Regulation of CD8 T Cell Immunity by CDK Inhibitor p27Kip1

描述（由申请人提供）：CD 8 T细胞在防御病毒感染（如HIV、B型和C型肝炎、爱泼斯坦巴尔病毒、巨细胞病毒和沙粒病毒）中发挥关键作用。因此，一个主要的驱动力是开发新的疫苗接种策略，可以引发有效的CD 8 T细胞记忆，并产生有效的保护性免疫，对这些病毒病原体。最近的研究表明，记忆性CD 8 T细胞前体作为效应细胞的一个子集存在于原发性T细胞应答的峰值。然而，与感受态记忆细胞不同，这些记忆前体处于细胞周期停滞的状态;它们缺乏通过稳态增殖自我更新或响应于抗原再调用而增殖的能力，并且提供差的保护性免疫。这些记忆前体在抗原清除后通过持续长达数周的过程开始分化程序，成为有能力的记忆细胞。尽管认识到效应记忆分化是CD 8 T细胞依赖性保护性免疫发展中的限速步骤，但记忆前体中增殖阻滞的诱导和随后逆转的机制尚不清楚。我们使用淋巴细胞性脉络丛脑膜炎病毒（LCMV）小鼠模型的研究表明，细胞周期蛋白依赖性激酶抑制剂p27 Kip 1的遗传缺陷不仅加速了记忆细胞从效应CD 8 T细胞的分化，而且还显著增加了CD 8 T细胞记忆的大小。基于这些发现，我们的中心假设是：p27 Kip 1通过对抗细胞周期的重新进入，阻止记忆性CD 8 T细胞从初级效应CD 8 T细胞分化为有丝分裂的线索。我们进一步假设，记忆CD 8 T细胞分化，包括p27 Kip 1的表达是由上游FOXO转录因子调节。本研究旨在阐明FOXO/p27 Kip 1通路在调节记忆性CD 8 T细胞分化中的关键作用，具体目的如下：1）验证p27 Kip 1在调节记忆性CD 8 T细胞分化中的作用是CD 8 T细胞自主性的假说; 2）验证FOXO转录因子调节记忆性CD 8 T细胞分化的假说。目的1和2将利用互补方法，包括TCR转基因（tg）小鼠、骨髓嵌合体、T细胞中p27 Kip 1或FOXO有条件缺陷的小鼠，和诱导型p27 Kip 1/FOXO tg小鼠，以确定p27 Kip 1和FOXO在调节表型和功能修饰的动力学中的体内作用，所述表型和功能修饰发生在从效应子分化为有能力的记忆的过程中的CD 8 T细胞中细胞这些研究将为记忆细胞分化的机制提供重要的见解，并有可能将p27 Kip 1和FOXO确定为开发新型免疫疗法的分子靶点，以加速和增强疫苗诱导的CD 8 T细胞依赖性保护性免疫对病毒感染的诱导。公共卫生：CD 8 T细胞在防御人类的几种病毒感染方面发挥着关键作用。在这项提案中，我们试图确定关键的分子调节因子（p27 Kip 1和FOXO），可以通过免疫疗法在CD 8 T细胞中进行调节，以增强疫苗诱导的保护性抗病毒免疫力。这些研究应有助于开发有效的疫苗，以保护人类免受病毒引起的疾病。

DESCRIPTION (provided by applicant): CD8 T cells play a crucial role in defense against viral infections like HIV, hepatitis B and C, Epstein Barr virus, cytomegalovirus, and arenaviruses. A major drive therefore has been to develop novel vaccination strategies that can elicit potent CD8 T cell memory and engender effective protective immunity against these viral pathogens. Recent studies have indicated that memory CD8 T cell precursors are present as a subset of effector cells at the peak of the primary T cell response. However, unlike competent memory cells, these memory precursors are in a state of cell cycle arrest; they lack the ability to self renew by homeostatic proliferation or proliferate in response to antigen re-call, and provide poor protective immunity. These memory precursors embark on a program of differentiation to become competent memory cells by a process lasting up to several weeks after antigen clearance. Despite the recognition that effector to memory differentiation is a rate-limiting step in the development of CD8 T cell-dependent protective immunity, the mechanisms underlying the induction and subsequent reversal of proliferative block in memory precursors are poorly defined. Our studies using the mouse model of lymphocytic choriomeningitis virus (LCMV) have shown that genetic deficiency of cyclin dependent kinase inhibitor p27Kip1 not only accelerated the differentiation of memory cells from effector CD8 T cells, but also significantly increased the magnitude of CD8 T cell memory. Based on these findings, our central hypothesis is: p27Kip1 impedes differentiation of memory CD8 T cells from primary effector CD8 T cells by opposing cell cycle re-entry in response to mitogenic cues. We further hypothesize that memory CD8 T cell differentiation including p27Kip1 expression is regulated by the upstream FOXO transcription factors. This proposal focuses on elucidating the key roles of FOXO/p27Kip1 pathway in regulating the differentiation of memory CD8 T cells through these specific aims: 1) To test the hypothesis that effects of p27Kip1 in regulating memory cell differentiation are CD8 T-cell autonomous 2) To test the hypothesis that FOXO transcription factors regulate differentiation of memory CD8 T cells. Aims 1 and 2 will utilize complementary approaches including TCR transgenic (tg) mice, bone marrow chimeras, mice that are conditionally deficient for p27Kip1 or FOXOs in T cells, and inducible p27Kip1/FOXO tg mice to determine the in vivo role of p27Kip1 and FOXOs in regulating the dynamics of phenotypic and functional modifications that occur in CD8 T cells during differentiation from effectors to competent memory cells. These studies would provide critical insight into the mechanisms governing memory cell differentiation, and also have the potential to identify p27Kip1 and FOXO as molecular targets for development of novel immunotherapies to accelerate and enhance the induction of vaccine-induced CD8 T cell-dependent protective immunity against viral infections. PUBLIC HEALTH RELEVENCE: CD8 T cells play a critical role in defense against several viral infections of humans. In this proposal, we seek to identify key molecular regulators (p27Kip1 and FOXO) that can be modulated in CD8 T cells by immunotherapies to enhance vaccine-induced protective anti-viral immunity. These studies should aid in the development of effective vaccines to protect humans against virally induced diseases.