Leukocyte Signaling in the Elderly and Vaccine Immunogenicity

描述（由申请方提供）：流感病毒感染继续导致老年人（65岁或以上）的死亡率不成比例，尽管该人群每年接受灭活三价流感疫苗（TIV）的免疫接种水平较高。TIV在老年人中的疗效和免疫原性总体上显著低于年轻成人，尽管由于尚不清楚的原因存在显著的个体差异。TIV含有血凝素（HA）和神经氨酸酶（NA）主要病毒表面蛋白并且是无佐剂的，其主要通过诱导针对HA的中和抗体来提供保护。由TIV诱导的流感特异性CD 4 T细胞应答可能有助于抗体应答，并且与CD 8 T细胞应答结合，可能在尽管接种疫苗但仍发生感染的情况下增强病毒清除。目前还不清楚在多大程度上纳入有效的佐剂，如阳离子脂质/DNA复合物（CLDC）的适应性免疫，可以提高免疫原性的老年人TIV和其他疫苗。一项由Juvaris Biotherapeutics，Inc.申办的II期人体研究，在472名老年男性和女性中评估了单次肌肉注射TIV/CLDC的安全性、有效性和剂量要求，以获得最佳的流感特异性体液和T细胞应答。作为本研究的一部分，GLP认证的流感特异性抗体以及基线和接种后多个时间点的CD 4和CD 8 T细胞应答测定正在进行中。这些免疫原性结果的可用性，连同多等份冻存PBMC的沿着，为确定本项目中老年人疫苗免疫原性的预测因子提供了独特的机会。中心假设是，在一些老年人中，对TIV的体液和T细胞免疫原性受损反映了抗原呈递细胞（APC）、B细胞和T细胞的信号传导的内在限制，并且这些限制反过来是微小RNA（miRNA）改变mRNA表达的结果。将通过确定这些老年受试者的TIV或TIV/CLDC疫苗接种后的免疫原性是否可以通过其单核细胞、B细胞和T细胞在细胞因子受体（例如，IL-21、先天免疫配体，例如，CpG DNA和抗原，即，B细胞和T细胞受体，如通过改进的磷酸流式细胞术测定所评估的。同时，将通过多色流式细胞术测定单核细胞、B细胞以及CD 4和CD 8 T细胞的主要亚群的频率，这在磷酸化流式细胞术结果的解释中将是重要的。然后将评价作为TIV或TIV/CLDC免疫原性最稳健预测因子的白细胞亚群和刺激物的基础和刺激依赖性mRNA和miRNA表达水平，重点关注已知或可能影响信号传导的变化。总之，这些研究将确定白细胞亚群信号传导水平在老年人中产生对TIV和TIV/CLDC的适应性免疫应答中的预测价值和生物学重要性。他们还将提供新的机制的见解，具体的mRNA和miRNA在调节信号和激活诱导的基因表达参与疫苗免疫原性的作用。 公共卫生相关性（由申请人提供）：在高风险人群（如老年人）中使用疫苗预防感染（如流感）的一个主要局限性是，很难预测免疫接种是否能有效地充分增强免疫应答，以保护该个体免受传染病的侵害。这项研究将确定免疫系统白色血细胞功能的测试，可用于预测老年人接种流感疫苗是否可能充分刺激免疫系统，为该个体提供免受感染的保护。这些测试将有助于更有效地使用现有的疫苗，也将有助于开发一种新的，更有效的流感疫苗，该疫苗使用佐剂，一种添加到疫苗中以增强免疫反应的物质。

DESCRIPTION (provided by applicant): Influenza viral infection continues to cause disproportionate mortality in the elderly (those 65 years of age or older) despite a high level of annual immunization of this population with inactivated trivalent influenza vaccine (TIV). The efficacy and immunogenicity of TIV in the elderly overall is substantially lower than in younger adults, although there is substantial individual variation for reasons that are not well understood. TIV, which contains the hemagglutinin (HA) and neuraminidase (NA) major viral surface proteins and is unadjuvanted, provides protection mainly by inducing neutralizing antibodies to HA. Influenza-specific CD4 T-cell responses induced by TIV may contribute to the antibody response and, in conjunction with CD8 T-cell responses, may enhance viral clearance in the event that infection occurs despite vaccination. It is unclear to what extent the inclusion of potent adjuvants for adaptive immunity, such as cationic lipid/DNA complexes (CLDC) could boost immunogenicity in the elderly to TIV and other vaccines. A phase II human study sponsored by Juvaris Biotherapeutics, Inc., is evaluating the safety, efficacy and dose requirement of a single intramuscular dose of TIV/CLDC for optimal influenza-specific humoral and T-cell responses in 472 elderly men and women. As part of this study, GLP-certified assays of influenza-specific antibody and CD4 and CD8 T-cell responses at baseline and at multiple time points post-vaccination are in progress. The availability of these immunogenicity results, along with multiple aliquots of cryopreserved PBMCs, provides a unique opportunity to determine predictors of vaccine immunogenicity in the elderly in this project. The central hypothesis is that impaired humoral and T-cell immunogenicity to TIV in some of the elderly reflects intrinsic limitations in the signaling of antigen-presenting cells (APCs), B cells, and T cells, and that these limitations are, in turn, the result of alterations of mRNA expression by microRNAs (miRNAs). This hypothesis will be tested by determining if immunogenicity after TIV or TIV/CLDC vaccination of these elderly subjects can be predicted by the capacity of their monocytes, B cells, and T cells to signal after engagement of receptors for cytokines, e.g., IL-21, innate immune ligands, e.g., CpG DNA, and antigen, i.e., the B-cell and T-cell receptors, as assessed by an improved phospho-flow cytometric assay. In parallel, the frequency of the major subsets of monocytes, B cells, and CD4 and CD8 T- cells will be determined by polychromatic flow cytometry, which will be important in the interpretation of the phospho-flow results. The leukocyte subsets and the stimuli that are most robust predictors of TIV or TIV/CLDC immunogenicity will then be evaluated for their basal and stimulus-dependent levels of mRNA and miRNA expression, focusing on changes that are known to or are likely to influence signaling. Together, these studies will define the predictive value and biological importance of the level of leukocyte subset signaling in generating adaptive immune responses to TIV and TIV/CLDC in the elderly. They will also provide new mechanistic insights as to the role that specific mRNAs and miRNAs play in regulating signaling and activation-induced gene expression involved in vaccine immunogenicity. PUBLIC HEALTH RELEVANCE (provided by applicant): A major limitation in using vaccines to prevent infections, such as influenza, in high-risk populations, such as the elderly, is that it is difficult to predict whether an immunization will be effective in sufficiently boosting the immune response to protect that individual from the infectious disease. This research will identify tests of the function of white blood cells of the immune system that can be used to predict whether vaccination of an elderly individual for influenza will be likely to sufficiently stimulate the immune system to provide that individual with protection from infection. These tests will help use current vaccines more effectively, and will also help in the development of a new, more potent vaccine for influenza that uses an adjuvant, a substance added to vaccines to boost the immune response.