Drug-like small molecule modulators of the Integrated Stress Response

描述（由申请人提供）：早期分泌途径中的蛋白质错误折叠通过两种不同的机制发挥其病理影响：内质网（ER）质量控制机制对错误折叠突变蛋白的保留和降解导致功能丧失表型（以溶酶体贮积病为例）。错误折叠蛋白质的产生和积累通过引起ER应激而威胁细胞器的完整性，导致细胞功能障碍和死亡，这被认为有助于神经退行性疾病、糖尿病和其他衰老疾病。ER中的蛋白质折叠环境由少量信号转导途径控制，并且这些信号转导途径以定型的方式响应ER应激，介导未折叠蛋白质应答（UPR）。最近的证据表明，普遍定期审议的信号水平是由全球参数和ER的不同蛋白质客户的平均需求定义的。因此，普遍定期审议不太可能针对任何一种引起突变的疾病的具体紧急情况进行微调。这方面的证据最近提出了自己的情况下，敲除正常基因的功能，在UPR改善生存条件下的ER压力。这种稳态失败的例子表明，UPR可以在病理生理条件下进行治疗性操纵。翻译起始因子2a（eIF2a）的调节磷酸化和去磷酸化是UPR的一个很好理解的和潜在的可塑性臂，被称为整合应激反应（ISR）。我们建议确定药物样的小分子，将提高和其他将减少ISR的活动。ISR的瞬时抑制可能压倒ER的质量控制机制，并促进酶活性突变蛋白向其功能区室的运输，减轻相关的功能表型丧失。ISR失活也可能证明有益于基于细胞培养的生物治疗剂的生产，所述生物治疗剂用于治疗由酶替代疗法的错误折叠引起的疾病。ISR的激活剂可能保护细胞和器官免受ER应激的致命后果，并可能被证明在治疗衰老疾病中有用。为了实现这些目标，我们将开发抑制ER应激诱导的eIF2a激酶PERK的化合物的高通量筛选（HTS）和二次筛选以评估化合物的效力、特异性、生物利用度和脱靶效应。在平行链中，我们将开发用于eIF2a去磷酸化的抑制剂和用于eIF2a激酶的激活剂的其他HTS测定，所述eIF2a激酶通过非经典机制起作用并激活ISR而不引起应激。因此，该提案的长期目标是创建一个药理学平台，用于操纵对错误折叠蛋白质的细胞反应。鉴于蛋白质错误折叠在人类疾病中的普遍作用，这些药物很可能成为未来医生治疗设备的一部分。

DESCRIPTION (provided by applicant): Protein misfolding in the early secretory pathway exerts it pathological affects by two distinct mechanisms: Retention and degradation of misfolded mutant proteins by the endoplasmic reticulum's (ER) quality control machinery leads to loss-of-function phenotypes (exemplified by lysosomal storage diseases). Production and accumulation of the misfolded protein threatens the integrity of the organelle by causing ER stress, leading to cell dysfunction and death, which is believed to contribute to neurodegenerative disorders, diabetes mellitus and other diseases of aging. The protein folding environment in the ER is controlled by a small number of signal transduction pathways and these respond to ER stress in a stereotyped fashion, mediating the unfolded protein response (UPR). Recent evidence suggests that the level of signaling in the UPR is defined by global parameters and by the averaged needs of the ER's diverse protein clientele. It is unlikely therefore that the UPR is finely tuned to the specific exigencies of any one mutation-causing disease. Evidence for this has recently presented itself as circumstances in which knockout of normal genes that function in the UPR improved survival under conditions of ER stress. Such examples of failure of homeostasis indicate that the UPR can be manipulated therapeutically in pathophysiological conditions. Regulated phosphorylation and dephosphorylation of translation initiation factor 2a (elF2a) is a well understood and potentially malleable arm of the UPR, referred to as the integrated stress response (ISR). We propose to identify drug like small molecules that will enhance and others that will reduce the ISR's activity. Transient inhibition of the ISR might overwhelm the quality control mechanism of the ER and promote trafficking of enzymatically active mutant proteins to their functional compartment, alleviating the associated loss of function phenotypes. ISR inactivation might also prove beneficial in the cell culture based production of biotherapeutics used to treat diseases caused by misfolding by enzyme replacement therapy. Activators of the ISR are likely to protect cells and organs against the lethal consequences of ER stress and may prove useful in treating diseases of aging. To accomplish these goals we will develop high throughput screens (HTS) for compounds that inhibit the ER stress inducible elF2a kinase PERK and secondary screens to evaluate the potency, specificity, bioavailability and off-target effects of the compounds. In a parallel strand we will develop HTS assays for inhibitors of elF2a dephosphorylation and others for activators of elF2a kinases, which function by non-canonical mechanisms and activate the ISR without causing stress. The long-term goal of this proposal is therefore to create a pharmacological platform for manipulating the cellular response to misfolded proteins. Given the pervasive role of protein misfolding in human diseases, such agents are likely to become part of the therapeutic armamentarium of future physicians.