Biomarkers of Parkinson Disease and Related Disorders

帕金森病（PD）和相关疾病的CSF生物标志物： PD和其他α-突触核蛋白病的诊断是临床的，在早期、轻度病例中往往不确定。我们假设，脑脊液（CSF）水平的神经元代谢产物的儿茶酚胺检测中枢儿茶酚胺缺乏症，从而帮助诊断这些疾病。我们测量了帕金森病和其他两种突触核蛋白病，多系统萎缩（MSA）和纯自主神经功能衰竭（PAF）的CSF儿茶酚，包括多巴胺，去甲肾上腺素，及其主要的神经元代谢物二羟苯乙酸（DOPAC）和二羟苯乙二醇（DHPG）的水平。PD、MSA和PAF组的CSF DOPAC和DHPG水平均低于对照组。PD组DOPAC低于PAF组，PAF组DHPG低于PD组。CSF DOPAC在分离近期发作的帕金森综合征患者与对照组中的敏感性为100%，特异性为89%。从这些发现中，我们得出结论，突触核蛋白病以中枢多巴胺和去甲肾上腺素缺乏的CSF神经化学证据为特征，PD和PAF涉及不同的多巴胺能与去甲肾上腺素能病变，并且CSF DOPAC提供了敏感的临床实验室手段来鉴定PD，甚至在疾病的早期（Goldstein等人，Brain 2012;135：1900-1913）。 我们与华盛顿大学的J. Zhang合作，正在探索PD、MSA和PAF的CSF蛋白质组生物标志物。磷酸化α-突触核蛋白（PS-129）是一种与PD发病机制密切相关的蛋白质，在PD患者和对照组中与总α-突触核蛋白一起沿着测定。CSF PS-129水平与PD严重程度相关，并且当与总突触核蛋白组合时，为临床重叠的帕金森病之间的鉴别诊断提供了高灵敏度和特异性（Wang等人，Science Transl Med 2012;4：121ra20）。 我们患者的额外CSF样本已发送给Zhang博士，以检测CSF PS-129/总α-突触核蛋白的增加，并将CSF PS-129/总α-突触核蛋白与PD、MSA或PAF患者的CSF儿茶酚相关联。 邻苯二酚水平的LCMS与LCED的比较： CSF中DOPAC的水平，尤其是多巴胺的水平非常低。我们比较了液相色谱和三重四极杆质谱（LCMS）与液相色谱和电化学检测（LCED）对人体生物液体（如血浆）中儿茶酚胺的高灵敏度测量。到目前为止，LCED已被证明比LCMS更灵敏和更可靠，用于检测低水平的儿茶酚胺，如多巴胺。 在这项工作的一个重要延伸，我们正在开发的方法来跟踪儿茶酚胺代谢，使用LCMS测量水平的13 C-标记的化合物后，管理的13 C-多巴胺。13 C是一种稳定的非放射性碳同位素。 PD风险的生物标志物： 我们正在测试具有多种PD风险因素的人是否具有大脑或外周中儿茶酚胺神经元损失的生物标志物，如果是，他们是否在随访期间发展为临床PD。通过一个独特的方案特定网站（https：pdrisk.ninds.nih.gov），迄今为止有超过50，000人访问，已确定了250多名具有多种PD统计风险因素的人。我们正在筛选符合条件的人，以确认他们的风险因素，然后让他们接受住院生物标志物检测，包括脑18F-DOPA PET，心脏18F-DOPA PET和CSF儿茶酚。因此，CSF DOPAC在具有多种风险因素的人中降低。 通过死后神经化学进行的生物标志物验证： PD和相关疾病的神经化学或神经影像学生物标志物的验证取决于尸检确认。我们正在测试是否有心脏交感神经去神经的神经影像学证据的患者心肌去甲肾上腺素浓度低。首先，与无路易体疾病的对照组相比，路易体疾病患者的心肌去甲肾上腺素含量降低了90%以上。 11 C-甲基reboxylate PET用于可视化去甲肾上腺素能神经支配： 直到最近，没有放射性配体的神经影像学成功，特异性可视化去甲肾上腺素能神经支配的中枢神经网站。11 C-Methylreboxylate（11 C-MRB）具有这些功能。在人类中，11 C-MRB给药后，使用高分辨率研究断层扫描仪（HRRT）进行正电子发射断层扫描（PET）可以显示蓝斑，即脑桥神经元簇，是大脑中去甲肾上腺素的主要来源。根据NIH临床方案03-N-0004的批准修正案，我们将在NIH临床中心引入11 C-MRB PET扫描，以检测和测量PD和相关疾病中去甲肾上腺素能病变的程度。

CSF Biomarkers of Parkinson Disease (PD) and Related Disorders: The diagnosis of PD and other alpha-synucleinopathies is clinical and often uncertain in early, mild cases. We hypothesized that cerebrospinal fluid (CSF) levels of neuronal metabolites of catecholamines would detect central catecholamine deficiency and thereby help diagnose these disorders. We measured CSF levels of catechols including dopamine, norepinephrine, and their main respective neuronal metabolites dihydroxyphenylacetic acid (DOPAC) and dihydroxyphenylglycol (DHPG) in PD and two other synucleinopathies, multiple system atrophy (MSA) and pure autonomic failure (PAF). The PD, MSA, and PAF groups all had lower CSF DOPAC and DHPG levels than did the controls. DOPAC was lower in PD than in PAF and DHPG lower in PAF than in PD. CSF DOPAC was 100% sensitive at 89% specificity in separating patients with recent onset of Parkinsonism from controls. From these findings we conclude that synucleinopathies feature CSF neurochemical evidence for central dopamine and norepinephrine deficiency, that PD and PAF involve differential dopaminergic vs. noradrenergic lesions, and that CSF DOPAC provides a sensitive clinical laboratory means to identify PD, even early in the disease (Goldstein et al., Brain 2012;135:1900-1913). In collaboration with J. Zhang, of the Univ. of Washington, we are exploring CSF proteomic biomarkers of PD, MSA, and PAF. Phosphorylated alpha-synuclein (PS-129), a protein