Towards a vaccine against Fasciola hepatica using FhSAP2 as an antigen

描述（由申请方提供）：蠕虫寄生虫肝片形吸虫引起肝吸虫病或片形吸虫病，从而影响人类以及绵羊、牛和山羊的健康。片形吸虫病每年造成的农业损失估计超过32亿美元。片形吸虫病也是一种主要的人畜共患疾病，估计全世界有1 700万人感染。三氯苯达唑是目前控制片形吸虫病最有效的药物，但治疗费用和感染后绵羊抗药性的出现是影响其疗效的主要因素。肝病表明，需要制定可持续的战略，如接种疫苗，以控制这一疾病。然而，尽管长期的研究，肝片吸虫疫苗尚未开发。 我们已经确定了一个新的F。称为FhSAP 2的肝抗原，当在弗氏完全佐剂（FCA）中皮下（SC）递送时，能够在用F.肝囊蚴FhSAP 2是一个分子量为11.5kDa的多肽，属于F.肝鞘脂激活蛋白样/NK-溶素蛋白家族。我们还证明了FhSAP 2-FCA制剂诱导的免疫与高水平的IgG 2a抗体和高水平的IFN 3相关，这是Th 1免疫应答的特征。这些结果提示FhSAP 2可能通过与CD 4 + Th 1细胞相关的机制介导FhSAP 2的保护作用。与大多数蠕虫一样，肝吸虫在传统上与Th 2免疫应答相关。本研究的目的是研究T辅助细胞1，T辅助细胞2反应的调节和对F。在肝片吸虫病的小鼠模型中用这些制剂治疗由FhSAP 2诱导的肝硬化。为了实现这一目标，我们建议在三年资助期内实现两个具体目标。在具体目标-1中，我们将研究当在含有免疫刺激序列的佐剂中SC递送时FhSAP 2的保护作用。具体地，我们将确定用Montanide TM ISM 1312和伊萨70 M中的FhSAP 2或捕获到ISCOM中的FhSAP 2与IL-12或CpG-ODN共同施用的初免-加强策略是否可以模拟或增强用CFA中的FhSAP 2获得的保护水平。这将导致疫苗配方的优化，用于未来的研究。在特定的目标-2中，我们将研究在目标-1中使用的疫苗制剂如何调节辅助性T细胞1（Th 1）-辅助性T细胞2（Th 2）应答。我们将观察Th 1/Th 2细胞因子谱、抗体亚类水平以及在naove小鼠中引发的B-和T-细胞、巨噬细胞（MX）和树突状细胞（DC）的活化状态，并且这些参数将与在特异性目的-1中获得的保护水平相关。结果将使我们能够测试我们的假设的保护作用的Th 1反应片形吸虫病。 如果所提出的目标得以实现，我们不仅将获得一种用于反刍动物试验的优化疫苗配方，而且还将提出一种新的概念，即如何诱导对F. hepatica，这将打开大门，进一步研究，以阐明免疫机制，使可能的Th 1免疫反应，是有效的，对多细胞吸虫，如F。肝脏。此外，这些研究可能作为应用这一概念在人类肝吸虫病原体引起的其他被忽视的疾病的研究的基础。 本SC 1申请的主要目的是过渡到非SCORE支持机制，并在此过程中确立自己作为独立竞争研究者的地位。 与公共卫生的关系：片形吸虫病是反刍动物的主要疾病，近年来也是人类的主要疾病。我们已经证明FhSAP 2抗原在片形吸虫病的小鼠和兔模型中诱导显著的保护作用。我们建议优化疫苗配方，提高疫苗效力。在将来这样做时，我们将能够在反刍动物中评估优化的疫苗。

DESCRIPTION (provided by applicant): The helminth parasite Fasciola hepatica causes liver fluke disease or fascioliasis, thereby affecting the health of humans, as well as sheep, cattle and goats, among others mammalians. Fascioliasis causes losses in agriculture estimated at >US3.2billion per year. Fascioliasis is also a major zoonotic disease and it is estimated that 17 million people worldwide are infected. The flukicide triclabendazole is the most effective drug to control fascioliasis; however, the cost of treatment and the emergence of drug resistance in sheep infected with F. hepatica suggest a need to develop sustainable strategies, such as vaccination for the control of this disease. However, despite the long-standing research, a vaccine against Fasciola hepatica has not yet been developed. We have identified a novel F. hepatica antigen termed FhSAP2 that when delivered subcutaneously (SC) in Freund's complete adjuvant (FCA) is able to induce partial immunity (>60% reduction in parasite burden and significant reduction of liver damage) in rabbits and mice challenged with F. hepatica metacercariae. FhSAP2 is an 11.5kDa polypeptide belonging to the F. hepatica saposin-like / NK-lysin protein family. We also demonstrated that the immunity induced by the FhSAP2-FCA formulation is associated with high levels of IgG2a antibodies and high levels of IFN3, which are signatures of a Th1 immune response. These results led us to hypothesize that the protection induced by FhSAP2 is mediated by a mechanism linked to CD4+ Th1 cells, which is a new concept in the field of fascioliasis since F. hepatica, like most helminthes, is traditionally associated to Th2 immune responses. The goal of the present study is to study the modulation of T-helper 1, T- helper 2 responses and the protective effect against F. hepatica induced by FhSAP2 with these formulations in the mouse model of fascioliasis. To address this goal we propose to fulfill two specific aims in the 3-year funding period. In specific aim-1 we will study the protective effect of FhSAP2 when delivered SC in adjuvants containing immunostimulatory sequences. Specifically, we will ascertain whether the prime-boost strategy of naove C57BL/6 (H-2b) mice with FhSAP2 in MontanideTM ISM 1312 and ISA 70M or trapped into ISCOMs co- administered with IL-12 or CpG-ODN could mimic or enhance the protection levels obtained with FhSAP2 in CFA. This will lead to an optimization of the vaccine formulation for future studies. In the specific aim-2 we will study how the vaccine formulations used in the aim-1 modulate the T helper 1(Th1)-T helper 2 (Th2) responses. We will look at the Th1/Th2 cytokine profile, the levels of antibody subclasses as well as the activation status of B- and T-cells, macrophages (MX) and dendritic cells (DCs) elicited in naove mice and these parameters will be correlated with the levels of protection obtained in specific aim-1. The outcome will allow us to test our hypothesis of the protective role of Th1 responses in fascioliasis. If the proposed aims are achieved, we not only would have in hand an optimized vaccine formulation to be assayed in ruminants but we will also put forward a new concept of how to induce protection against F. hepatica, which will open the door to further studies to elucidate the immune mechanisms that make possible the Th1 immune responses which are effective against a multicellular fluke like F. hepatica. Moreover, these studies might serve as the basis for the application of this concept in the study of other neglected diseases caused by human liver fluke pathogens. The main objective of PI with this SC1 application is to make a transition to a non-SCORE support mechanism and in doing so establish herself as an independent competitive investigator. PUBLIC HEALTH RELEVANCE: Fasioliasis is a major disease of ruminants and more recently humans. We have demonstrated that an FhSAP2 antigen induces significant protection in a mouse and rabbit model of fascioliasis. We propose to optimize the vaccine formulation and improve the efficacy of the vaccine. In doing so in the future we will be able to evaluate the optimized vaccine in ruminants.