Vitamin A adjuvant to enhance gut immunity and rotavirus vaccines in neonates

描述（由申请人提供）：轮状病毒（RV）在世界范围内引起婴儿腹泻，大多数死亡发生在非洲和亚洲。新批准的口服RV疫苗在贫困国家的婴儿中具有未经证实或较低的效力。它们的高成本仍然是普遍采用的障碍。在发展中国家，普遍存在的维生素A缺乏症（VAD）危及儿童健康、肠道免疫反应和疫苗有效性。世卫组织已将婴儿维生素A补充与免疫规划联系起来，以减少VAD并增加疫苗覆盖率。补充维生素a对RV疫苗的影响尚不清楚。我们的创新和新颖策略是使用补充维生素a作为新生儿口服疫苗的佐剂。新的发现表明，维生素a是肠道免疫反应的关键调节因子，而VAD损害肠道免疫反应。肠道树突状细胞（dc）将膳食中的维生素a转化为维甲酸（RA）， dc和/或RA与关键信号分子（细胞因子、炎症介质）协同作用，在T细胞和B细胞上印记肠道归巢（CCR9, a4¿7），促进同型转换为IgA并增强IgA的产生。在婴儿和仔猪中，粪便或血清IgA抗体或肠道中IgA抗体分泌细胞与预防RV腹泻相关。肠道dc协调肠道免疫反应的关键上调（通过Teffector， B细胞和介质）对肠道病原体或下调（通过免疫调节性T细胞和介质如TGF¿）对食物抗原和共生体。我们假设由炎症或TLR信号组成的“危险信号”对于激活dc以克服肠道下调和增加对口服疫苗的免疫反应至关重要。为了验证这一点，我们将使用在肠细胞中复制的减毒RV（增加促炎，减少TGF -细胞因子）作为口服疫苗与VitA佐剂一起增强肠道IgA反应。另外，非复制的轮状病毒样颗粒（VLP）疫苗可能需要TLR信号（多肌苷-多胞酸合成dsRNA， TLR3配体）和VitA来模拟RV dsRNA的作用。使用VitA作为佐剂，我们将比较口服RV减毒活疫苗（Aim 1）和VLP （Aim 2）的免疫应答。疫苗的效力将在VAD和充满维生素a的新生猪（Gn）中进行测试，这些猪受到毒性人类RV的攻击（Aim 3）。新生儿Gn仔猪在胃肠道生理、粘膜免疫反应和对人RV腹泻的易感性方面与婴儿相似。猪像人一样代谢维生素a，维生素a的水平可以模拟婴儿的VAD。为了阐明免疫机制，我们将评估先天性（DC， NK/NKT细胞，IFNa，炎症因子）和适应性T （Teff/Treg比率）和B细胞（IgA）肠道免疫反应，包括肠道DC和细胞因子谱及其与保护的相关性。了解VAD对肠道免疫的影响对于提高发展中国家VAD婴儿口服疫苗的疗效至关重要。如果成功，在婴儿中（按照世卫组织的建议）对现有RV口服疫苗进行补充维生素a的战略实施可能是可行的，并且可以在发展中国家提供更具成本效益的疫苗（剂量更低、更少）。一个创新的结果将是维生素a佐剂的普遍应用，不仅适用于RV疫苗，也适用于其他粘膜疫苗，如HIV疫苗。

DESCRIPTION (provided by applicant): Rotavirus (RV) causes diarrhea in infants worldwide with most deaths in Africa and Asia. Newly licensed oral RV vaccines have unproven or lower efficacies in infants in impoverished countries. Their high costs remain obstacles to universal adoption. In developing countries, widespread vitamin A deficiencies (VAD) compromise children's health, gut immune responses and vaccine effectiveness. WHO has linked vitamin A (VitA) supplementation in infants to immunization programs to reduce VAD and increase vaccine coverage. The impact of VitA supplementation on RV vaccines is unknown. Our innovative and novel strategy is to use supplemental VitA as an adjuvant with oral vaccines in neonates. New discoveries have revealed that VitA is a key regulator of intestinal immune responses and that VAD impairs gut immune responses. Gut dendritic cells (DCs) convert dietary VitA to retinoic acid (RA) and DCs and/or RA in concert with key signaling molecules (cytokines, inflammatory mediators), imprint gut homing (CCR9, a4¿7) on T and B cells, promote isotype switching to IgA and enhance IgA production. In infants and piglets, fecal or serum IgA antibodies or IgA antibody secreting cells in the gut correlate with protection against RV diarrhea. Gut DCs orchestrate critical upregulation of intestinal immune responses (via Teffector, B cells and mediators) to enteric pathogens or downregulation (via immunoregulatory T cells and mediators like TGF¿) to food antigens and commensals. We hypothesize that "danger signals" consisting of inflammatory or TLR signaling are essential to activate DCs to overcome intestinal downregulation and increase immune responses to oral vaccines. To test this, we will use attenuated RV that replicates in enterocytes, (increase proinflammatory, decrease TGF¿ cytokines) as oral vaccine with VitA adjuvant to enhance gut IgA responses. Alternatively nonreplicating rotavirus-like-particle (VLP) vaccines may require TLR signals (polyinosine-polycytidylic acid synthetic dsRNA, TLR3 ligand) with VitA to mimic the effect of RV dsRNA. Using