Alzheimer’s disease (AD) is characterized by the presence of tau protein inclusions and amyloid beta (Aβ) plaques in the brain, with Aβ peptides generated by cleavage of the amyloid precursor protein (APP) by BACE1 and γ-secretase. We previously described a primary rat neuron assay in which tau inclusions form from endogenous rat tau after seeding cells with insoluble tau isolated from the human AD brain. Here, we used this assay to screen an annotated library of ∼8700 biologically active small molecules for their ability to reduce immuno-stained neuronal tau inclusions. Compounds causing ≥30% inhibition of tau aggregates with <25% loss of DAPI-positive cell nuclei underwent further confirmation testing and assessment of neurotoxicity, and non-neurotoxic hits were subsequently analyzed for inhibitory activity in an orthogonal ELISA that quantified multimeric rat tau species. Of the 173 compounds meeting all criteria, a subset of 55 inhibitors underwent concentration-response testing and 46 elicited a concentration-dependent reduction of neuronal tau inclusions that were distinct from measures of toxicity. Among the confirmed inhibitors of tau pathology were BACE1 inhibitors, several of which, along with γ-secretase inhibitors/modulators, caused a concentration-dependent lowering of neuronal tau inclusions and a reduction of insoluble tau by immunoblotting, although they did not decrease soluble phosphorylated tau species. In conclusion, we have identified a diverse set of small molecules and related targets that reduce neuronal tau inclusions. Notably, these include BACE1 and γ-secretase inhibitors, suggesting that a cleavage product from a shared substrate, such as APP, might affect tau pathology.

阿尔茨海默病 (AD) 的特征是大脑中存在 tau 蛋白内含物和 β 淀粉样蛋白 (Aβ) 斑块，其中 Aβ 肽是由 BACE1 和 γ 分泌酶裂解淀粉样前体蛋白 (APP) 产生的。我们之前描述了一种原代大鼠神经元测定，其中在用从人 AD 大脑中分离出的不溶性 tau 接种细胞后，内源性大鼠 tau 形成 tau 包涵体。在这里，我们使用这种测定法筛选了约 8700 个生物活性小分子的注释库，以了解它们减少免疫染色神经元 tau 内含物的能力。对 tau 聚集体产生 ≥30% 抑制且 DAPI 阳性细胞核损失 <25% 的化合物进行了进一步的确认测试和神经毒性评估，随后在定量多聚体大鼠 tau 物种的正交 ELISA 中分析非神经毒性化合物的抑制活性。在满足所有标准的 173 种化合物中，其中 55 种抑制剂接受了浓度反应测试，其中 46 种引起了神经元 tau 包含物的浓度依赖性减少，这与毒性测量不同。在已证实的 tau 病理学抑制剂中，有 BACE1 抑制剂，其中几种抑制剂与 γ-分泌酶抑制剂/调节剂一起，导致神经元 tau 包涵体浓度依赖性降低，并通过免疫印迹法减少不溶性 tau 蛋白，尽管它们不会降低可溶性磷酸化蛋白。 tau 物种。总之，我们已经确定了一组不同的小分子和相关靶点，可以减少神经元 tau 内含物。值得注意的是，这些抑制剂包括 BACE1 和 γ 分泌酶抑制剂，这表明来自共享底物（例如 APP）的裂解产物可能会影响 tau 病理学。