Immunomedics, Inc., Morris Plains, New Jersey, USA.
Bioconjug Chem. 2012 Mar 21;23(3):538-47. doi: 10.1021/bc200608e. Epub 2012 Feb 10.
Radiolabeling compounds with positron-emitting radionuclides often involves a time-consuming, customized process. Herein, we report a simple lyophilized kit formulation for labeling peptides with (18)F, based on the aluminum-fluoride procedure. The prototype kit contains IMP485, a NODA (1,4,7-triazacyclononane-1,4-diacetate)-MPAA (methyl phenylacetic acid)-di-HSG (histamine-succinyl-glycine) hapten-peptide, [NODA-MPAA-D-Lys(HSG)-D-Tyr-D-Lys(HSG)-NH(2)], used for pretargeting, but we also examined a similar kit formulation for a somatostatin-binding peptide [IMP466, NOTA-D-Phe-Cys-Phe-D-Trp-Lys-Thr-Cys-Throl] bearing a NOTA ligand to determine if the benefits of using a kit can be extended to other AlF-binding peptides. The NODA-MPAA ligand forms a single stable complex with (AlF)(2+) in high yields. In order to establish suitable conditions for a facile kit, the formulation was optimized for pH, peptide to Al(3+) ratio, bulking agent, radioprotectant, and the buffer. For optimal labeling, the kit was reconstituted with an aqueous solution of (18)F(-) and ethanol (1:1), heated at 100-110 °C for 15 min, and then simply and rapidly purified using one of two equally effective solid-phase extraction (SPE) methods. Al(18)F-IMP485 was isolated as a single isomer complex, in high yield (45-97%) and high specific activity (up to 223 GBq/μmol), within 20 min. The labeled product was stable in human serum at 37 °C for 4 h and in vivo, urine samples showed the intact product was eliminated. Tumor targeting of the Al(18)F-IMP485 in nude mice bearing human colon cancer xenografts pretargeted with an anti-CEACAM5 bispecific antibody showed very low uptake (0.06% ± 0.02 ID/g) in bone, further illustrating its stability. At 1 h, pretargeted animals had high Al(18)F-IMP485 tumor uptake (28.1% ± 4.5 ID/g), with ratios of 9 ± 4, 123 ± 38, 110 ± 43, and 120 ± 108 for kidney, liver, blood and bone, respectively. Tumor uptake remained high at 3 h postinjection, with increased tumor/nontumor ratios. The NOTA-somatostatin-binding peptide also was fluorinated with good yield and high specific activity in the same kit formulation. However, yields were somewhat lower than those achieved with IMP485 containing the NODA-MPAA ligand, likely reflecting this ligand's superior binding properties over the simple NOTA. These studies indicate that (18)F-labeled peptides can be reproducibly prepared as stable Al-F complexes with good radiochemical yield and high specific activity using a simple, one-step, lyophilized kit followed by a rapid purification by SPE that provides the (18)F-peptide ready for patient injection within 30 min.
放射性标记化合物与正电子发射放射性核素通常涉及耗时且定制化的过程。在此，我们报告了一种基于铝-氟化物程序，用于用 (18)F 标记肽的简单冻干试剂盒配方。该原型试剂盒包含 IMP485，一种 NODA（1,4,7-三氮杂环壬烷-1,4-二乙酸）-MPAA（甲基苯乙酸）-二-HSG（组氨酸-琥珀酰-甘氨酸）半抗原-肽，[NODA-MPAA-D-Lys（HSG）-D-Tyr-D-Lys（HSG）-NH（2）]，用于前靶向，但我们还检查了类似的试剂盒配方用于结合生长抑素的肽 [IMP466，NOTA-D-Phe-Cys-Phe-D-Trp-Lys-Thr-Cys-Throl]，带有 NOTA 配体，以确定使用试剂盒的好处是否可以扩展到其他 AlF 结合肽。NODA-MPAA 配体以高产率形成与 (AlF)（2+）的单一稳定配合物。为了为简便的试剂盒建立合适的条件，对配方进行了优化，以获得最佳的 pH 值、肽与 Al（3+）的比例、膨胀剂、辐射保护剂和缓冲液。为了获得最佳的标记效果，试剂盒用 (18)F（-）和乙醇（1:1）的水溶液重新配制，在 100-110°C 下加热 15 分钟，然后使用两种同样有效的固相萃取 (SPE) 方法之一进行简单快速的纯化。Al（18）F-IMP485 作为单一异构体配合物分离，产率高（45-97%），比活度高（高达 223GBq/μmol），在 20 分钟内完成。标记产物在 37°C 下在人血清中 4 小时稳定，体内，尿液样本显示完整的产物被消除。在携带人结肠癌异种移植物的裸鼠中进行的 Al（18）F-IMP485 肿瘤靶向实验，使用抗 CEACAM5 双特异性抗体进行前靶向，结果显示在骨骼中的摄取非常低（0.06%±0.02 ID/g），进一步证明了其稳定性。在 1 小时时，前靶向动物的 Al（18）F-IMP485 肿瘤摄取率（28.1%±4.5 ID/g）较高，肾脏、肝脏、血液和骨骼的摄取率分别为 9±4、123±38、110±43 和 120±108。注射后 3 小时肿瘤摄取仍保持较高水平，肿瘤/非肿瘤比值增加。NOTA-生长抑素结合肽也以相同的试剂盒配方以良好的产率和高比活度进行了放射性标记。然而，产率略低于含有 NODA-MPAA 配体的 IMP485 获得的产率，可能反映了这种配体的结合性能优于简单的 NOTA。这些研究表明，（18）F 标记的肽可以使用简单的一步冻干试剂盒以稳定的 Al-F 配合物形式重现性地制备，该试剂盒随后通过 SPE 进行快速纯化，在 30 分钟内为患者提供（18）F-肽。
