Evaluation of 64Cu-Labeled Bifunctional Chelate鈥揃ombesin Conjugates

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• 作者：Samia Ait-Mohand ; Patrick Fournier ; Ve虂ronique Dumulon-Perreault ; Garry E. Kiefer ; Paul Jurek ; Cara L. Ferreira ; Franc抬ois Be虂nard ; Brigitte Gue虂rin
• 刊名：Bioconjugate Chemistry
• 出版年：2011
• 出版时间：August 17, 2011
• 年：2011
• 卷：22
• 期：8
• 页码：1729-1735
• 全文大小：831K
• 年卷期：v.22,no.8(August 17, 2011)
• ISSN：1520-4812

文摘

Several bifunctional chelates (BFCs) were investigated as carriers of ⁶⁴Cu for PET imaging. The most widely used chelator for ⁶⁴Cu labeling of BFCs is DOTA (1,4,7,10-tetraazacyclododecane-N,N鈥?i>,N鈥?i>,N鈥测€测€?tretraacetic acid), even though this complex exhibits only moderate in vivo stability. In this study, we prepared a series of alternative chelator鈥損eptide conjugates labeled with ⁶⁴Cu, measured in vitro receptor binding affinities in human breast cancer T47D cells expressing the gastrin-releasing peptide receptor (GRPR) and compared their in vivo stability in mice. DOTA-, NOTA-(1,4,7-triazacyclononane-1,4,7-triacetic acid), PCTA-(3,6,9,15-tetraazabicyclo[9.3.1]pentadeca-1(15),11,13-triene-3,6,9-triacetic acid), and Oxo-DO3A-(1-oxa-4,7,10-triazacyclododecane-4,7,10-triacetic acid) peptide conjugates were prepared using H₂N-Aoc-[lass="smallcaps">d-Tyr⁶,尾Ala¹¹,Thi¹³,Nle¹⁴]bombesin(6鈥?4) (BBN) as a peptide template. The BBN moiety was selected since it binds with high affinity to the GRPR, which is overexpressed on human breast cancer cells. A convenient synthetic approach for the attachment of aniline-BFC to peptides on solid support is also presented. To facilitate the attachment of the aniline-PCTA and aniline-Oxo-DO3A to the peptide via an amide bond, a succinyl spacer was introduced at the N-terminus of BBN. The partially protected aniline-BFC (p-H₂N-Bn-PCTA(Ot-Bu)₃ or p-H₂N-Bn-DO3A(Ot-Bu)₃) was then coupled to the resulting N-terminal carboxylic acid preactivated with DEPBT/ClHOBt on resin. After cleavage and purification, the peptide-conjugates were labeled with ⁶⁴Cu using [⁶⁴Cu]Cu(OAc)₂ in 0.1 M ammonium acetate buffer at 100 掳C for 15 min. Labeling efficacy was >90% for all peptides; Oxo-DO3A-BBN was incubated an additional 150 min at 100 掳C to achieve this high yield. Specific activities varied from 76 to 101 TBq/mmol. Competition assays on T47D cells showed that all BFC-BBN complexes retained high affinity for the GRPR. All BFC-BBN ⁶⁴Cu-conjugates were stable for over 20 h when incubated at 37 掳C in mouse plasma samples. However, in vivo, only 37% of the ⁶⁴Cu/Oxo-DO3A complex remained intact after 20 h while the ⁶⁴Cu/DOTA-BBN complex was completely demetalated. In contrast, both ⁶⁴Cu/NOTA- and ⁶⁴Cu/PCTA-BBN conjugates remained stable during the 20 h time period. Our results indicate that it is possible to successfully conjugate aniline-BFC with peptide on solid support. Our data also show that ⁶⁴Cu-labeled NOTA- and PCTA-BBN peptide conjugates are promising radiotracers for PET imaging of many human cancers overexpressing the GRP receptor.