High Throughput Screening for Small Compounds to Inhibit the Expression of Strept

描述(申请人提供)：细菌中普遍存在的抗生素耐药性正日益引起人们的关注，成为一种主要的公共卫生威胁。目前的抗生素会导致目标细菌死亡或生长停滞。因此，抗生素的使用施加了强大的选择压力，有利于抗生素耐药菌株。新型抗菌试剂可以抑制病原菌的毒力，而不需要选择抗生素耐药性，为传染病的治疗提供了一种很有前途的替代方法。A组链球菌(GAS)是一种重要的人类病原体，每年影响全球数百万人。链激酶(SK)是一种主要的气体毒力因子，可以激活人的纤溶酶原。在我们以前的研究中，我们已经确定链激酶/纤溶酶原相互作用是GAS发病的关键因素。我们建议利用这一观察结果，设计新的抗菌试剂来治疗气体感染。在初步研究中，我们已经筛选了55,000个小化合物来抑制SK在GAS中的表达，并确定了23个候选的Hit化合物。利用NIH分子库和成像路线图倡议，提出了一种额外的高通量筛选，用于SK表达抑制剂的高达500,000多个小化合物。基于生长的筛选将被优化为使用SK启动子控制的具有卡那霉素抗性基因的GAS菌株来筛选能够抑制卡那霉素抗性表达的小分子化合物，这些化合物将作为SK表达抑制剂的先导化合物。未来，将研究候选化合物对GAS基因表达的全局影响，为识别靶标提供线索。在已发表的报告中已经证明可以调控SK表达的两个GAS双组分系统将作为主要候选目标进行测试。此外，我们以前的研究中建立的一些小鼠气体感染模型将被用来阐明候选化合物在体内对气体毒力的影响。以上研究收集的数据将进一步加深我们对SK在气体感染中的贡献的理解，并寻找能够抑制气体毒力的小分子化合物。因此，可以探索通过干扰气体毒力而不过度引入耐药性选择压力来治疗细菌感染的替代方法，以补充抗生素治疗。

DESCRIPTION (provided by applicant): The widespread occurrence of antibiotic resistance among bacteria is causing increasing concern as a major public health threats. Current antibiotics cause death or growth arrest in the target bacteria. As a result, antibiotic use exerts strong selective pressure to favor antibiotic resistant strains. Novel antimicrobial reagents that suppress pathogen virulence without selecting for antibiotic resistance provide a promising alternative approach for treatment of infectious diseases. Group A Streptococcus (GAS) is an important human pathogen affecting millions of people globally each year. The streptokinase (SK) is a major GAS virulence factor that activates human plasminogen. In our previous studies, we have established the streptokinase/plasminogen interaction as a critical factor in GAS pathogenesis. We propose to take advantage of this observation and design novel antimicrobial reagents for the treatment of GAS infection. In the preliminary study, we have screened 55,000 small compounds for inhibitors of SK expression in GAS and 23 candidate hit compounds have been identified. An additional high throughput screen of up to 500,000 more small compounds for SK expression inhibitors is proposed by taking advantage of the NIH Molecular Libraries and Imaging roadmap initiative. A growth based screen will be optimized to use a GAS strain with kanamycin resistance gene under control of SK promoter to screen for small compounds that can inhibit kanamycin resistance expression, which will serve as lead compounds for SK expression inhibitors. In the future, global effects of candidate compounds on GAS gene expression will be studied to provide clues for identification of the targets. Two GAS two-component systems that have been demonstrated to regulate SK expression in published reports will be tested as prime candidate targets. In addition, a number of murine GAS infection models established in our previous studies will be used to elucidate the effects of candidate compounds on GAS virulence in vivo. Data collected from the above studies will further our understanding of the contribution of SK to GAS infection and identify small compounds that can inhibit GAS virulence. As a result, alternative approach to treat bacterial infection by interfering with GAS virulence without unduly introducing selection pressure for resistance can be explored to supplement antibiotic treatment.