Selection of T-cell subset epitopes against F. hepatica using next generation phage display technology

使用下一代噬菌体展示技术选择针对肝片吸虫的 T 细胞子集表位

• 批准号: BB/M018520/1
• 负责人: Kevin Gough
• 金额: $ 18.96万
• 依托单位: University of Nottingham
• 依托单位国家: 英国
• 项目类别: Research Grant
• 财政年份: 2015
• 资助国家: 英国
• 起止时间: 2015 至 无数据
• 项目状态: 已结题
• 来源: https://gtr.ukri.org/projects?ref=BB%2FM018520%2F1
• 关键词: Selection; cell; subset; epitopes; against

Parasitic disease is a major cause of welfare and health issues in farmed livestock. Furthermore, the losses they cause and the need to provide veterinary care or drug treatments lead to massive economic losses in the UK and globally, e.g. US$2000M/yr for F. hepatica. The control of these diseases is primary through the application of drugs, however prolonged exposure of parasites has led to a rapid loss of drug efficacy due to the emergence of drug-resistant parasites. This has occurred in multiple parasite species and in response to multiple drug classes. Furthermore the continued use of some classes of drugs is no longer allowed due to the risk of entry of drug residues into the food chain. Moving forward the need for intensification of the domestic and international food supply will place increased pressure on farmers to ensure maximal output from their livestock. Correct control of parasitic disease is one route by which sustainable intensification can take place without compromising the health and welfare of livestock and ensuring economic returns. Prophylactic vaccination is the most suitable method by which this can be achieved as there are no concerns regarding drug residues or parasite resistance within the context of vaccination. Identification and selection of successful vaccine targets for helminths has proven difficult and so progress in bringing such vaccines to market is slow. Helminths are complex pathogens which have multiple modes of evading the host immune response and so develop long-lived infections. This is accompanied by a dampening of the host immune response to avoid host and disease pathology. The manipulation of the host immune response by helminths, such as F. hepatica, makes selection of vaccine candidates eliciting protective immunity difficult. Where moderate protection has been achieved in the case of F. hepatica this has involved a mixed immune response, involving antibody and cell-mediated protection. These opposite arms of the response are orchestrated by T-helper 1 (Th1) and T-helper 2 (Th2) cells. These cells differ in their recognition of parasite epitopes, and the time at which they appear during infection, with Th1 cells peaking during the first two weeks of infection and Th2 cells peaking 6-8 weeks after infection.We propose to isolate these cells from experimentally infected cattle and identify the parasite proteins they recognise by use of a novel technology called next generation phage display (NGPD). Animals experimentally infected with F. hepatica will develop robust immune responses generating large quantities of Th1 and Th2 cells in circulation. These can be identified by use of fluorescently tagged antibodies which recognize specific cell surface molecules which are unique to each cell type. These cells will be used to pan a phage display peptide library binding those for which they are specific. This will be conducted in two iterative rounds of panning to enrich the most frequently recognised peptides. Enriched peptides will be ranked by statistical analysis prior to being sythesised for testing using in vitro functional assays to determine the optimal composition of a multi-epitope vaccine. Synthesised peptides will be used in a flourescent screen. This will confirm those most commonly recognised by Th1 and Th2 cells in a mutually exclusive fashion. They will also be used to drive responses from Th1 and Th2 cells measuring production of key marker molecules to quantify the magnitude of response they can drive. The development of this tool and its related technologies will enable us to deliver a multi-target vaccine for protection against Fasciola hepatica. Furthermore, we will be in a position to apply this much needed technology to combat other production-limiting infections enabling progress in the development of vaccines for the livestock sector.

