PARK14/Calcium signaling as a novel biomarker for Parkinson disease

帕金森病（PD）是一种无法治愈的神经退行性疾病，其进展缓慢（无临床症状）， 在PD相关运动功能障碍的首次症状发作前数年，绝大多数 的PD病例（约85%）是特发性（idPD），目前，没有特异性生物标志物用于早期诊断 这种毁灭性的疾病。Bolotina博士实验室的最新发现（Zhou等人，Nature 通信，2016）导致识别出PARK 14/PLA 2g 6中先前未知的缺陷- 依赖性Ca 2+信号传导（PLA 2g 6/Ca 2+），其可以在idPD患者的外周细胞中检测到，并且可以 用于开发全新的生物标志物策略。我们还确定，这些缺陷会导致 多巴胺能神经元的进行性死亡和年龄依赖性PD样运动功能障碍的发展， 一种新的小鼠模型，非常接近人类的idPD。重要的是，我们发现人类idPD与 随着PLA 2g 6（L）表达/功能的显著丧失和钙池操纵的Ca 2+内流（SOCE）的损害， 我们可以在idPD患者试验组的原代皮肤成纤维细胞（PSF）中检测到。 在此，我们假设血小板和/或PSF中PLA 2g 6/Ca 2+信号传导的特征缺陷可以用于 作为检测老年人idPD的新生物标志物。我们正在寻求对一项试点计划的支持， 重点验证我们新的基于PLA 2g 6/Ca 2+的生物标志物的特异性，并将测试使用 这些生物标志物在人血小板和/或皮肤成纤维细胞中作为检测idPD的新方法。我们的研究 团队是唯一有资格进行拟议研究的团队：Bolotina博士（PI）拥有独特的知识， 用于在人类受试者的试验组中验证PLA 2g 6/Ca 2+生物标志物的实验平台，以及圣博士 Hilaire（临床合作研究者）目前参与帕金森病进展标志物倡议（PPMI）研究， 并有一个完善的平台，供捐助者招募和样本收集。所有方法均已确立， 由PI和合作研究者发表。初步数据完全支持我们的假设，并证明了我们的可行性。 提议我们建议的具体目标是： 目的1：使用原发性腹膜透析患者的现有样本验证特征性PLA 2g 6/Ca 2+缺陷对idPD的特异性。 皮肤成纤维细胞来自NINDS储存库。PLA 2g 6（L）、钙库操纵的Ca ~（2+）内流和ER的表达和功能 将使用分子、生物化学和成像技术分析Ca 2+水平，并在成纤维细胞中进行比较 来自idPD、家族性PD（LRRK 2或GBA突变）、阿尔茨海默病、亨廷顿病、 和对照组的非神经系统捐献者。所有样本将从NINDS人细胞和数据库中获得。 目的2：测试是否可以在idPD的血小板中检测到PLA 2g 6/Ca 2+信号转导中的特征缺陷 患者活血小板和皮肤成纤维细胞样本将从老年患者的试点组中获得 诊断为早期idPD，并与年龄匹配的对照组非神经系统供体进行比较。 将使用分子、生物化学和成像技术分析PLA 2g 6/Ca 2+信号传导。

Parkinson’s disease (PD) is an incurable neurodegenerative disorder that progresses silently (without clinical manifestations) for years prior to onset of the first symptoms of PD-associated motor dysfunction. The vast majority of PD cases (about 85%) are idiopathic (idPD), and currently, there are no specific biomarkers for early diagnosis of this devastating disease in aging humans. Recent discoveries in Dr. Bolotina’s lab (Zhou et al, Nature Communications, 2016) resulted in identification of the previously unknown defect(s) in PARK14 / PLA2g6- dependent Ca2+ signaling (PLA2g6/Ca2+), which could be detected in peripheral cells from idPD patients, and may be used for development of a brand new biomarker strategy. We also established that such defects lead to progressive demise of dopaminergic neurons and development of age-dependent PD-like motor dysfunction in a new mouse model that closely mimics idPD in humans. Importantly, we found a strong association of human idPD with significant loss of PLA2g6(L) expression/function and impairment of the store-operated Ca2+ entry (SOCE), which we could detect in primary skin fibroblasts (PSFs) from a pilot group of idPD patients. Here we hypothesize that signature defects in PLA2g6/Ca2+ signaling in platelets and/or PSFs could be used as a novel biomarker for detection of idPD in aging humans. We are seeking support for a pilot program that will focus on validation of the specificity of our new PLA2g6/Ca2+-based biomarker, and will test the feasibility of using these biomarkers in human platelets and/or skin fibroblasts as a novel approach for detection of idPD. Our research team is uniquely qualified for proposed studies: Dr. Bolotina (PI) has unique knowledge and established experimental platform for validation of PLA2g6/Ca2+ biomarkers in the pilot groups of human subjects, and Dr. Saint- Hilaire (clinical Co-Investigator) is currently involved in the Parkinson’s Progression Markers Initiative (PPMI) study, and has a well-established platform for donor recruitment and sample collection. All approaches are established and published by PI and Co-Investigator. Preliminary data fully support our hypothesis and demonstrate feasibility of our proposal. Specific Aims of our proposal are: Aim 1: To validate specificity of the signature PLA2g6/Ca2+ defects for idPD using existing samples of primary skin fibroblasts from NINDS repository. Expression and function of PLA2g6(L), store-operated Ca2+ entry and ER Ca2+ levels will be analyzed using molecular, biochemical, and imaging techniques, and compared in fibroblasts from the patients with idPD, familial PD (mutations in LRRK2 or GBA), Alzheimer’s disease, Huntington’s disease, and control non-neurologic donors. All samples will be obtained from NINDS human cell and data repository. Aim 2: To test if the signature defects in PLA2g6/Ca2+ signaling could be detected in platelets from idPD patients. The samples of live platelets and skin fibroblasts will be obtained from a pilot group of aged patients diagnosed with early stages of idPD, and compared with the age-matched group of control non-neurologic donors. PLA2g6/Ca2+ signaling will be analyzed using molecular, biochemical, and imaging techniques.