critically involved in the pathogenesis of PD, was measured along with total alpha-synuclein in patients with PD and controls. CSF PS-129 levels correlated with PD severity and when combined with total synuclein provided high sensitivity and specificity for differential diagnosis among clinically overlapping Parkinsonian disorders (Wang et al., Science Transl Med 2012;4:121ra20). Additional CSF samples from our patients have been sent to Dr. Zhang, to test for increased CSF PS-129/total alpha-synuclein and correlate CSF PS-129/total alpha-synuclein with CSF catechols in the same patients with PD, MSA, or PAF. Comparison of LCMS vs LCED for Levels of Catechols: CSF levels of DOPAC and especially of dopamine are extremely low. We compared liquid chromatography and triple quadrupole mass spectroscopy (LCMS) vs. liquid chromatography and electrochemical detection (LCED) for high-sensitivity measurements of catecholamines in human biological fluids such as plasma. So far LCED has proven more sensitive and more reliable than LCMS for detection of low levels of catecholamines such as dopamine. In an important extension of this work we are developing means to track catecholamine metabolism using LCMS by measuring levels of 13C-labeled compounds after administration of 13C-dopamine. 13C is a stable, non-radioactive isotope of carbon. Biomarkers of Risk of PD: We are testing whether people with multiple risk factors for PD have biomarkers of loss of catecholamine neurons in the brain or periphery and if so whether they develop clinical PD during follow-up. Via a unique Protocol-specific website (https://pdrisk.ninds.nih.gov) visited by more than 50,000 people to date, more than 250 people with multiple statistical risk factors for PD have been identified. We are screening eligible people to confirm their risk factors and then admitting them for inpatient biomarkers testing including brain 18F-DOPA PET, cardiac 18F-dopamine PET, and CSF catechols. Preliminarily, CSF DOPAC is decreased in people with multiple risk factors. Biomarkers Validation by Post-mortem Neurochemistry: Validation of neurochemical or neuroimaging biomarkers of PD and related disorders depends on post-mortem confirmation. We are testing whether patients with neuroimaging evidence of cardiac sympathetic denervation have low myocardial norepinephrine concentrations. Preliminarily, myocardial norepinephrine content is decreased by more than 90% in patients with Lewy body diseases compared to controls without Lewy body diseases. 11C-Methylreboxetine PET to Visualize Noradrenergic Innervation: Until recently, no radioligand for neuroimaging successfully and specifically visualized central neural sites of noradrenergic innervation. 11C-Methylreboxetine (11C-MRB) has these capabilities. In humans, after 11C-MRB administration positron emission tomographic (PET) scanning with a High Resolution Research Tomograph (HRRT) can visualize the locus ceruleus, the pontine cluster of neurons that is the main source of norepinephrine in the brain. Under an approved Amendment to NIH Clinical Protocol 03-N-0004 we are introducing 11C-MRB PET scanning at the NIH Clinical Center, to detect and measure the extent of noradrenergic lesions in PD and related disorders.