Development of a Novel PET Tracer for Imaging Microglial Function in Alzheimer's Disease

摘要 阿尔茨海默病(AD)是最常见的痴呆症形式，也是65岁以上成年人死亡的主要原因。 不幸的是，早期诊断和治疗发展的进步因缺乏药物而受到阻碍。 敏感的生物标志物能够检测早期的、功能相关的神经分子变化：尽管数据 全基因组关联研究揭示了与健康的先天免疫功能丧失有关的基因 作为AD的主要危险因素，仍然需要生物标记物来研究神经免疫功能 在这种疾病发生之前和发展过程中。正电子发射断层扫描(PET)是一种非常 灵敏的分子成像模式非常适合于研究这种生物标志物，并建立了对非 对生化过程的侵入性体内审问。现有的神经炎症的PET生物标志物(例如， 转位蛋白18 kDa[TSPO]、CB2、CSF1R、P2X7)存在显著缺陷，包括 阐明中枢神经系统(CNS)中多种细胞类型的功能作用和/或表达。 在中枢神经系统内，腺苷二磷酸受体P2Y12R仅在先天的小胶质细胞上表达。 中枢神经系统的免疫效应细胞，并驱动与小胶质细胞相关的趋化和形态变化 激活。通常被认为是动态平衡小胶质细胞的生物标志物，P2Y12的表达 在急性(如脂多糖挑战)和慢性(如阿尔茨海默病)中均表现为减少 神经炎。晚期AD患者的死后人脑组织显示出 小胶质细胞上的P2Y12R表达减少和几乎完全不表达 淀粉样β斑块。尽管是一个极好的药理靶标，但目前有 无中枢神经系统可穿透的P2Y12R PET示踪剂。为了解决这一未得到满足的需求，我确定了临床候选药物 AZD1283是一种有希望的PET示踪剂。最近，我设计了一种策略，用放射性标记这种分子 C-11(t1/2=20.4min)，合成了[11C]AZD1283，体外在人血浆中表现出很高的稳定性。 在这里，我将比较[11C]AZD1283和TSPO PET示踪剂[11C]DPA-713，以了解它们测量变化的能力 在两种神经炎症小鼠模型中的小胶质细胞(目标1)。此外，我将评估翻译后的 [11C]AZD1283对健康非人灵长类动物的显影和人体体外放射自显影的潜力 AD脑组织(目标2)。通过追求这些目标获得的经验将使我能够直接发展一套技能 适用于未来开发针对免疫系统的示踪剂和疗法的独立研究。这就做 在世界神经炎症PET专家Michelle James博士的指导下进行这项工作， 斯坦福大学病理学教授、医学博士托马斯·蒙廷的额外指导。他们的指导，在 再加上我在斯坦福大学出色的培训环境和可用的资源，这是非常重要的 有助于成功完成这一项目，并最终实现向职业生涯的成功过渡 作为一名创新和有效的独立内科科学家。

SUMMARY Alzheimer’s disease (AD) is the most common form of dementia and a major cause of death in adults over 65. Unfortunately, advancements in early diagnosis and treatment development have been hindered by a paucity of sensitive biomarkers enabling detection of early, functionally relevant, neuromolecular changes: Although data from genome wide association studies have revealed genes relating to a loss of healthy innate immune function as a major risk factor for AD, there remains a need for biomarkers to investigate neuro-immune function preceding and during the development of this disease. Positron Emission Tomography (PET) is an extremely sensitive molecular imaging modality well suited to studying such biomarkers, with established utility for non- invasive in vivo interrogation of biochemical processes. Existing PET biomarkers of neuroinflammation (e.g., the translocator protein 18 kDa [TSPO], CB2, CSF1R, P2X7) suffer from significant drawbacks including a poorly elucidated functional role and/or expression across multiple cell types in the central nervous system (CNS). Within the CNS, the adenosine diphosphate receptor P2Y12R is expressed exclusively on microglia, the innate immune effector cells of the CNS, and drives chemotaxis and morphological changes associated with microglial activation. Generally considered a biomarker of homeostatic microglia, P2Y12 expression has been demonstrated to decrease in both acute (e.g., lipopolysaccharide challenge) and chronic (e.g., AD) neuroinflammation. Postmortem human brain tissue from advanced AD patients demonstrated a global reduction in P2Y12R expression and a near total absence of P2Y12R expression on microglia surrounding amyloid-beta plaques. Despite being an extremely well characterized pharmacological target, there are currently no CNS-penetrable P2Y12R PET tracers. Aiming to address this unmet need, I identified clinical drug candidate AZD1283 as a promising possible PET tracer. Recently, I devised a strategy to radiolabel this molecule with carbon-11 (t1/2=20.4 min), synthesized [11C]AZD1283 and showed it to be highly stable in vitro in human plasma. Here, I will compare [11C]AZD1283 with TSPO PET tracer [11C]DPA-713 for their ability to measure alterations in microglia in two murine models of neuroinflammation (Aim 1). Additionally, I will assess the translational potential of [11C]AZD1283 through imaging healthy non-human primates and in vitro autoradiography of human AD brain tissue (Aim 2). The experience gained by pursuing these aims will allow me to develop a skillset directly applicable to future independent research developing tracers and therapies that target the immune system. I will conduct this work under the mentorship of Michelle James, PhD, a world expert in neuroinflammation PET, with additional mentorship from Thomas Montine, MD, PhD, the chair of Pathology at Stanford. Their mentorship, in conjunction with the excellent training environment and resources available to me at Stanford, are highly conducive to successful completion of this project, and will ultimately enable a successful transition to a career as an innovate and effective independent physician scientist.