibody. For in vivo studies we used 67Ga-IMP 241 as a surrogate for 68Ga-IMP 241, in view of the short, 68-min half-life of the 68Ga nuclide. The 67Ga-IMP 241 was successfully pretargeted to human colon tumor xenografts in athymic mice with both the chemical and the diabody bispecific proteins. High tumor-to-normal tissue ratios for 67Ga uptake were found for all tissues at 1 to 6 h after injection of 67Ga-IMP 241. When using the BS1.5H diabody for pretargeting, tumor-to-blood, tumor-to-liver, and tumor-to-lung ratios of 67Ga-IMP 241 at 1 and 3 h after injection were 41:1 and 137:1, 51:1 and 106:1, and 16:1 and 46:1, respectively. Conclusion: The general approach described, along with the new compositions and the labeling methods we have developed, may eventually allow for use of 68Ga-labeled specific targeting agents in a routine clinical PET application.

【 授权许可】

Unknown