Molecular Imaging Center, Department of Radiology, University of Southern California, Los Angeles, California 90033, United States.
Bioconjug Chem. 2011 Feb 16;22(2):256-63. doi: 10.1021/bc100388g. Epub 2011 Jan 18.
Robust chelating stability under biological condi-tions is critical for the design of copper-based radiopharmaceuticals. In this study, the stabilities of (64)Cu-DOTA and diamsar (two bifunctional Cu-64 chelators (BFCs)) conjugated DGEA peptides were evaluated. The in vitro stabilities of (64)Cu-DOTA-DGEA, (64)Cu-DOTA-Ahx-DGEA, and (64)Cu-Z-E(diamsar)-Ahx-DGEA were evaluated in PBS. A carboxyl-protected DOTA-DGEA was also synthesized to study the potential inter- and intramolecular interactions between DOTA and the carboxylate groups of DGEA peptide. microPET imaging of (64)Cu-DOTA-DGEA and (64)Cu-Z-E(diamsar)-Ahx-DGEA were performed in PC-3 prostate tumor model to further investigate the in vivo behavior of the tracers. DOTA-DGEA, DOTA-Ahx-DGEA, Z-E(diamsar)-Ahx-DGEA, and protected DOTA-DGEA peptides were readily obtained, and their identities were confirmed by MS. (64)Cu(2+) labeling was performed with high radiochemical yields (>98%) for all tracers after 1 h incubation. Stability experiments revealed that (64)Cu-DOTA-DGEA had unexpectedly high (64)Cu(2+) dissociation when incubated in PBS (>55% free (64)Cu(2+) was observed at 48 h time point). The (64)Cu(2+) dissociation was significantly reduced in the carboxyl-protected (64)Cu-DOTA-DGEA complex but not in the (64)Cu-DOTA-Ahx-DGEA complex, which suggests the presence of competitive binding for (64)Cu(2+) between DOTA and the carboxyl groups of the DGEA peptide. In contrast, no significant (64)Cu(2+) dissociation was observed for (64)Cu-Z-E(diamsar)-Ahx-DGEA in PBS. For microPET imaging, the PC-3 tumors were clearly visualized with both (64)Cu-DOTA-DGEA and (64)Cu-Z-E(diamsar)-Ahx-DGEA tracers. However, (64)Cu-DOTA-DGEA demonstrated 5× higher liver uptake than (64)Cu-Z-E(diamsar)-Ahx-DGEA. This biodistribution variance could be attributed to the chelating stability difference between these two tracers, which correlated well with the PBS stability experiments. In summary, the in vitro and in vivo evaluations of (64)Cu-Z-E(diamsar)-Ahx-DGEA and (64)Cu-DOTA-DGEA have demonstrated the significantly superior Cu-chelation stability for the diamsar derivative compared with the established DOTA chelator. The results also suggest that diamsar may be preferred for Cu chelation especially when multiple carboxylic acid groups are present. Free carboxyl groups may naturally compete with DOTA for (64)Cu(2+) binding and therefore reduce the complex stability.
在生物条件下具有稳健的螯合稳定性对于设计基于铜的放射性药物至关重要。在这项研究中，评估了（64）Cu-DOTA 和二脒沙（两种双功能 Cu-64 螯合剂（BFCs））缀合的 DGEA 肽的稳定性。在 PBS 中评估了（64）Cu-DOTA-DGEA、（64）Cu-DOTA-Ahx-DGEA 和（64）Cu-Z-E（二脒沙）-Ahx-DGEA 的体外稳定性。还合成了一种羧基保护的 DOTA-DGEA，以研究 DOTA 与 DGEA 肽的羧酸盐之间的潜在分子内和分子间相互作用。在 PC-3 前列腺肿瘤模型中进行了（64）Cu-DOTA-DGEA 和（64）Cu-Z-E（二脒沙）-Ahx-DGEA 的 microPET 成像，以进一步研究示踪剂的体内行为。DOTA-DGEA、DOTA-Ahx-DGEA、Z-E（二脒沙）-Ahx-DGEA 和保护的 DOTA-DGEA 肽很容易获得，并通过 MS 确认其身份。所有示踪剂在 1 小时孵育后，（64）Cu（2+）标记均具有>98%的高放射化学产率。稳定性实验表明，（64）Cu-DOTA-DGEA 在 PBS 孵育时具有出人意料的高（64）Cu（2+）解离（在 48 小时时间点观察到超过 55%的游离（64）Cu（2+））。在羧基保护的（64）Cu-DOTA-DGEA 络合物中，（64）Cu（2+）的解离显着减少，但在（64）Cu-DOTA-Ahx-DGEA 络合物中未观察到，这表明 DOTA 和 DGEA 肽的羧基之间存在（64）Cu（2+）的竞争性结合。相比之下，（64）Cu-Z-E（二脒沙）-Ahx-DGEA 在 PBS 中未观察到显着的（64）Cu（2+）解离。对于 microPET 成像，（64）Cu-DOTA-DGEA 和（64）Cu-Z-E（二脒沙）-Ahx-DGEA 示踪剂均可清晰地显示 PC-3 肿瘤。然而，（64）Cu-DOTA-DGEA 在肝脏中的摄取量比（64）Cu-Z-E（二脒沙）-Ahx-DGEA 高 5 倍。这种生物分布差异可能归因于这两种示踪剂之间的螯合稳定性差异，这与 PBS 稳定性实验很好地相关。总之，（64）Cu-Z-E（二脒沙）-Ahx-DGEA 和（64）Cu-DOTA-DGEA 的体外和体内评价表明，与已建立的 DOTA 螯合剂相比，二脒沙衍生物的 Cu 螯合稳定性显着提高。结果还表明，特别是当存在多个羧酸基团时，二脒沙可能更适合用于 Cu 螯合。游离的羧基基团可能会自然与 DOTA 竞争结合（64）Cu（2+），从而降低络合物的稳定性。
