Ligand discovery for delineating cholesterol homeostasis in the brain

阿尔茨海默病（AD）是一种长期的神经退行性疾病，在美国所有死亡原因中排名第六，其特征是β淀粉样蛋白沉积和神经原纤维缠结。目前，还没有药物可以阻止或逆转疾病的进展，所有创造这种疗法的努力都失败了。最近的研究表明，大脑中胆固醇稳态异常与包括阿尔茨海默病在内的几种神经退行性疾病密切相关。CYP46A1酶将脑胆固醇转化为24s -羟基胆固醇是维持脑胆固醇稳态的主要消除机制。CYP46A1紊乱与阿尔茨海默病的生理病理有关。因此，CYP46A1的药理学调节代表了一种有吸引力的阿尔茨海默病治疗方法。正电子发射断层扫描（PET）能够定量体内生化过程，合适的CYP46A1配体将大大提高我们对AD生理病理条件下CYP46A1介导的胆固醇稳态的理解，否则无法通过体外（破坏性）分析。通过PET定量测定活AD脑组织中CYP46A1的含量，可以评估AD新疗法的分布、靶点参与和剂量占用情况。到目前为止，还没有成功的例子证明CYP46A1成像用于临床应用，这表明我们在体内研究这一靶点的能力显着不足。因此，我们建议开发一种新的PET配体来填补这一空白，作为AD的第一个翻译成像工具。

Alzheimer’s disease (AD) is a long-term neurodegenerative disorder that ranks sixth in the leading cause of all deaths in the United States and features amyloid β protein deposition and neurofibrillary tangles. At present, there are no drugs available to halt or reverse disease progression, and all efforts to create such therapies have failed. Recent studies have demonstrated that abnormalities of cholesterol homeostasis in the brain are strongly associated with several neurodegenerative diseases, including AD. The CYP46A1 enzymatic conversion of brain cholesterol into 24S-hydroxycholesterol is the major elimination mechanism to maintain brain cholesterol homeostasis. Disturbances in CYP46A1 is implicated in the AD physiopathology. Therefore, pharmacological modulation of CYP46A1 represents an attractive AD therapeutic approach. Positron emission tomography (PET) is capable of quantifying biochemical processes in vivo, and a suitable CYP46A1 ligand would substantially improve our understanding of CYP46A1-mediated cholesterol homeostasis under AD physiopathological conditions otherwise inaccessible by ex vivo (destructive) analysis. Quantification of CYP46A1 in living AD brain by PET would provide the assessment of distribution, target engagement and dose occupancy of new AD therapeutics. To date, no successful examples have been demonstrated to image CYP46A1 for clinical use, representing a significant deficiency of our ability to study this target in vivo. Therefore, we propose to develop a novel PET ligand that can fill this void, as the first translational imaging tool for AD. Our ligand [11C]CYP-507 demonstrated the first prototype for imaging towards CYP46A1, but was discontinued due to marginal binding specificity and low brain penetration. In our next generation, we successfully identified a lead molecule, CYP-812, which showed high binding affinity and high selectivity. An 11C-isotopologue of CYP-812 was synthesized and preliminary PET studies confirmed that we have overcome the major obstacles for CYP46A1 ligand development by achieving: 1) reasonable and regional-specific brain uptake; 2) moderate target specificity. Though CYP-812 is a promising lead, further optimizations aimed at higher brain permeability, improved potency and binding specificity with proper brain kinetics are sought for translational cross-species (rodents and nonhuman primates) imaging studies to achieve optimal CYP46A1 quantification for AD research. As specific goals, we will design and prepare a focused library of CYP46A1 modulators amenable for labeling with 11C or 18F, and evaluate their ability to quantify CYP46A1 activity and changes during drug challenge in rodents and nonhuman primates, as well as autoradiography and biological validation in postmortem human brain tissues. The impact of this work is not only to develop the first potent and selective CYP46A1 PET ligand for the study of AD-related biological processes, but also ultimately, via PET imaging validation in higher species, to advance this ligand for potential clinical translation and monitor target response of novel AD neurotherapeutics.