Mechanisms of Immune Memory Regulation by PD-1

描述(由申请人提供)：了解免疫记忆是如何形成的，对于我们设计针对各种传染病和癌症的有效疫苗的能力至关重要。我们已经证明，免疫记忆的定量和定性性质是在感染或免疫后早期决定的，抗原的持续时间是一个关键的决定因素。我们还发现了新的细胞表面标志物，可以成功地预测T细胞扩增初期的记忆结果。此外，使用这些细胞表面标志物来区分记忆命中注定的细胞，我们发现效应器分化的程度与记忆潜力呈负相关。我们的研究涉及全基因组微阵列对记忆命运型和终末分化的效应细胞的比较，有趣的是，记忆命运型效应器在其表面表达更高水平的抑制性受体，程序性死亡-1(PD-1)。这一观察结果促使我们假设PD-1对注定要记忆的CD8 T细胞施加刹车，以防止过度刺激并促进记忆分化。由于PD-1广泛表达于CD4T细胞、CD8T细胞、B细胞、单核细胞、树突状细胞等，我们建立了CD8T细胞特异性的PD-1基因敲除小鼠，专门研究PD-1信号在CD8T细胞记忆反应中的作用。在混合嵌合体环境中，野生型(WT)和PD-1基因敲除(KO)CD8 T细胞存在于同一WT小鼠中，我们发现PD-1 KO CD8 T细胞在急性病毒感染后扩增并分化为类似于WT CD8 T细胞的效应细胞。然而，它们的生存潜力有很大的不同-PD-1 KO CD8 T细胞在病原体清除后死亡更多，并且PD-1 KO CD8 T细胞比WT CD8 T细胞产生最少的记忆库。这个非常集中的R03提案的目标有两个：第一个目标：确定PD-1信号何时起作用来调节记忆细胞的存活。将结合全基因组转录组分析和抗体阻断来确定PD-1信号的时间和来源，以调节急性感染期间免疫记忆的寿命。目的2：确定PD-1信号如何促进记忆细胞的寿命。为此，我们将通过细胞因子信号转导来评估PD-1表达对T细胞增殖的抑制作用。 了解抑制性PD-1信号是否对T细胞的刺激和增殖起到“刹车”作用，从而促进记忆寿命。虽然PD-1在慢性感染中调节免疫病理的作用已被公认，但其在急性感染中的作用尚不清楚。完成后，这些研究将代表着我们在理解调节免疫记忆寿命的因素方面向前迈出的重要一步，并将开辟PD-1信号领域作为疫苗设计的一个新的研究领域，我们希望在后续R01提案的支持下继续进行这一领域。PHS 398/2590(06/09版)页面续格式页面

DESCRIPTION (provided by applicant): Understanding how immunological memory is formed is central to our ability to design efficacious vaccines against a variety of infectious diseases ad cancer. We have shown that quantitative and qualitative properties of immunologic memory are decided early after infection or immunization, and that the duration of antigen is a key determinant. We also identified novel cell surface markers that can successfully predict memory outcome during initial stages of T cell expansion. Moreover, using these cell surface markers to distinguish memory- fated cells, we found that the extent of effector differentiation correlates negatively with memory potential. Our studies involving genome-wide microarray comparisons of memory-fated and terminally differentiated effector cells intriguingly revealed that memory-fated effectors expressed higher levels of inhibitory receptor, programmed death-1 (PD-1) on their surface. This observation prompted us to hypothesize that PD-1 exerts brakes on memory-fated CD8 T cells to prevent overstimulation and promote memory differentiation. Because PD-1 is expressed widely on CD4 T cells, CD8 T cells, B cells, monocytes, dendritic cells, etc., we generated CD8 T cell-specific PD-1 knockout mice, to specifically address the role of PD-1 signaling in CD8 T cell memory responses. In a mixed chimera setting where wild-type (WT) and PD-1 knockout (KO) CD8 T cells were present in the same WT mouse, we found that PD-1 KO CD8 T cells expanded and differentiated into effector cells similar to WT CD8 T cells following an acute viral infection. However, there was a dramatic difference in their survival potential - PD-1 KO CD8 T cells underwent greater death after pathogen clearance and minimal memory reservoirs were generated from PD-1 KO CD8 T cells compared to WT CD8 T cells. The goal of this very focused R03 proposal is two-fold - Aim 1: To determine when PD-1 signals function to regulate memory cell survival. A combination of genome-wide transcriptome analyses and antibody blockade will be employed to determine the timing and source of PD-1 signals in regulating longevity of immune memory during acute infections. Aim 2: To determine how PD-1 signals promote longevity of memory cells. In this specific aim, inhibitory effects of PD-1 expression on T cell proliferation vis a vis cytokine signal transduction will be evaluated to understand whether inhibitory PD-1 signals exert "brakes" on T cell stimulation and proliferation to promote memory longevity. While the role of PD-1 in regulating immunopathology in chronic infections is well established, its function in acute infections is not known. When completed, these studies will represent a major step forward in our understanding of factors regulating longevity of immunologic memory, and will crack open the field of PD-1 signaling as a novel research area for vaccine design, which we hope to pursue under the auspice of a follow-up R01 proposal. PHS 398/2590 (Rev. 06/09) Page Continuation Format Page