Preclinical evaluation of psoralen inactivation as a platform for developing a killed, whole cell vaccine for Enterotoxigenic E. coli (ETEC)

西科特项目摘要/摘要 产肠毒素大肠埃希菌(ETEC)、福氏志贺菌和空肠弯曲菌为优势菌 发展中国家胃肠道(GI)疾病的发病率和死亡率的原因。急性呼吸窘迫综合征 由这些药物引起的腹泻与发育缺陷和慢性胃肠道综合征有关。 需要对幼儿和成年旅行者以及部署在军队中的人使用安全的疫苗 预防这些微生物引起的感染和感染后的后遗症。福尔马林灭活是当前的趋势 生产灭活型全细胞细菌疫苗的标准。这种方法不需要基因 对诱导保护性免疫反应的抗原的操作或广博的知识。然而， 福尔马林使蛋白质交联，有可能破坏诱导保护性免疫的抗原表位。 在活体细菌感染的背景下的免疫反应。我们建议测试一种替代方案的可行性 为肠道细菌生产安全的全细胞疫苗的灭活方法。补骨脂素是一种可光激活的物质 可逆地嵌入到核酸中的药物。用长波紫外线照射后 (P+UVA)，在嘧啶残基形成共价链间交联，防止基因组复制。自.以来 P+UVA通过交联核酸而不是蛋白质来灭活细菌，我们假设蛋白质 抗原将以原生形式保存，产生具有卓越免疫原性的疫苗。我们有 已成功使用P+UVA灭活ETEC。有了这项建议，我们将通过比较来延长ETEC的工作 P+UVA对福尔马林灭活的ETEC抗原保存和免疫原性的影响 建立了体外检测和小鼠模型。对这两种失活方法的直接比较分析是 需要评估P+UVA方法作为肠道细菌敏捷平台的价值和可行性 疫苗，更广泛地说，用于预防影响患者的抗药性细菌感染的疫苗 在医院环境中的护理。 。 1

Westcott.Project Summary/Abstract Enterotoxigenic E. coli (ETEC), Shigella flexneri and Campylobacter jejuni are the predominant bacterial causes of morbidity and mortality from gastrointestinal (GI) disease in the developing world. Episodes of acute diarrhea caused by these agents have been linked to developmental defects and chronic GI syndromes. Vaccines that are safe for use in young children as well as adult travelers and deployed military are needed to prevent infection and post-infection sequellae caused by these organisms. Formalin inactivation is the current standard for generating killed, whole cell bacterial vaccines. This approach does not require genetic manipulations or extensive knowledge of antigens that induce protective immune responses. However, formalin crosslinks proteins, which has the potential to destroy antigenic epitopes that induce protective immune responses in the context of live bacterial infection. We propose to test the feasibility of an alternative inactivation method to generate safe, whole cell vaccines for enteric bacteria. Psoralen is a photoactivatable drug that reversibly intercalates into nucleic acids. Following irradiation with long wavelength UV light (P+UVA), covalent inter-strand crosslinks form at pyrimidine residues, preventing genome replication. Since P+UVA inactivates bacteria by crosslinking nucleic acids rather than proteins, we hypothesize that protein antigens will be preserved in a native form, generating vaccines with superior immunogenicity. We have successfully inactivated ETEC using P+UVA. With this proposal we will extend the ETEC work by comparing P+UVA to formalin-inactivated ETEC with respect to antigen preservation and immunogenicity using established in vitro assays and murine models. A direct comparative analysis of the 2 inactivation methods is needed to assess the value and feasibility of the P+UVA approach as an agile platform for enteric bacterial vaccines, and more generally for vaccines to prevent antibiotic-resistant bacterial infections that impact patient care in hospital settings. . 1