Engineered R-type pyocins as bactericidal agents against E.coli O157:H7

描述(由申请人提供)：我们的总体目标是开发针对E.ColiO157：H7的杀菌R型脓菌素，作为一种针对感染和污染的特定治疗方法。肠出血性大肠杆菌(EHEC)是一组通过食物和水传播的病原体，可导致从非出血性腹泻到溶血性尿毒症综合征(HUS)等各种疾病。大肠杆菌O157：H7既会引起大规模暴发，也会引起零星感染，从而对公众健康构成威胁。这里提出的工作是基于我们的发现，R型腐菌素是一种高度特异和高效的杀菌蛋白复合体，已被证明对动物的全身细菌感染有效。虽然这些杀菌剂对其目标生物体具有高度特异性，但通过用噬菌体相关的尾部蛋白修饰它们的尾部纤维，这些杀菌剂可以重定向到其他细菌。我们的领先候选产品AvR2-V10由噬菌体V10的尾部纤维与R2-型腐菌素融合而成，因此嵌合颗粒对所有测试的O157大肠杆菌菌株具有杀菌特异性-即噬菌体的特异性。在这里提出的工作中，我们将推进AvR2-V10作为抗O157：H7大肠杆菌制剂的开发。我们假设R型腐菌素可以在E.ColiBL21系统中合成和生产，产生有效的治疗E.ColiO157：H7感染的药物。为了实现我们的目标，我们将通过在E.ColiBL21中以可调节的方式生产AvR2-V10来进行临床前测试，使其符合GLP标准(特定目标1)。我们将在模拟与O157：H7感染相关的人类病理的幼兔模型中评估其疗效(特定目标2)。该候选产品旨在用作专门针对E.ColiO157的治疗或预防药物，并能在流行情况下提供有效的反应，无论是意外感染还是故意大规模感染。公共卫生相关性：O157大肠杆菌可导致严重的肠道感染和潜在的死亡，并以生物恐怖袭击或意外食品或水传播疫情的形式对公共卫生构成威胁。我们已经利用我们的平台技术设计了一种有效的杀菌剂AvR2-V10，专门针对E.ColiO157。这种试剂现在被开发为治疗或预防大肠杆菌O157感染或污染的药物，在流行情况下提供有效的反应。在此，我们建议开发一种简单、高产的该制剂的生产方法，并在兔模型上测试其效果。

DESCRIPTION (provided by applicant): Our overall Goal is to develop our bactericidal R-type pyocin targeting E.coli O157:H7, as a specific therapeutic against infection and contamination. Enterohemorrhagic E. coli (EHEC) are a group of food- and waterborne pathogens that cause illnesses ranging from non-bloody diarrhea to hemolytic uremic syndrome (HUS). E. coli O157:H7 causes both large outbreaks as well as sporadic infections, and thus poses a threat to public health. The work proposed here is based on our findings that R-type pyocins are highly specific and highly potent bactericidal protein complexes that have been shown to be efficacious for systemic bacterial infections in animals. While highly specific to their target organism, these bactericidal agents can be retargeted to other bacteria by modifying their tail fibers with the related tail proteins of bacteriophages. Our leading product candidate, AvR2-V10, consists of the tail fiber of the bacteriophage ?V10 fused to an R2-type pyocin, such that the chimeric particle has the bactericidal specificity against all tested O157 E. coli strains - i.e. the specificity of the bacteriophage. In the work proposed here, we will advance the development of AvR2-V10 as an anti E. coli O157:H7 agent. We hypothesize that R-type pyocins can be synthesized and produced in an E. coli BL21 system, yielding an effective therapeutic against E. coli O157:H7 infection. In order to achieve our goals, we will conduct preclinical testing of AvR2-V10 by producing it in a regulatable manner in E. coli BL21, such that it abides to GLP standards (Specific Aim #1). We will evaluate its efficacy in an infant rabbit model which mimics the human pathology associated with O157:H7 infection (Specific Aim #2). The product candidate is intended to be used as a therapeutic or prophylactic that specifically targets E.coli O157 and can offer a potent response in cases of epidemic, either accidental or intentional mass infection. PUBLIC HEALTH RELEVANCE: E. coli O157 can cause severe intestinal infections and potentially death, and poses a threat to public health in the form of a bioterrorist attack or accidental food or waterborne outbreak. We have used our platform technology to engineer a potent bactericidal agent, AvR2-V10, to specifically target E.coli O157. This agent is now being developed as a therapeutic or prophylactic against E.coli O157 infection or contamination, offering a potent response in cases of epidemic. Herein, we propose to develop a simple, high yield production method for this agent, and test it's efficacy in a rabbit model.