The microtubule-associated protein Tau is a driver of neuronal dysfunction in Alzheimer’s disease and other tauopathies. In this process, Tau initially undergoes subtle changes to its abundance, subcellular localization and a vast array of post-translational modifications including phosphorylation that progressively result in the protein’s somatodendritic accumulation and dysregulation of multiple Tau-dependent cellular processes. Given the various loss- and gain-of-functions of Tau in disease and the brain-wide changes in the proteome that characterize tauopathies, we asked whether targeting Tau would restore the alterations in proteostasis observed in disease. Therefore, by phage display, we generated a novel pan-Tau antibody, RNJ1, that preferentially binds human Tau and neutralizes proteopathic seeding activity in multiple cell lines and benchmarked it against a clinically tested pan-Tau antibody, HJ8.5 (murine version of tilavonemab). We then evaluated both antibodies, alone and in combination, in the K3 tauopathy mouse model, showing reduced Tau pathology and improvements in neuronal function following 14 weekly treatments, without obtaining synergy for the combination. These effects were more pronounced in female mice. To investigate the molecular mechanisms contributing to improvements in neuronal function, we employed quantitative proteomics, phosphoproteomics and kinase prediction analysis to first establish alterations in K3 mice relative to wild-type controls at the proteome level. In female K3 mice, we found 342 differentially abundant proteins, which are predominantly involved in metabolic and microtubule-associated processes, strengthening previously reported findings of defects in several functional domains in multiple tauopathy models. We next asked whether antibody-mediated Tau target engagement indirectly affects levels of deregulated proteins in the K3 model. Importantly, both immunotherapies, in particular RNJ1, induced abundance shifts towards a restoration to wild-type levels (proteostasis). A total of 257 of 342 (∼75%) proteins altered in K3 were closer in abundance to wild-type levels after RNJ1 treatment, and 73% after HJ8.5 treatment. However, the magnitude of these changes was less pronounced than that observed with RNJ1. Furthermore, analysis of the phosphoproteome showed an even stronger restoration effect with RNJ1, with ∼82% of altered phosphopeptides in K3 showing a shift to wild-type levels, and 75% with HJ8.5. Gene set over-representation analysis further confirmed that proteins undergoing restoration are involved in biological pathways affected in K3 mice. Together, our study suggests that a Tau immunotherapy-induced restoration of proteostasis links target engagement and treatment efficacy.

中文翻译：

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蛋白质稳态是 Tau 免疫疗法的基本原则


微管相关蛋白 Tau 是阿尔茨海默病和其他 tau 蛋白病中神经元功能障碍的驱动因素。在这个过程中，Tau 蛋白的丰度最初发生了微妙的变化，亚细胞定位和大量的翻译后修饰，包括磷酸化，这些修饰逐渐导致蛋白质的生长树突状积累和多个 Tau 依赖性细胞过程的失调。鉴于 Tau 在疾病中的各种功能丧失和获得以及表征 tau 病的蛋白质组的全脑变化，我们询问靶向 Tau 是否会恢复在疾病中观察到的蛋白质稳态改变。因此，通过噬菌体展示，我们生成了一种新的 pan-Tau 抗体 RNJ1，该抗体优先结合人 Tau 并中和多种细胞系中的蛋白病接种活性，并将其与临床测试的 pan-Tau 抗体 HJ8.5（tilavonemab 的小鼠版本）进行了基准测试。然后，我们在 K3 tau 蛋白病小鼠模型中评估了单独和联合使用的抗体，显示 14 周治疗后 Tau 病理减少和神经元功能改善，但没有获得联合治疗的协同作用。这些影响在雌性小鼠中更为明显。为了研究有助于神经元功能改善的分子机制，我们采用定量蛋白质组学、磷酸化蛋白质组学和激酶预测分析，首先在蛋白质组水平上确定 K3 小鼠相对于野生型对照的改变。在雌性 K3 小鼠中，我们发现了 342 种差异丰富的蛋白质，这些蛋白质主要参与代谢和微管相关过程，加强了先前报道的多种 tau 蛋白病模型中多个功能域缺陷的发现。 接下来，我们询问抗体介导的 Tau 靶标参与是否间接影响 K3 模型中失调蛋白的水平。重要的是，两种免疫疗法，特别是 RNJ1，都诱导了丰度向恢复到野生型水平（蛋白质稳态）的转变。在 K3 中改变的 342 个蛋白中，共有 257 个 （∼75%） 在 RNJ1 处理后丰度更接近野生型水平，在 HJ8.5 处理后为 73%。然而，这些变化的幅度不如 RNJ1 观察到的那么明显。此外，磷酸化蛋白质组分析显示 RNJ1 具有更强的恢复作用，K3 中 ∼82% 的改变磷酸肽显示转变为野生型水平，而 HJ8.5 中 75% 的磷酸肽。基因集过度代表分析进一步证实，正在恢复的蛋白质参与 K3 小鼠受影响的生物途径。总之，我们的研究表明，Tau 免疫疗法诱导的蛋白质稳态恢复与靶点参与和治疗效果有关。