Department of Nuclear Medicine, Peking University First Hospital, No. 8 St. Xishiku, Beijing, 100034, China.
Mol Imaging Biol. 2019 Apr;21(2):286-296. doi: 10.1007/s11307-018-1234-7.
Tc-99m- and I-131-labeled arginine-arginine-leucine (RRL) peptides have shown the feasibility of tumor imaging in our previous studies. However, there have been no reports using RRL peptide for positron emission tomography (PET) imaging. In this study, RRL was radiolabeled with Ga-68 under optimized reaction conditions to develop a better specific and effective tumor imaging agent.
RRL was synthesized and conjugated to a bifunctional chelating agent (DOTA-NHS), then radiolabeled with Ga-68. Labeling yield was optimized by varying pH, temperature, and reaction time and the stability was evaluated in human fresh serum. Cellular uptakes of [Ga]DOTA-RRL and FITC-conjugated RRL in HepG2 cells were evaluated using a gamma counter, confocal microscopy, and flow cytometry. PET images and biodistribution were performed in HepG2 tumor-bearing mice after injection of [Ga]DOTA-RRL or [Ga]GaCl at different time points. Further, blocking study was investigated using cold RRL.
The labeling yield of [Ga]DOTA-RRL was 80.6 ± 3.9 % with a pH of 3.5-4.5 at 100 °C for 15 min. The cellular uptake of [Ga]DOTA-RRL in HepG2 cells was significantly higher than that of [Ga]GaCl (P < 0.05). Moreover, the high fluorescent affinity of FITC-conjugated RRL in HepG2 cells was shown using confocal microscopy and flow cytometry. After injection of [Ga]DOTA-RRL in HepG2 tumor-bearing mice, tumor regions exhibited high radioactive accumulation over 120 min and the highest uptake at 30 min. After blocked with cold RRL, HepG2 tumors could not be visualized. [Ga]GaCl was unable to show tumor images at any time point. The biodistribution results confirmed the PET imaging and blocking results.
Our study successfully prepared [Ga]DOTA-RRL with a high labeling yield under the optimized reaction conditions and demonstrated its potential role as a PET imaging agent for tumor-targeted diagnosis.
在我们之前的研究中，Tc-99m-和 I-131 标记的精氨酸-精氨酸-亮氨酸（RRL）肽已显示出肿瘤成像的可行性。然而，目前还没有使用 RRL 肽进行正电子发射断层扫描（PET）成像的报道。在这项研究中，我们在优化的反应条件下用 Ga-68 标记 RRL，以开发出更好的特异性和有效的肿瘤成像剂。
合成 RRL 并与双功能螯合剂（DOTA-NHS）缀合，然后用 Ga-68 标记。通过改变 pH 值、温度和反应时间来优化标记产率，并在人新鲜血清中评估其稳定性。使用伽马计数器、共聚焦显微镜和流式细胞术评估 HepG2 细胞中 [Ga]DOTA-RRL 和 FITC 缀合的 RRL 的细胞摄取。在 HepG2 荷瘤小鼠中注射 [Ga]DOTA-RRL 或 [Ga]GaCl 后，在不同时间点进行 PET 图像和生物分布研究。此外，还通过冷 RRL 进行了阻断研究。
在 pH 值为 3.5-4.5、100°C 下反应 15min 的情况下，[Ga]DOTA-RRL 的标记产率为 80.6±3.9%。HepG2 细胞中 [Ga]DOTA-RRL 的摄取明显高于 [Ga]GaCl（P<0.05）。此外，共聚焦显微镜和流式细胞术显示 FITC 缀合的 RRL 在 HepG2 细胞中具有高荧光亲和力。在 HepG2 荷瘤小鼠中注射 [Ga]DOTA-RRL 后，肿瘤部位在 120min 内表现出高放射性积聚，在 30min 时摄取量最高。用冷 RRL 阻断后，HepG2 肿瘤无法显影。[Ga]GaCl 无法在任何时间点显示肿瘤图像。生物分布结果证实了 PET 成像和阻断结果。
我们在优化的反应条件下成功制备了具有高标记产率的 [Ga]DOTA-RRL，并证明了其作为肿瘤靶向诊断的 PET 成像剂的潜力。
