Adapting Secretory Proteostasis through Pharmacologic IRE1 Activation

蛋白质内稳态(或蛋白质稳态)的失衡与病因学上的- 各种疾病，包括糖尿病、系统性淀粉样变性疾病、心脏病和衰老相关疾病 神经退行性疾病，如阿尔茨海默病(AD)。目前还没有针对这些疾病的治疗方法 疾病，促使人们努力了解疾病的发病机制，并开发新的方法来 缓解相关的致病蛋白平衡失衡。因为细胞蛋白质组的整体完整性是 活性和功能的中心方面，细胞进化出一个蛋白质平衡网络，由折叠和 降解因子，以确保新生或错误折叠的多肽正确折叠，或在以下情况下促进它们的降解 折叠不能实现。内质网(ER)未折叠蛋白反应(UPR)在 在维持细胞内和细胞外蛋白质平衡方面的作用，因为内质网环境是 分泌蛋白质的折叠。重要的是，许多蛋白质与蛋白平衡相关的疾病有关 包括AD在内的蛋白通过分泌途径运输，因此与内质网蛋白稳态相互作用 环境。因此，普遍定期审议是操纵不稳定、与疾病有关的疾病水平的一个有吸引力的目标。 蛋白质。这种反应通常是在内质网错误折叠蛋白负荷增加的情况下激活的 管腔(即，内质网应激)，这是一种信号，由肌醇需要的内质网膜蛋白感知和转导 酶1(IRE1)、蛋白激酶样内质网蛋白激酶(PERK)和激活转录 因子6(ATF6)，进而诱导蛋白抑制转录程序，以减轻未折叠的蛋白质负荷。 本研究项目特别关注普遍定期审议的IRE1信令臂，该信令已被广泛 使用遗传和化学遗传方法研究证明激活的治疗潜力 对于多种病因多样的疾病来说，这是一条重要的途径。虽然这些方法具有变革性 为了研究IRE1信号，目前可用于药理激活这一途径的化合物有 受到其固有的混杂和细胞毒性的限制。对于这个研究项目，我使用了一种基于荧光素酶的 结合转录和蛋白质组学研究的高通量筛选方法以鉴定新的 IRE1信号轴的小分子激活剂，具有明确的作用机制(目标1)。通过这些 经过努力，我优先选择化合物474作为IRE1的一种有前途的一流、无毒和特异的激活剂 多种细胞培养模型中的信号轴。为了刻画其治疗潜力， 药物激活IRE1，我将把这个化合物和其他化合物应用于AD的细胞模型以研究其作用 失稳淀粉样前体蛋白(APP)和β-淀粉样蛋白(Abβ)多肽水平及其下游的研究 相关细胞系细胞健康的生理学读数(目标2)。这些努力将展示其潜力 用于药物激活IRE1以缓解AD和AD所涉及的与疾病相关的蛋白平衡失衡 其他疾病。

Imbalances in protein homeostasis (or proteostasis) are implicated in the onset and pathogenesis of etiologically- diverse diseases including diabetes, systemic amyloid disease, heart disease, and aging-related neurodegenerative disorders such as Alzheimer’s Disease (AD). There are currently no treatments for these diseases, prompting an effort to both understand disease pathogenesis and develop novel approaches to mitigate the associated pathogenic proteostasis imbalances. As the overall integrity of the cellular proteome is a central facet of viability and function, the cell evolved a proteostasis network comprised of folding and degradation factors to ensure proper folding of nascent or misfolded peptides, or to promote their degradation if folding cannot be achieved. The endoplasmic reticulum (ER) Unfolded Protein Response (UPR) plays a crucial role in maintaining both intra- and extra-cellular proteostasis, as the ER environment is a main checkpoint for the folding of secreted proteins. Importantly, many proteins implicated in proteostasis-associated diseases including AD are trafficked through the secretory pathway and therefore interface with the ER proteostasis environment. Thus, the UPR is an attractive target for manipulating the levels of destabilized, disease-relevant proteins. This response is normally activated under circumstances of increased misfolded protein load in the ER lumen (i.e., ER stress), a signal which is sensed and transduced by the ER membrane proteins Inositol Requiring Enzyme 1 (IRE1), Protein kinase-like Endoplasmic Reticulum Kinase (PERK), and Activating Transcription Factor 6 (ATF6), which in turn induce proteostatic transcriptional programs to mitigate the unfolded protein load. This research project focuses specifically on the IRE1 signaling arm of the UPR, which has been extensively studied using genetic and chemical genetic approaches to demonstrate the therapeutic potential for activation of this pathway for multiple, etiologically-diverse diseases. While these approaches have been transformative for studying IRE1 signaling, compounds currently available for pharmacologic activation of this pathway are limited by their inherent promiscuity and cellular toxicity. For this research project, I utilize a luciferase-based high-throughput screening approach in conjunction with transcriptomic and proteomic studies to identify novel small molecule activators of the IRE1 signaling axis with a defined mechanism of action (Aim 1). Through these efforts, I have prioritized compound 474 as a promising first-in-class, non-toxic and specific activator of the IRE1 signaling axis in multiple cell culture models. In an effort to characterize the therapeutic potential of pharmacologic IRE1 activation, I will apply this compound and others to cellular models of AD to study effects on destabilized Amyloid Precursor Protein (APP) and Amyloid beta (Abβ) peptide levels as well as downstream physiological readouts of cellular health in relevant cell lines (Aim 2). These efforts will demonstrate the potential for pharmacologic IRE1 activation to mitigate disease-relevant imbalances in proteostasis implicated in AD and other diseases.