Evaluation of Memory Responses and Biomarkers from a Phase IEnterotoxigenic Escherichia coli (ETEC) Intramuscular SubunitVaccine with dmLT Adjuvant

产肠毒素大肠杆菌(ETEC)是引起儿童、旅行者和军人细菌性感染性腹泻的主要原因 危险地区的军事人员。因此，疫苗的开发将有利于公众健康。 一种策略是使用定居因子的亚基与不耐热毒素(LT)的类毒素相结合。最近， 进行了首例人类安全性和免疫原性1期疫苗试验(NCT03404674)。有了剂量- CS6亚单位抗原CSSBA联合LT-R192G/L211A(DmLT)肌肉注射递增。不是 报道了严重的不良反应，我们在几个队列中观察到了很强的体液免疫原性， 与dmLT剂量显著相关。然而，对临床试验样本的完整分析，包括体液和细胞 记忆力不足。正如本疫苗试验表明的那样，dmLT不仅是一种LT类毒素，而且是一种有效的佐剂， 激发对联合递送抗原的免疫力；然而，我们对分子的理解存在差距 与dmLT共同传递的抗原启动疫苗接种结果的机制。 这项建议的目的是扩大对ETEC第一阶段临床的血清和PBMC样本的分析 并确定指导疫苗结果的关键生物标记物和分子机制。在建议的 我们的目标是探索(1)疫苗剂量如何调节记忆力、寿命和多样性的发展 体液反应；(2)疫苗接种如何改变持久细胞免疫的发展；(3)早期 信号事件可作为免疫的生物标志物；以及(4)免疫过程中的分子机制 塑造疫苗接种结果。 为此，我们将分析我们存储的临床试验样本并执行一些复杂的分析， 包括使用相关ETEC 2b期疫苗样本的转录和代谢组学分析- 效力试验，以确定疫苗诱导保护的签名，然后我们将使用第一阶段试验进行验证 样本。此外，我们将用细胞分析和小鼠模型来验证研究结果。这些发现将会有所帮助 随着ETEC疫苗的开发，人类通过非肠道途径使用，也提供了机制 洞察指导疫苗接种结果的关键事件。

Enterotoxigenic E. coli (ETEC) is a major cause of bacterial infectious diarrhea in children, travelers and deployed military personnel in risk areas. As such, development of a vaccine would be advantageous for public health. One strategy is to use subunits of colonization factors combined with toxoids of heat-labile toxin (LT). Recently, a first-in-humans safety and immunogenicity Phase 1 vaccine trial (NCT03404674) was conducted. with dose- escalating intramuscular delivery of CS6-subunit antigen CssBA combined with LT-R192G/L211A (dmLT). No serious adverse events were reported and we observed strong humoral immunogenicity in several cohorts, notably related to dmLT dose. Yet a complete analysis of clinical trial samples, including humoral and cellular memory is lacking. As this vaccine trial indicates, dmLT is not only an LT toxoid but also a potent adjuvant that stimulates immunity to co-delivered antigens; however, there is a gap in our understanding of molecular mechanisms responsible for initiating vaccination outcomes with antigens co-delivered with dmLT. The objective of this proposal is to expand analysis on serum and PBMC samples from an ETEC Phase 1 clinical trial and to define the key biomarkers and molecular mechanisms directing vaccine outcomes. In the proposed studies we aim to explore (1) how vaccination doses modulated development of memory, longevity and diversity of the humoral response; (2) how vaccination altered development of durable cellular immunity; (3) whether early signaling events can serve as biomarkers of immunity; and (4) what molecular mechanisms during immunization shape vaccination outcomes. To do so we will analyze our stored clinical trial samples and perform a number of sophisticated analyses, including transcriptional and metabolomics assays using samples from a related ETEC Phase 2b vaccine- efficacy trail to define signatures of vaccine induced protection which we will then validate using the Phase 1 trial samples. In addition, we will validate findings with cellular analyses and mouse models. These findings will help with the development of an ETEC vaccine for human use by parenteral route and also provide mechanistic insight into key events directing vaccination outcomes.