Molecular Probe Dynamics Laboratory, Center for Molecular Imaging Science (CMIS), RIKEN, 6-7-3 Minatojima-minamimachi, Chuo-ku, Kobe, Hyogo, 650-0047, Japan.
Ann Nucl Med. 2013 Aug;27(7):625-39. doi: 10.1007/s12149-013-0728-2. Epub 2013 Apr 20.
Targeted photodynamic therapy (PDT) is necessary for preventing the side effects associated with PDT, such as photosensitivity caused by the distribution of photosensitizers into normal tissues. In the development of targeted PDT agents, a simple evaluation system of in vivo pharmacokinetics, as well as target cell uptake, is absolutely imperative. We hypothesized that (64)Cu chelation with porphyrin photosensitizer-biomacromolecule conjugates may become a simple and versatile labeling strategy for this purpose.
Protoporphyrin IX (PPIX) and a bombesin (BBN) analog, that interacts with the gastrin-released peptide (GRP) receptor, were used as a photosensitizer and tumor-targeting peptide, respectively. Then, a conjugate of PPIX and BBN analog linked via short polyethylene glycol (PPIX-PEG6-BBN analog) was synthesized and used as a targeted PDT agent. In addition, a (64)Cu-chelated PPIX-PEG6-BBN analog was synthesized under optimized reaction conditions. Lastly, cell uptake study and PET image-based pharmacokinetic analyses of the PPIX-PEG6-BBN analog were carried out in a human prostate cancer cell line, PC-3, which highly expresses the GRP receptor, and PC-3 tumor-bearing mice.
It was confirmed that degradation (thought to be based on radiolysis) occurs, and large amounts of (64)Cu-labeling compounds are wasted in the reaction mixture. Interestingly, the addition of ethanol into the reaction mixture provides an effective solution for this problem. As for cell uptake study, the [(64)Cu]PPIX-PEG6-BBN analog demonstrated significantly higher uptake for PC-3 cells than [(64)Cu]PPIX and, in addition, the uptake of [(64)Cu]PPIX-PEG6-BBN analog was significantly inhibited by adding excess cold BBN analog peptide. PET image-based pharmacokinetic evaluation revealed that [(64)Cu]PPIX-PEG6-BBN analog and [(64)Cu]PPIX rapidly accumulate into the liver and kidney, circulate in blood for a long time compared with normal peptides, and distribute at a low level in the tumor. This result suggested that in vivo biodistribution of PPIX-PEG6-BBN analog is mainly dependent on the lipophilicity of PPIX. Ex vivo measurements of radioactivity distribution after PET studies showed that although there was no remarkable difference in the tumor/skin ratio of radioactivity between [(64)Cu]PPIX-PEG6-BBN analog and [(64)Cu]PPIX, the pancreas (an organ that also expresses GRP receptors)/skin ratio was significantly higher in the case of [(64)Cu]PPIX-PEG6-BBN analog.
We report on the successful synthesis of (64)Cu-chelated porphyrin photosensitizers and their tumor-targeting peptide conjugates under conditions in which radiolysis is suppressed. This labeling strategy with porphyrin photosensitizers may be of value for the rapid development of ideal targeted PDT agents.
靶向光动力疗法（PDT）对于预防 PDT 相关的副作用是必要的，例如由于光敏剂分布到正常组织中而引起的光过敏。在靶向 PDT 剂的开发中，体内药代动力学的简单评估系统以及靶细胞摄取是绝对必要的。我们假设（64）Cu 螯合物与卟啉光敏剂-生物大分子缀合物的结合可能成为一种简单而通用的标记策略。
原卟啉 IX（PPIX）和一种与胃泌素释放肽（GRP）受体相互作用的蛙皮素（BBN）类似物分别用作光敏剂和肿瘤靶向肽。然后，通过短的聚乙二醇（PPIX-PEG6-BBN 类似物）合成 PPIX 和 BBN 类似物的缀合物，并将其用作靶向 PDT 剂。此外，在优化的反应条件下合成了（64）Cu 螯合的 PPIX-PEG6-BBN 类似物。最后，在高表达 GRP 受体的人前列腺癌细胞系 PC-3 和 PC-3 荷瘤小鼠中进行了 PPIX-PEG6-BBN 类似物的细胞摄取研究和基于 PET 图像的药代动力学分析。
已证实降解（据推测基于放射分解）会发生，并且大量（64）Cu 标记化合物会在反应混合物中浪费。有趣的是，向反应混合物中添加乙醇为该问题提供了有效的解决方案。至于细胞摄取研究，（64）Cu]PPIX-PEG6-BBN 类似物对 PC-3 细胞的摄取明显高于（64）Cu]PPIX，并且，（64）Cu]PPIX-PEG6-BBN 类似物的摄取通过添加过量冷 BBN 类似物肽可显著抑制。基于 PET 图像的药代动力学评估表明，（64）Cu]PPIX-PEG6-BBN 类似物和（64）Cu]PPIX 迅速积聚到肝脏和肾脏中，与正常肽相比，在血液中循环很长时间，并且在肿瘤中分布水平较低。该结果表明，PPIX-PEG6-BBN 类似物的体内生物分布主要取决于 PPIX 的亲脂性。PET 研究后放射性分布的体外测量表明，尽管（64）Cu]PPIX-PEG6-BBN 类似物与（64）Cu]PPIX 之间在肿瘤/皮肤放射性比之间没有明显差异，但（64）Cu]PPIX-PEG6-BBN 类似物的胰腺（也表达 GRP 受体）/皮肤比明显更高。
我们报告了在抑制放射分解的条件下成功合成了（64）Cu 螯合卟啉光敏剂及其肿瘤靶向肽缀合物。这种带有卟啉光敏剂的标记策略对于快速开发理想的靶向 PDT 剂可能具有价值。
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