VitA as adjuvant, we will compare immune responses to live attenuated (Aim 1) with VLP (Aim 2) oral RV vaccines. Vaccine efficacy will be tested in VAD versus VitA replete neonatal gnotobiotic (Gn) pigs challenged with virulent human RV (Aim 3). Neonatal Gn piglets mimic infants in their gastrointestinal physiology, mucosal immune responses and susceptibility to human RV diarrhea. Pigs metabolize VitA like humans and VitA levels can be manipulated to mimic VAD in infants. To elucidate immune mechanisms, we will assess innate (DCs, NK/NKT cells, IFNa, inflammatory cytokines) and adaptive T (Teff/Treg ratio) and B cell (IgA) intestinal immune responses, including gut DC and cytokine profiles and their correlation with protection. Understanding the impact of VAD on gut immunity is critical to improve oral vaccine efficacy in VAD infants in developing countries. If successful, strategic implementation of supplemental VitA in infants (as recommended by WHO) with existing RV oral vaccines may be feasible and could provide more cost-effective vaccines (lower, fewer doses) in developing countries. An innovative outcome would be universal application of VitA adjuvants not only to RV vaccines, but also to other mucosal vaccines such as for HIV. RELEVANCE: Rotavirus (RV) causes 660,000 childhood deaths annually worldwide, with most (90%) in Africa and Asia. Although live oral rotavirus vaccines are licensed, their unproven or lower effectiveness in infants in impoverished countries, where they are most needed, and high costs remain obstacles to their universal adoption. In developing countries, widespread malnutrition with micronutrient deficiencies (vitamin A, etc) and multiple concurrent infections compromise children's health, gut immune responses and vaccine effectiveness. New discoveries have revealed that vitamin A is a key regulator of intestinal immune responses and that vitamin A deficiency impairs gut immune responses. We will test supplemental vitamin A as adjuvant with live or nonreplicating oral rotavirus vaccines in vitamin A deficient versus vitamin A replete neonatal gnotobiotic pigs. Our studies will elucidate the immune responses and correlates of protection to challenge with virulent human rotavirus. Neonatal gnotobiotic piglets will be used as a disease model because they are the only animal susceptible to human rotavirus diarrhea. Moreover they are similar to infants in anatomy, gastrointestinal physiology, nutrient requirements/metabolism, and development of mucosal immunity. Also swine metabolize retinol comparable to humans. Thus the effects of vitamin A on the porcine immune system most closely mimic that in humans, and piglets best mirror responses of infants to rotavirus diarrhea and vaccines. An understanding of impacts of vitamin A deficiency on gut immune responses is critical to improve oral vaccine efficacy in vitamin A compromised infants in developing countries. If successful, immediate implementation of supplemental vitamin A in infants (as recommended by WHO) with the existing RV vaccines may be feasible and could provide more cost-effective vaccines (lower, fewer doses) in developing countries. Vitamin A adjuvants may be universally applicable, not only to rotavirus vaccines, but also to other mucosal vaccines for humans such as HIV.