Fantoni Enrico Raffaele, Dal Ben Diego, Falzoni Simonetta, Di Virgilio Francesco, Lovestone Simon, Gee Antony
Department of Imaging Sciences and Biomedical Engineering, King's College London, St Thomas' Hospital, 4th floor Lambeth Wing, SE1 7EH, London, UK.
School of Pharmacy, Medicinal Chemistry Unit, University of Camerino, via S. Agostino 1, 62032, Camerino, MC, Italy.
EJNMMI Res. 2017 Dec;7(1):31. doi: 10.1186/s13550-017-0275-2. Epub 2017 Apr 4.
The P2X7 receptor has been shown to play a fundamental role in the initiation and sustenance of the inflammatory cascade. The development of a novel fluorine-18 PET tracer superior and with a longer half-life to those currently available is a promising step towards harnessing the therapeutic and diagnostic potential offered by this target. Inspired by the known antagonist A-804598, the present study outlines the design via molecular docking, synthesis and biological evaluation of the novel P2X7 tracer [F]EFB. The tracer was radiolabelled via a three-step procedure, in vitro binding assessed in P2X7-transfected HEK293 and in B16 cells by calcium influx assays and an initial preclinical evaluation was performed in a lipopolysaccharide (LPS)-injected rat model of neuroinflammation.
The novel tracer [F]EFB was synthesised in 210 min in 3-5% decay-corrected radiochemical yield (DC RCY), >99% radiochemical purity (RCP) and >300 GBq/μmol and fully characterised. Functional assays showed that the compound binds with nM K to human, rat and mouse P2X7 receptors. In vivo, [F]EFB displayed a desirable distribution profile, and while it showed low blood-brain barrier penetration, brain uptake was quantifiable and displayed significantly higher mean longitudinal uptake in inflamed versus control rat CNS regions.
[F]EFB demonstrates strong in vitro affinity to human and rodent P2X7 and limited yet quantifiable BBB penetration. Considering the initial promising in vivo data in an LPS rat model with elevated P2X7 expression, this work constitutes an important step in the development of a radiotracer useful for the diagnosis and monitoring of clinical disorders with associated neuroinflammatory processes.
P2X7受体已被证明在炎症级联反应的启动和维持中起关键作用。开发一种新型的氟-18正电子发射断层显像（PET）示踪剂，其性能优于现有示踪剂且半衰期更长，是挖掘该靶点治疗和诊断潜力的一个有前景的步骤。受已知拮抗剂A-804598的启发，本研究通过分子对接、合成及生物学评价概述了新型P2X7示踪剂[F]EFB的设计。该示踪剂通过三步程序进行放射性标记，通过钙内流测定法在转染P2X7的人胚肾293（HEK293）细胞和B16细胞中评估体外结合情况，并在注射脂多糖（LPS）的大鼠神经炎症模型中进行了初步临床前评价。
新型示踪剂[F]EFB在210分钟内合成，校正衰变后的放射化学产率（DC RCY）为3 - 5%，放射化学纯度（RCP）>99%，比活度>300 GBq/μmol，并进行了全面表征。功能测定表明该化合物与人、大鼠和小鼠的P2X7受体结合的解离常数（Kd）为纳摩尔级别。在体内，[F]EFB显示出理想的分布特征，虽然其血脑屏障穿透率较低，但脑摄取量可定量，且在炎症大鼠中枢神经系统区域的平均纵向摄取量明显高于对照区域。
[F]EFB对人和啮齿动物的P2X7受体具有很强的体外亲和力，且血脑屏障穿透率有限但可定量。鉴于在P2X7表达升高的LPS大鼠模型中获得的初步有前景的体内数据，这项工作是开发一种用于诊断和监测伴有神经炎症过程的临床疾病的放射性示踪剂的重要一步。
