Purpose

[¹⁸F]FHBG has been used as a positron emission tomography (PET) imaging tracer for the monitoring of herpes simplex virus type 1 thymidine kinase (HSV1-tk), a reporter gene for cell and gene therapy in humans. However, this tracer shows inadequate blood-brain barrier (BBB) penetration and, therefore, would be limited for accurate quantification of reporter gene expression in the brain. Here, we report the synthesis and evaluation of 9-(4-[¹⁸F]fluoro-3-(hydroxymethyl)butyl)-2(phenylthio)-6-oxopurine ([¹⁸F]FHBT) as a new PET tracer for imaging reporter gene expression of HSV1-tk and its mutant HSV1-sr39tk, with the aim of improved BBB penetration.

Procedures

[¹⁸F]FHBT was prepared by using a tosylate precursor and [¹⁸F]KF. The cellular uptake of [¹⁸F]FHBT was performed in HSV1-sr39tk-positive (+) or HSV1-sr39tk-negative (−) MDA-MB-231 breast cancer cells. The specificity of [¹⁸F]FHBT to assess HSV1-sr39tk expression was evaluated by in vitro blocking studies using 1 mM of ganciclovir (GCV). Penetration of [¹⁸F]FHBT and [¹⁸F]FHBG across the BBB was assessed by dynamic PET imaging studies in normal mice.

Results

The tosylate precursor reacted with [¹⁸F]KF using Kryptofix2.2.2 followed by deprotection to give [¹⁸F]FHBT in 10 % radiochemical yield (decay-corrected). The uptake of [¹⁸F]FHBT in HSV1-sr39tk (+) cells was significantly higher than that of HSV1-sr39tk (−) cells. In the presence of GCV (1 mM), the uptake of [¹⁸F]FHBT was significantly decreased, indicating that [¹⁸F]FHBT serves as a selective substrate of HSV1-sr39TK. PET images and time-activity curves of [¹⁸F]FHBT in the brain regions showed similar initial brain uptakes (~ 12.75 min) as [¹⁸F]FHBG (P > 0.855). Slower washout of [¹⁸F]FHBT was observed at the later time points (17.75 − 57.75 min, P > 0.207).

Conclusions

Although [¹⁸F]FHBT showed no statistically significant improvement of BBB permeability compared with [¹⁸F]FHBG, we have demonstrated that the 2-(phenylthio)-6-oxopurine backbone can serve as a novel scaffold for developing HSV1-tk/HSV1-sr39tk reporter gene imaging agents for additional research in the future.

中文翻译：

---

9-(4-[18F] 氟-3-(羟甲基)丁基)-2-(苯硫基)-6-氧嘌呤作为突变型单纯疱疹病毒 1 型胸苷激酶报告基因成像的新型 PET 试剂的合成和表征。

目的

[ ¹⁸ F] FHBG 已被用作正电子发射断层扫描 (PET) 成像示踪剂，用于监测单纯疱疹病毒 1 型胸苷激酶 (HSV1-tk)，这是一种用于人类细胞和基因治疗的报告基因。然而，这种示踪剂显示血脑屏障 (BBB) 渗透不足，因此，对于大脑中报告基因表达的准确量化将受到限制。在这里，我们报告了 9-(4-[ ¹⁸ F] 氟-3-(羟甲基)丁基)-2(苯硫基)-6-氧嘌呤 ([ ¹⁸ F]FHBT) 作为一种新的 PET 成像示踪剂的合成和评估HSV1-tk 及其突变体 HSV1-sr39tk 的报告基因表达，目的是提高 BBB 渗透率。

程序

[ ¹⁸ F]FHBT是通过使用甲苯磺酸盐前体和[ ¹⁸ F]KF制备的。[ ¹⁸ F]FHBT的细胞摄取在HSV1-sr39tk-阳性(+)或HSV1-sr39tk-阴性(-)MDA-MB-231乳腺癌细胞中进行。[ ¹⁸ F]FHBT 评估 HSV1-sr39tk 表达的特异性通过使用 1 mM 更昔洛韦 (GCV) 的体外阻断研究进行评估。[ ¹⁸ F]FHBT 和[ ¹⁸ F]FHBG 穿过血脑屏障的渗透通过动态PET 成像研究在正常小鼠中进行评估。

结果

甲苯磺酸盐前体使用Kryptofix2.2.2与[ ¹⁸ F]KF反应，然后去保护以10％放射化学产率(衰减校正)得到[ ¹⁸ F]FHBT。HSV1-sr39tk (+) 细胞对[ ¹⁸ F]FHBT的摄取显着高于HSV1-sr39tk (-) 细胞的摄取。在GCV (1 mM) 存在下，[ ¹⁸ F]FHBT的摄取显着降低，表明[ ¹⁸ F]FHBT 作为HSV1-sr39TK 的选择性底物。PET图像和[时间-活性曲线¹⁸ F] FHBT在大脑区域中显示出相似的初始脑吸收（〜12.75分钟）为[ ¹⁸ F] FHBG（P  > 0.855）。[ ^{18 的}较慢冲洗在较晚的时间点（17.75 - 57.75 分钟，P > 0.207）观察到 F]FHBT 。

结论

尽管[ ¹⁸ F] FHBT显示出与比较BBB通透性没有统计学显著改善[ ¹⁸ F] FHBG，我们已经表明，2-（苯硫基）-6- oxopurine骨干可以作为新型支架用于开发HSV1-TK / HSV1 -sr39tk 报告基因显像剂用于未来的进一步研究。