DEVELOPMENT OF VESICULAR ACETYLCHOLINE TRANSPORTER IMAGING AGENTS FOR PET

描述（由申请人提供）：本项目的最终目标是开发正电子发射断层扫描（PET）成像剂，以测量人脑中囊泡乙酰胆碱转运蛋白（VAChT）表达的水平。VAChT是胆碱能神经元功能的一种独特的新标记物。这种蛋白质是测量胆碱能神经元损失的关键生物标志物。神经退行性疾病如阿尔茨海默病、唐氏综合征和帕金森病最常见的生理特征之一是中枢胆碱能系统中的胆碱能缺陷。VAChT表达水平的变化也发生在抗精神病药物治疗和药物成瘾治疗期间。迄今为止，没有临床上合适的放射性示踪剂成像VAChT与PET已被确定。因此，对VAChT相对于σ受体具有高亲和力和高选择性以及良好药代动力学特征的放射性示踪剂将对PET社区具有巨大的用途。尽管（+）-[18 F]FBT可用于麻醉恒河猴的VAChT成像，但（+）-FBT对σ受体的高亲和力和从人类受试者脑中的缓慢洗脱速率表明其在临床上不适用于VAChT成像。为了找到可接受的VAChT PET示踪剂，我们在vesamicol结构中的C区和B区之间引入酮基（参见图1）。与此同时，我们利用了苯并维索霉素类似物比维索霉素类似物增加的VAChT结合亲和力。我们将通过引入F、Br、I等不同的取代基来进一步探索C区域，以确定用碳-11或氟-18等PET同位素标记的位置。此外，我们将应用我们的酮策略来开发具有降低的σ亲和力的新FBT类似物。我们的几个新化合物具有高VAChT亲和力和高选择性。其中，（-）-4-氟苯甲酰基苯并维索霉素（（-）-FBBV）是一个有希望的候选物：Ki-VAChT = 4.1 1 ± 0.6 nM，Ki-s1 = 660 ± 1120 nM，Ki-s2 = 320 ± 135 nM。（-）-[18 F]FBBV最初在大鼠和猴中进行了体内评价。最初的结果非常令人鼓舞，但其相对较高的亲脂性可能是PET成像研究中产生高信噪比的挑战。该项目的目标是进行结构-活性关系研究，旨在优化亲脂性与VAChT结合亲和力（Ki值）和选择性。为了实现这一目标，分子模拟和QSAR研究将用于设计新的化合物。将用F-18或C-11放射合成潜在化合物，并在大鼠中进行评价，以确认体内与VAChT的特异性结合。本次申请的具体目的是：1。合成并测量目标化合物的亲和力以鉴定对VAChT具有高亲和力（Ki <10 nM）和对VAChT具有高选择性的配体。符合该标准的外消旋混合物将被拆分为旋光异构体，并测定各异构体的亲和力。2.符合特定目标1和3的碳-11或氟-18标记放射性示踪剂的放射合成。测量放射性示踪剂的亲油性，以确定其log P值是否在~ 1.0范围内 3.0. 4.在大鼠中进行生物分布和脑摄取研究，以选择潜在的放射性示踪剂进行进一步的体内评价。公共卫生相关性神经退行性疾病是美国老年人群中最常见的疾病。确定一种新型的PET示踪剂来成像VAChT，可用于临床测量胆碱能神经元的损失或恢复，对于临床评估胆碱能缺陷疾病的严重程度或针对胆碱能功能障碍患者的治疗反应将非常有用。

DESCRIPTION (provided by applicant): The ultimate goal of this project is to develop Positron Emission Tomography (PET) imaging agents to measure the level of vesicular acetylcholine transporter (VAChT) expression in the human brain. VAChT is a unique and novel marker for cholinergic neuron function. This protein is a key biomarker for measuring the loss of cholinergic neurons. One the most common physiological characteristics of neurodegenerative diseases such as Alzheimer s disease, Down s syndrome, and Parkinson s disease is the cholinergic deficit in the central cholinergic system. Changes in the level of VAChT expression also occur during antipsychotic therapy and treatment of drug addiction. To date, no clinically suitable radiotracer for imaging VAChT with PET has been identified. Therefore, a radiotracer that possesses a high affinity and high selectivity for VAChT versus sigma receptors and good pharmacokinetic characteristics would be of tremendous use to the PET community. Although (+)-[18F]FBT was useful for imaging the VAChT in anesthetized rhesus monkeys, the high affinity of (+)-FBT for sigma receptors and the slow rate of washout from brain in human subjects indicated that it is not clinically useful for imaging the VAChT. In order to find an acceptable VAChT PET tracer, we incorporated a ketone group between C region and B region in the structure of vesamicol (see Figure 1). At the same time, we exploited the increased VAChT binding affinity of the benzovesamicol analogs than the vesamicol analogs. We will further explore the C region by introducing different substituted group such as F, Br, I and others in order to identify a position to be labeled with PET isotopes such as carbon-11 or fluorine-18. Additionally we will apply our ketone strategy to develop new FBT analogs with reduced sigma affinity. Several of our new compounds have both high VAChT affinity and high selectivity. One of them, (-)-4-fluorobenzoyl benzovesamicol ((-)-FBBV) is a promising candidate: Ki-VAChT = 4.1 1 0.6 nM, Ki-s1 = 660 1 120 nM, Ki-s2 = 320 1 35 nM. (-)-[18F]FBBV was initially evaluated in vivo in rats and monkeys. The initial results are very encouraging, but its relatively high lipophilicity could be a challenge for generating a high signal : noise ratio in PET imaging studies. The goal of this project is to conduct a structure-activity relationship study aimed at optimizing lipophilicity versus VAChT binding affinity (Ki value) and selectivity. To achieve the goal, molecular modeling and QSAR studies will be used for designing the new compounds. The potential compounds will be radiosynthesized with F-18 or C-11 and evaluated in rats to confirm the specific binding to VAChT in vivo. The Specific aims of this grant application are: 1. Synthesize and measure the affinities of the target compounds to identify ligands having a high affinity for the VAChT (Ki <10 nM) and high selectivity for VAChT. Racemic mixtures meeting this criterion will be resolved into optical isomers and the individual affinities will be determined. 2. Radiosynthesize of Carbon-11 or Fluorine-18 labeled radiotracers meeting Specific Aim 1 and 3. Measure the lipophilicity of the radiotracers to determine if their log P value is in the range ~ 1.0 3.0. 4. Perform biodistribution and brain uptake studies in rats to select the potential radiotracers for further in vivo evaluation. PUBLIC HEALTH RELEVANCE Neurodegenerative diseases are the most prevalent diseases of aged population in USA. Identifying a novel PET tracer of imaging VAChT that could be used on clinic to measure the loss or recovery of cholinergic neuron will be tremendous useful on clinic assessing the severity of cholinergic deficient diseases or responding to treatments targeting on the cholinergic dysfunction patients.