A novel vaccine to prevent Clostridium difficile colitis

一种预防艰难梭菌结肠炎的新型疫苗

• 批准号: 8668719
• 负责人: EDGAR C. BOEDEKER
• 金额: --
• 依托单位: ALBUQUERQUE VA MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 2013
• 资助国家: 美国
• 起止时间: 2013-04-01 至 2017-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8668719
• 关键词: Antibiotics; Antibodies; Antigens; Attenuated; Bacterial Adhesins; Binding; C-terminal; Clostridium difficile; Colitis; Development; Disease; Elderly; Engineering; Escherichia coli; Escherichia coli EHEC; Escherichia coli Infections; General Population; Genes; Genetic Techniques; Goals; Healthcare Systems; Hospitals; Immune system; Immunity; Immunization; Incidence; Infection; Intestines; Laboratories; Lead; Lesion; Life; Measurement; Modeling; Molecular Genetics; Mus; Nosocomial Infections; Organism; Patients; Phase; Plasmids; Production; Proliferating; Proteins; Recurrence; Reproduction spores; Risk; Safety; Serum; Shiga Toxin; Surface; System; Systemic disease; Terminal Repeat Sequences; Toxin; United States Department of Veterans Affairs; Vaccines; Veterans; Virulence; immunogenic; immunogenicity; mouse model; novel therapeutics; novel vaccines; oral vaccine; prevent; prophylactic; protective efficacy; public health relevance; receptor binding; safety testing; vaccine efficacy; vector; vector vaccine

DESCRIPTION (provided by applicant): The need for novel therapeutic and prophylactic approaches to Clostridium difficile (C. difficile) infections in the VA Health Care system, is urgent. The overall goal of this proposal is to develop live oral vaccines which express C. difficile toxin antigens which will induce protective mucosal, and systemic, immunity to the toxins in order to prevent Clostridium difficile-associated disease (CDAD). To achieve this, an attenuated strain of E.coli which we have developed as a vaccine vector for antigen delivery will be engineered to express the immunogenic C-terminal receptor binding portions of the major virulence determinants of C.difficile, the A and B toxins (TcdA and TcdB). To evaluate vaccine efficacy we will use a recently described antibiotic (cefoperazone)-treated mouse model of CDAD(70) which we have validated in our laboratory. To express protective toxin antigens in immunogenic form in our E.coli vector strain, we will incorporate them into the passenger domain of autotransporter(28) proteins to deliver them to the mucosal immune system as we have done for other toxin antigens. This proposal has 3 specific aims: The first specific aim is to express immunogenic components of the TcdA and TcdB toxins of C. difficile as antigens in our attenuated E. coli vector ZCR533(6). The C-terminal binding regions of the toxins will be expressed as fusions with the passenger domain of the autotransporter EspP, the intrinsic type V secretion system(28) of O157:H7 E.coli, and introduced on a medium copy number plasmid. The second specific aim is to determine the safety and mucosal immunogenicity of the vaccine constructs by intranasal, and intragastric, immunization of mice, followed by measurement of the binding and toxin neutralizing capacity ofserum and mucosal anti-TcdA and/or anti-TcdB antibodies. The third specific aim is to determine the protective efficacy of the vaccine constructs in C57/Bl6 mice using a newly described antibiotic-treated mouse model(70) for C. difficile challenge. We have recently validated this model in our laboratory. Our vaccine vector will be ZCR533, an attenuated E.coli strain derived from an O157:H7 enterohemorrhagic E.coli (EHEC) by deleting its genes for shiga toxin production and truncating its major adhesin, intimin, such that it can no longer induce attaching /effacing lesions, but can still induce anti-intimin antibody(8,75). We will use standard molecular genetics techniques to prepare C. difficile vaccine constructs by expressing the C terminal repeats comprising the binding domains of the TcdA and TcdB toxins, into the EspP autotransporter(28) on a multicopy plasmid. Our recent studies have indicated the validity of this approach to immunization against toxin antigens, since the similarly-sized shiga toxin B subunit antigen is immunogenic, and protective against shiga toxin producing E.coli (STEC) infection, when expressed in the autotransporter EspP on the surface of the organism, or secreted into the medium. To test the safety, immunogenicity and protective efficacy of the vaccine constructs we will utilize mouse models of intranasal, and intragastric, immunization(12) and a new cefoperazone-treated mouse model (70) of CDAD, which we have validated in our laboratory, for C. difficille challenge. These studies could lead to the development of a new class of safe and effective vaccines directed against C.difficile toxins to prevent CDAD.

描述（由申请人提供）： 迫切需要对VA卫生保健系统中艰难梭菌（艰难梭菌）感染的新型治疗和预防性方法。该提案的总体目标是开发表达艰难梭菌毒素抗原的活疫苗，这些疫苗将诱导保护性粘膜，并对毒素进行全身性免疫，以防止艰难梭菌相关疾病（CDAD）。为了实现这一目标，我们已将其作为抗原递送的疫苗载体开发的衰减菌株将被设计为表达c.difficigile，A和B毒素（TCDA和TCDB）的主要毒力决定因素的免疫原性C末端受体结合部分。为了评估疫苗功效，我们将使用最近描述的经CDAD的抗生素（头孢曲松）处理的小鼠模型（70），我们在实验室中验证了。为了在我们的大肠杆菌载体菌株中以免疫原性形式表达保护性毒素抗原，我们将它们纳入自转运蛋白（28）蛋白的乘客结构域，以将其输送到粘膜免疫系统中，就像我们为其他毒素抗原所做的那样。 该提议具有3个特定目的：第一个具体目的是表达艰难梭菌的TCDA和TCDB毒素的免疫原性成分，作为我们衰减的大肠杆菌载体ZCR533中的抗原（6）。毒素的C末端结合区域将以与自动转移剂ESPP的乘客结构域的融合表示，O157：H7 e.coli的内在型V分泌系统（28），并在中拷贝数质粒上引入。 第二个具体目的是通过鼻内和胃内的小鼠免疫来确定疫苗构建体的安全性和粘膜免疫原性，然后测量结合和毒素中和毒素的能力和粘膜和粘膜抗TCDA和/或抗TCDA和/或抗TCDB抗体。 第三个具体目的是使用新描述的抗生素处理的小鼠模型（70）来确定C57/BL6小鼠在C57/BL6小鼠中的保护性疗效（艰难梭菌挑战）。我们最近在我们的实验室中验证了该模型。 我们的疫苗载体将是ZCR533，这是一种源自O157：H7肠肠肠癌e.coli（EHEC）的衰减。我们将使用标准的分子遗传学技术来制备艰难梭菌疫苗构建体，以表达包含TCDA和TCDB毒素的结合结构域的C末端重复剂，并在多拷贝质粒上进入ESPP自转运蛋白（28）。我们最近的研究表明，这种对毒素抗原的免疫方法的有效性，因为类似大小的志贺毒素B亚基抗原具有免疫原性，并且针对产生E.coli（STEC）感染的Shiga毒素的保护性，在自自转移者ESPP中表达在自身转移者ESPP的表面上。为了测试疫苗构建体的安全性，免疫原性和保护性疗效，我们将利用鼻内的小鼠模型和胃内的免疫，免疫（12）和新的头孢曲松治疗的CDAD的新型CDAD的小鼠模型（70），我们在我们的实验室中已经验证了C. Loodille Checkally验证的CDAD。这些研究可能导致开发针对C. c. c. codexin的新的安全有效疫苗，以防止CDAD。