Data_Sheet_1_GW5074 Increases Microglial Phagocytic Activities: Potential Therapeutic Direction for Alzheimer’s Disease.docx

Microglia, the resident immune cells of the central nervous system (CNS), are responsible for maintaining homeostasis in the brain by clearing debris and are suggested to be inefficient in Alzheimer’s Disease (AD), a progressive neurodegenerative disorder for which there is no disease-modifying drug. Besides pathological approaches, unbiased evidence from genome-wide association studies (GWAS) and gene network analysis implicate genes expressed in microglia that reduce phagocytic ability as susceptibility genes for AD. Thus, a central feature toward AD therapy is to increase the microglial phagocytic activities while maintaining synaptic integrity. Here, we developed a robust unbiased high content screening assay to identify potential therapeutics which can reduce the amyloid-beta (Aβ1–42) load by increasing microglial uptake ability. Our screen identified the small-molecule GW5074, an inhibitor of c-RAF, a serine/threonine kinase, which significantly increased the Aβ1–42 clearance activities in human monocyte-derived microglia-like (MDMi) cells, a microglia culture model that recapitulates many genetic and phenotypic aspects of human microglia. Notably, GW5074 was previously reported to be neuroprotective for cerebellar granule cells and cortical neurons. We found that GW5074 significantly increased the expression of key AD-associated microglial molecules known to modulate phagocytosis: TYROBP, SIRPβ1, and TREM2. Our results demonstrated that GW5074 is a potential therapeutic for AD, by targeting microglia.

小胶质细胞（Microglia）是中枢神经系统（CNS）的驻留免疫细胞，通过清除细胞碎屑维持脑内稳态，且在阿尔茨海默病（AD）中功能低下；阿尔茨海默病是一种尚无疾病修饰治疗药物的进行性神经退行性疾病。除病理学研究手段外，全基因组关联研究（GWAS）与基因网络分析的无偏倚证据表明，小胶质细胞中表达的、可降低吞噬能力的基因为AD易感基因。因此，AD治疗的核心方向之一是在维持突触完整性的前提下，增强小胶质细胞的吞噬活性。本研究建立了一套稳健的无偏倚高内涵筛选实验体系，用于筛选可通过提升小胶质细胞摄取能力以降低β淀粉样蛋白（Aβ1–42）负荷的潜在治疗药物。通过该筛选体系，我们发现小分子化合物GW5074——一种丝氨酸/苏氨酸激酶c-RAF的抑制剂——可显著提升人单核细胞源性小胶质样（MDMi）细胞对Aβ1–42的清除活性；该小胶质细胞培养模型可重现人类小胶质细胞的诸多遗传与表型特征。值得注意的是，此前已有研究报道GW5074对小脑颗粒细胞与皮层神经元具有神经保护作用。我们进一步发现，GW5074可显著上调已知可调控吞噬作用的AD相关关键小胶质细胞分子的表达：TYROBP、SIRPβ1与TREM2。本研究结果证实，GW5074可通过靶向小胶质细胞，成为一款潜在的AD治疗药物。