HTP chemical genomic screens to identify positive regulators of the Unfolded Prot

描述（由申请人提供）：我们对病理机制的理解取得了巨大进展，最近发现分泌途径中有缺陷的蛋白质加工是许多遗传和环境疾病的组成部分。从糖尿病、阿尔茨海默病和帕金森病到血友病和溶酶体贮积病的各种疾病状态都以折叠缺陷或从内质网（ER）的转运受损为特征。非常重要的是，最近发现蛋白质合成的失调可能是多种人类癌症的致病性转移的关键组成部分。当错误折叠的蛋白质在ER腔中积累时，细胞激活未折叠蛋白质反应（UPR）以清除错误折叠的蛋白质并恢复稳态蛋白质加工。当压力延长或强烈时，UPR采用导致细胞死亡的遗传途径。这些拟议的研究集中于鉴定和表征UPR的这两个不同分支的药物样小分子调节剂。我们将研究以下假设：1）IRE 1-XBP 1的激活- UPR的（适应性）臂将促进其中细胞被错误折叠或分泌不良的蛋白质的积累所累的病理中的延长的恢复机会;和2）PERK-eIF 21-CHOP的特异性和稳健活化（凋亡）臂将加剧UPR并导致具有过度活跃的分泌机制的恶性细胞中的细胞死亡。在我们的初步研究中，我们已经成功地筛选了具有稳定表达荧光素酶构建体的中国卵巢（CHO）细胞系的66，000个化合物文库，所述荧光素酶构建体在UPR的IRE 1-XBP 1或PERK-eIF 21-CHOP臂上单独报告。在与专家医药，有机/合成和计算化学家团队合作后，我们已经确定了一些有效调节UPR的命中。我们现在建议通过重复筛选具有更大化学多样性和复杂性的文库来扩展我们的探针数据库，目的是鉴定更特异和有效的UPR调节探针。 公共卫生相关性：拟议研究的长期目标是确定特异性激活未折叠蛋白反应（UPR）的适应性或细胞死亡臂的新型小分子。我们将描述：1）适应性激活剂作为治疗剂用于治疗由异常蛋白质折叠和/或运输引起的疾病的潜力;和2）诱导凋亡的化合物杀死头颈癌细胞的能力。通过所提出的研究鉴定的小分子UPR激动剂/拮抗剂将提供治疗患有疾病的患者的方便的具有成本效益的手段，所述疾病的治疗限于最低限度有效的基因治疗或广泛的昂贵的酶替代治疗。

DESCRIPTION (provided by applicant): Tremendous advances in our understanding of pathologic mechanisms have recently revealed that defective protein processing within the secretory pathway is an integral component of many genetic and environmental diseases. Diverse disease states ranging from diabetes, Alzheimer's disease, and Parkinson's disease, to hemophilia and lysosomal storage diseases have all been characterized by folding defects or impaired transport from the endoplasmic reticulum (ER). Very importantly, it has recently come to light that deregulation of protein synthesis is likely a key component in the pathogenic metastasis of a variety human cancers. When misfolded protein accumulates in the ER lumen, the cell activates the Unfolded Protein Response (UPR) to clear the malfolded proteins and restore homeostatic protein processing. When a stress is prolonged or robust, the UPR employs a genetic pathway that results in cell death. These proposed studies are focused to identify and characterize drug-like small molecule regulators of these two distinct arms of the UPR. We will investigate the hypotheses that: 1) Activation of the IRE1-XBP1 (adaptive) arm of the UPR will facilitate a prolonged recovery opportunity in pathologies where cells are burdened with an accumulation of mis-folded or poorly secreted proteins; and 2) Specific and robust activation of the PERK-eIF21-CHOP (apoptotic) arm will exacerbate the UPR and lead to cell death in malignant cells with hyper-active secretory machinery. In our preliminary studies, we have successfully screened a 66,000 compound library with Chinese Hamster Ovary (CHO) cell lines stably expressing luciferase constructs that report individually on the IRE1-XBP1 or PERK-eIF21-CHOP arms of the UPR. Having worked with a team of expert Medicinal, Organic/Synthetic and Computational Chemists we have identified some hits that productively modulate the UPR. We now propose to expand our probe database by repeating the screen with libraries of greater chemical diversity and complexity with the goal of identifying more specific and potent UPR-regulating probes. PUBLIC HEALTH RELEVANCE: The long-term objective of the proposed studies is to identify novel small molecules that specifically activate adaptive or cell death arms of the unfolded protein response (UPR). We will characterize: 1) the potential of the adaptive activators as therapeutic agents for treating diseases caused by aberrant protein folding and/or trafficking; and 2) the ability of the apoptosis-inducing compounds to kill head and neck cancer cells. Small molecule UPR agonists/antagonists identified through the proposed studies will provide a convenient cost-effective means of treating patients suffering from diseases for which therapies are limited to minimally effective gene therapy or extensive costly enzyme replacement therapies.