寄生虫病是养殖牲畜福利和健康问题的主要原因。此外，它们造成的损失和提供兽医护理或药物治疗的需要导致英国和全球的巨大经济损失，例如，F.肝。这些疾病的控制主要是通过应用药物，然而，由于抗药性寄生虫的出现，寄生虫的长期暴露导致药物功效迅速丧失。这发生在多种寄生虫种属中，并对多种药物类别产生反应。此外，由于药物残留进入食物链的风险，不再允许继续使用某些类别的药物。今后，需要加强国内和国际粮食供应，这将给农民带来更大的压力，以确保牲畜的最大产出。正确控制寄生虫病是可持续集约化的途径之一，而不会损害牲畜的健康和福利并确保经济回报。预防性疫苗接种是实现这一目标的最合适方法，因为在疫苗接种的背景下，不存在药物残留或寄生虫耐药性问题。确定和选择蠕虫的成功疫苗靶标已被证明是困难的，因此将此类疫苗推向市场的进展缓慢。蠕虫是一种复杂的病原体，具有多种逃避宿主免疫反应的模式，因此会产生长期感染。这伴随着宿主免疫应答的抑制以避免宿主和疾病病理。蠕虫对宿主免疫反应的操纵，如F。肝癌，使选择疫苗候选人引发保护性免疫困难。在F.这涉及混合免疫应答，包括抗体和细胞介导的保护。这些相反的反应臂由辅助性T细胞1（Th 1）和辅助性T细胞2（Th 2）协调。这些细胞在识别寄生虫表位方面有所不同，并且它们在感染过程中出现的时间不同，Th 1细胞在感染的前两周达到峰值，Th 2细胞在感染后6-8周达到峰值。我们建议从实验感染的牛中分离这些细胞，并通过使用称为下一代噬菌体展示（NGPD）的新技术来识别它们识别的寄生虫蛋白。实验感染F.肝细胞将产生强大的免疫应答，在循环中产生大量的Th 1和Th 2细胞。这些可以通过使用荧光标记的抗体来识别，所述抗体识别对于每种细胞类型独特的特定细胞表面分子。这些细胞将用于淘选噬菌体展示肽文库，结合它们特异性的那些肽。这将在两轮迭代淘选中进行，以富集最常识别的肽。富集的肽将在合成之前通过统计分析进行排序，以使用体外功能测定进行测试，以确定多表位疫苗的最佳组成。合成的肽将用于荧光素筛选。这将以相互排斥的方式确认Th 1和Th 2细胞最常识别的那些。它们还将用于驱动Th 1和Th 2细胞的反应，测量关键标志物分子的产生，以量化它们可以驱动的反应幅度。该工具及其相关技术的开发将使我们能够提供多靶点疫苗来保护肝片吸虫。此外，我们将能够应用这一急需的技术来防治其他限制生产的感染，从而在畜牧业疫苗的开发方面取得进展。

项目成果

Fasciola hepatica, TGF-ß and host mimicry: the enemy within.

肝片形吸虫、TGF-β 和宿主拟态：内部的敌人。

• DOI: 10.1016/j.mib.2018.09.002
• 发表时间: 2018
• 期刊: Current opinion in microbiology
• 影响因子: 5.4
• 作者: Musah-Eroje M

A rapid IL-17 response to Cryptosporidium parvum in the bovine intestine.

• DOI: 10.1016/j.vetimm.2017.07.009
• 发表时间: 2017-09
• 期刊: Veterinary immunology and immunopathology
• 影响因子: 1.8
• 作者: Drinkall E;Wass MJ;Coffey TJ;Flynn RJ
• 通讯作者: Flynn RJ

The Chronic Stages of Bovine Fasciola hepatica Are Dominated by CD4 T-Cell Exhaustion.

牛fasciola Hepatica的慢性阶段以CD4 T细胞耗尽为主。

• DOI: 10.3389/fimmu.2017.01002
• 发表时间: 2017
• 期刊: Frontiers in immunology
• 影响因子: 7.3
• 作者: Sachdev D;Gough KC;Flynn RJ
• 通讯作者: Flynn RJ

Fasciola hepatica infection reduces Mycobacterium bovis burden and mycobacterial uptake and suppresses the pro-inflammatory response.

• DOI: 10.1111/pim.12326
• 发表时间: 2016-07
• 期刊: Parasite immunology
• 影响因子: 2.2
• 作者: Garza-Cuartero L;O'Sullivan J;Blanco A;McNair J;Welsh M;Flynn RJ;Williams D;Diggle P;Cassidy J;Mulcahy G
• 通讯作者: Mulcahy G