A high throughput screening assay for the identification of SUMOylation inhibitor

描述(由申请人提供)：这项提案的总体目标是进行高通量筛选分析(HTS)以确定SUMO化的抑制剂。近年来，由SUMO(小泛素样修饰物)家族进行的蛋白质修饰被认为是一种重要的翻译后修饰，在基因转录、细胞周期进展、DNA修复、病毒和细菌感染以及神经退行性疾病的发生发展中发挥着重要的作用。SUMO化抑制剂将作为研究SUMO化在不同细胞功能中的作用的迫切需要的试剂，这些作用在很大程度上仍不清楚。这些抑制剂还将对癌症和病毒感染等疾病具有治疗潜力。然而，这样的抑制剂还没有报道，科学界也还没有得到。我们已经开发了几种方法来鉴定这种抑制剂。主要的HTS检测使用Alpha筛选技术。二次筛查试验使用荧光共振能量转移(FRET)来消除在Alpha筛查中获得的假阳性。此外，泛素化试验将被用来识别抑制SUMO化而不是其他泛素样修饰系统的特异性HITS。筛选中确定的抑制剂的直接应用是：1)探索SUMO化在不同细胞功能中的作用，如在DNA修复途径中；2)在已被证明在癌症和病毒感染等疾病中发挥重要作用的细胞和动物模型中测试这些抑制剂。我们的长期目标是在本研究确定的HITS的构效关系研究的基础上，开发有效的SUMO化抑制剂。同源泛素-蛋白酶体途径的抑制剂在剖析泛素-蛋白酶体途径在许多细胞功能中的作用方面发挥了关键作用，并作为抗癌治疗药物取得了令人惊讶的成功。苏莫化抑制剂在研究中具有广泛影响的类似前景，并可能导致建立一种新的范例，开发治疗癌症和传染病等毁灭性疾病的药物。公共卫生相关性： 这项提案的总体目标是进行高通量筛选分析(HTS)以确定SUMO化的抑制剂。

DESCRIPTION (provided by applicant): The overall goal of this proposal is to perform high throughput screening assays (HTS) to identify inhibitors for SUMOylation. Protein modification by the SUMO (Small Ubiquitin-like MOdifier) family of proteins has recently been established as an important post- translational modification that plays an essential role in many functions including gene transcription, cell cycle progression, DNA repair, viral and bacterial infection, and the development of neurodegenerative diseases. Inhibitors of SUMOylation will serve as much needed reagents for investigating the role of SUMOylation in different cellular functions, which remain largely unclear. The inhibitors will also have therapeutic potential for diseases, such as cancer and viral infection. However, such inhibitors have not been reported and are not yet available to the scientific community. We have developed several assays to identify such inhibitors. The primary HTS assay uses the ALPHA screening technology. The secondary screening assay uses fluorescence resonance energy transfer (FRET) to eliminate false positive hits obtained in the ALPHA screen. Additionally, an ubiquitination assay will be used to identify the hits that are specific for inhibiting SUMOylation and not other ubiquitin-like modification systems. The immediate applications of the inhibitors identified in the screen are: 1) to probe the role of SUMOylation in different cellular functions, such as in the DNA repair pathways, 2) to test the inhibitors in cellular and animal models of diseases where SUMOylation has been shown to play important roles, such as in cancer and viral infections. Our long term goal is to develop potent SUMOylation inhibitors based on the structure-activity relationship studies of the hits identified from this study. Inhibitors of the homologous ubiquitin-proteosome pathway have been critical in dissecting the role of the ubiquitin-proteosome pathway in many cellular functions and are surprisingly successful as anti-cancer therapeutic agents. SUMOylation inhibitors hold similar promise in having a broad impact in research and could lead to the establishment of a new paradigm in developing therapeutic agents for devastating diseases, such as cancer and infectious diseases. PUBLIC HEALTH RELEVANCE: The overall goal of this proposal is to perform high throughput screening assays (HTS) to identify inhibitors for SUMOylation.