Development of novel imidazole and indane-indole based PET radioligands for imaging of cannabinoid CB2 receptors

大麻素受体2(CB2)在多种病理条件下表达上调，是神经影像研究的重要靶点。尽管靶向CB2受体的重要性很高，各个工作组也做出了相当大的努力，但到目前为止还没有合适的放射性示踪剂的报道。该研究项目的目标是设计一种18F标记的CB2 PET示踪剂，由于CB2受体在健康大脑中的表达水平较低，因此具有亚纳摩尔亲和力和高选择性，是强制性的特征。最近，申请人开发了亚纳摩尔仿射和高选择性的氟化衍生物14a(CB2KI=0.3 nM)，适用于18F-Label的后期掺入。其放射性氟化和体内/体外评估是该项目的第一个主要目标之一。同时，针对一种具有更好的类药物性质的脑靶向衍生物，该分子将进行进一步的修饰。通过进行详细的合成孔径雷达研究，一种具有优越性能的新的氟化衍生物将在体内靶向大脑中的CB2受体。它是否适合成像病变大脑中的CB2受体，最终将在脂多糖诱导的小鼠神经炎症模型中进行研究。该项目的第二部分将侧重于开发基于(R)-N-(6-cyano-2，2-dimethyl-2，3-dihydro-1H-inden-1-yl)-5-fluoro-1H-indole-2-carboxamide((R)-(-)-15)结构的CB2放射性示踪剂。化合物(R)-(-)-15被选为潜在的先导分子，因为它与CB2受体有很高的亲和力。初步结果证实，(-)对映体具有较低的纳米分子CB2亲和力和良好的选择性。它在吲哚5-位含有一种氟，对于18F-标记的亲核SN2引入是失活的。在这个位置引入18F-标记的努力将考虑文献中报道的最新方法，例如使用高价碘离子作为离开基团。考虑到在温和的反应条件下更高的CB2亲和力以及更高的放射性氟化产率，将为最后一步引入氟-18寻找更多的SN2活化位置。在这个研究项目中开发的最好的化合物将进行放射性氟化和生物调查。放射性标记的CB2配体的靶向性和药代动力学将在体外和体内野生型动物中进行评估。通过对最有前景的CB2放射性配基的动态PET研究，将在神经炎症的小鼠模型中评估CB2PET成像用于可视化神经炎症过程的适用性。这项研究提案中阐述的策略有望开发出第一种适合于用PET进行CB2受体活体成像的放射性示踪剂。

Cannabinoid receptor type 2 (CB2) represent a target with increasing importance for neuroimaging due to its upregulation in various pathological conditions. Despite the high importance of targeting the CB2 receptors and considerable efforts of various working-groups, no suitable radiotracer has been reported to date. The goal of the research project is the design of an 18F-labeled CB2 PET-tracer with sub-nanomolar affinity and high selectivity, mandatory features due to the low expression level of the CB2 receptors in the healthy brain. Recently, the applicant developed the sub-nanomolar affine and highly selective fluorinated derivative 14a (CB2Ki = 0.3 nM), suitable for the late stage incorporation of the 18F-label. Its radiofluorination and in vivo/vitro assessment is one of the first, main goals of this project. In parallel, aiming at a derivative with improved drug-like properties for targeting brain, the molecule will be further modified. By performing the thoroughly elaborated SAR study, a new fluorinated derivative with superior properties for the in-vivo targeting the CB2 receeptors in the brain will be died. Its suitability to image CB2 receptors in diseased brain will be finally investigated in a murine mouse model of neuroinflamation induced by LPS. A second part of the project will focus on the development of a CB2 radiotracer based on the structure of (R)-N-(6-cyano-2,2-dimethyl-2,3-dihydro-1H-inden-1-yl)-5-fluoro-1H-indole-2-carboxamide ((R)-(–)-15). Compound (R)-(–)-15 was selected from the literature as potential lead molecule due to its high affinity towards the CB2 receptors. Preliminary results confirmed a low nanomolar CB2 affinity and good selectivity of the (–) enantiomer. It contains a fluorine at the indole 5-position which is inactivated for the nucleophilic SN2 introduction of the 18F-label. Efforts to introduce the 18F-label at this position will consider the newest methods reported in the literature, for example by using hypervalent iodonium illides as leaving group. Aiming at higher CB2 affinity as well as higher radiofluorination yields under mild reactions conditions, more activated positions for SN2 will be investigated for the last step introduction of fluorine-18. The best compounds developed during this research project will be radiofluorinated and biologically investigated. Target specificity and pharmacokinetics of the radiolabeled CB2 ligands will be assessed in vitro and in vivo in wildtype animals. The suitability of CB2 PET-imaging to visualize neuroinflammatory processes by dynamic PET studies of the most promising CB2 radioligand will be evaluated in a murine model of neuroinflammation.The strategy elaborated in this research proposal is promising the development of the first radiotracer suitable for the in vivo imaging of the CB2 receptors with PET.