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

描述(申请人提供)：轮状病毒(RV)导致全球婴儿腹泻，非洲和亚洲死亡人数最多。新批准的口服RV疫苗在贫困国家的婴儿中的效果未经证实或较低。它们的高昂成本仍然是普遍采用的障碍。在发展中国家，普遍的维生素A缺乏症危害儿童的健康、肠道免疫反应和疫苗效力。世卫组织已将婴儿补充维生素A(VITA)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对轮状病毒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，而维生素A会损害肠道免疫反应。肠道树突状细胞(DC)与关键信号分子(细胞因子、炎症介质)协同作用，将饮食中的维生素A转化为维甲酸(RA)，并在T和B细胞上印迹肠道归巢(CCR9，A4？7)，促进同型转化为IgA，促进IgA的产生。在婴儿和仔猪中，肠道中的粪便或血清IgA抗体或IgA抗体分泌细胞与预防轮状病毒腹泻有关。肠道树突状细胞协调对肠道病原体的肠道免疫反应的关键上调(通过T效应、B细胞和介体)或对食物抗原和共生体的下调(通过免疫调节T细胞和介体，如转化生长因子)。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强对口服疫苗的免疫反应至关重要。为了测试这一点，我们将使用在肠道细胞中复制的减毒轮状病毒(增加促炎作用，减少转化生长因子)作为口服疫苗，并使用维生素A佐剂来增强肠道IgA反应。或者，非复制型轮状病毒样颗粒(VLP)疫苗可能需要带有VITA的TLR信号(多肌苷-多胞酸合成dsRNA，TLR3配体)来模拟轮状病毒dsRNA的作用。以VITA为佐剂，比较VLP口服轮状病毒减毒活疫苗(AIM 1)和VLP口服RV疫苗的免疫应答。疫苗效力将在VAD和VITA全新生诺生菌(Gn)猪身上进行测试，这些猪受到强毒人轮状病毒的攻击(目标3)。新生的Gn仔猪在胃肠道生理、粘膜免疫反应和对人轮状病毒腹泻的易感性方面与婴儿相似。猪像人类一样代谢维生素A，维生素A水平可以被操纵来模拟婴儿的维生素AD。为了阐明免疫机制，我们将评估先天性(DC、NK/NKT细胞、IFNA、炎性细胞因子)和适应性T细胞(TJeff/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婴儿的口服疫苗疗效至关重要。如果成功，在现有RV口服疫苗的婴儿中战略性实施补充VITA(如世卫组织建议的那样)可能是可行的，并可以在发展中国家提供更具成本效益的疫苗(更低、更少的剂量)。一个创新的结果将是VITA佐剂不仅广泛应用于轮状病毒疫苗，而且还应用于其他粘膜疫苗，如艾滋病毒。 相关性：轮状病毒(RV)每年在全世界造成66万儿童死亡，其中大多数(90%)在非洲和亚洲。尽管口服轮状病毒活疫苗获得了许可，但在最需要它们的贫穷国家的婴儿中，它们未经证实或效力较低，而且高昂的成本仍然是普遍采用它们的障碍。在发展中国家，普遍的营养不良和微量营养素缺乏(维生素A等)和多重并发感染损害了儿童的健康、肠道免疫反应和疫苗效力。新的发现表明，维生素A是肠道免疫反应的关键调节因子，缺乏维生素A会损害肠道免疫反应。我们将在维生素A缺乏和维生素A充足的新生灵芝猪身上测试补充维生素A作为活体或非复制型口服轮状病毒疫苗的佐剂。我们的研究将阐明对强毒人类轮状病毒攻击的免疫反应和保护的相关性。新生灵知生菌小猪将被用作疾病模型，因为它们是唯一容易感染人类轮状病毒腹泻的动物。此外，它们在解剖、胃肠生理、营养需求/代谢和粘膜免疫发育方面与婴儿相似。此外，猪代谢视黄醇的能力与人类相当。因此，维生素A对猪免疫系统的影响最接近于人类，而仔猪最好地反映了婴儿对轮状病毒腹泻和疫苗的反应。了解维生素A缺乏对肠道免疫反应的影响对于提高发展中国家维生素A缺乏症婴儿口服疫苗的疗效至关重要。如果成功，立即利用现有轮状病毒疫苗在婴儿中实施补充维生素A(按照世卫组织的建议)可能是可行的，并可以在发展中国家提供更具成本效益的疫苗(更低、更少剂量)。维生素A佐剂可能普遍适用，不仅适用于轮状病毒疫苗，也适用于其他人类黏膜疫苗，如艾滋病毒。

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.