Phosphodiesterase 7B (PDE7B) inhibition has been considered as a therapeutic target for the treatment of several neurological disorders. Currently, there are no radio-labeled tracers available to determine receptor occupancy (RO) of this target. Developing such a tracer could greatly facilitate the identification of viable PDE7B inhibitors. In the current study, a liquid chromatography tandem mass spectrometry (LC─MS/MS) method was utilized to evaluate the brain distribution of unlabeled tracer candidates following intravenous micro-dosing. This novel approach resulted in an accelerated identification of a potential novel RO tracer for PDE7B. The identified molecule, Compound 30, showed reasonable target-tissue specificity (striatum/cerebellum ratio of 2.2) and suitable uptake (0.25% of the injected dose/g brain tissue) as demonstrated in rats dosed with the unlabeled compound. Compound 30 was subsequently labeled with tritium (3H). In vitro characterization of 3H-Compound 30 demonstrated that this compound possessed a high target affinity with a subnanomolar Kd (0.8 nM) and a Bmax of 58 fmol/mg of protein using rat brain homogenate. Intravenous microdosing of 3H-Compound 30 showed preferential binding in the rat striatum, consistent with the mRNA distribution of PDE7B. In vitro displacement study with other structurally distinct PDE7B target-specific inhibitors using rat brain homogenate indicated that 3H-Compound 30 is an ideal tracer for Ki analysis. This is the first report of a preclinical tracer for PDE7B. With further characterization, Compound 30 may ultimately show the appropriate properties required to be further developed as a PDE7B PET ligand for clinical studies.

中文翻译：

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使用LC─MS/ MS快速鉴定新型磷酸二酯酶7B示踪剂，用于受体占用研究。

磷酸二酯酶7B（PDE7B）抑制已被视为治疗多种神经系统疾病的治疗靶标。当前，没有放射性标记的示踪剂可用于确定该靶标的受体占有率（RO）。开发这样的示踪剂可以大大促进可行的PDE7B抑制剂的鉴定。在本研究中，采用液相色谱串联质谱法（LC─MS/ MS）评估静脉内微量给药后未标记示踪剂候选物的大脑分布。这种新颖的方法导致了对PDE7B潜在的新型RO示踪剂的加速鉴定。鉴定出的分子化合物30显示出合理的靶组织特异性（纹状体/小脑比例为2.2）和适当的摄取（0。如在注射了未标记化合物的大鼠中所证实的，为注射剂量的25％/每克脑组织）。随后用30（3H）标记化合物30。3H-化合物30的体外表征表明，使用大鼠脑匀浆后，该化合物与亚纳摩尔Kd（0.8 nM）具有很高的靶标亲和力，Bmax为58 fmol / mg。3H化合物30的静脉内微剂量显示大鼠纹状体中的优先结合，与PDE7B的mRNA分布一致。使用大鼠脑匀浆对其他结构不同的PDE7B靶标特异性抑制剂进行的体外置换研究表明3H-化合物30是进行Ki分析的理想示踪剂。这是PDE7B临床前示踪剂的首次报道。经过进一步的表征，