Enhancement of HIV-specific CD4+ T cell function by blockade of the PD-1 pathway

描述(申请人提供)：在慢性人类免疫缺陷病毒(HIV)感染期间，HIV特异性T细胞无法控制病毒复制，部分原因是它们失去了增殖、产生细胞因子和裂解受感染目标细胞的能力。因此，HIV特异性T细胞的功能恢复对于限制疾病进展至关重要，也可以与治疗性疫苗接种结合使用，以增加疫苗产生的HIV特异性T细胞反应的功能。大多数试图通过阻断或刺激细胞表面标志物来恢复功能失调的HIV特异性T细胞的活性的尝试都没有成功。然而，最近，CD28家族的抑制性受体死亡蛋白1(PD-1)的信号传导被证明显著增加了慢性HIV感染者和淋巴细胞性脉络膜脑膜炎病毒(LCMV)感染小鼠的病毒特异性CD8 T细胞的频率和功能。LCMV是慢性病毒感染导致T细胞功能障碍的模型系统。我们的初步数据显示，在慢性HIV感染期间，PD-1在HIV特异的CD4T细胞上也被特异性上调，并且阻断也增加了HIV特异的CD4T细胞的增殖。然而，PD-1阻断对HIV特异性CD4T细胞细胞因子的产生和成熟的影响知之甚少。PD-1途径的阻断确实导致慢性感染小鼠的LCMV病毒载量大幅下降，但由于没有经过验证的慢性HIV感染的小鼠模型，因此很难确定同样的情况是否适用于HIV感染。我们建议使用体外和体内双管齐下的方法来确定阻断PD-1连接是否是恢复或增强慢性HIV感染者HIV特异性CD4T细胞功能的可行治疗方法。我们假设，阻断HIV感染者外周血单个核细胞(PBMC)中PD-1的结合将在体外特异性地增强HIV特异性CD4T细胞的功能，同时阻断PD-1连接并激活共刺激分子CD28、4-1BB或OX40的单抗的“鸡尾酒”将导致比单独阻断PD-1更大的功能增强。此外，我们将使用一种新的人源化Rag2-/-？C/-小鼠(RAG-HU)HIV感染模型来检测阻断PD-1结扎对体内HIV病毒载量和CD4计数的影响。Rag2-/-3c/-小鼠与人CD34造血干细胞重组，CD34干细胞分化为能够增强免疫反应的人免疫细胞。我们最近证明，这些小鼠对艾滋病毒感染是允许的。我们假设，阻断HIV感染的RAG-HU小鼠中的PD-1结合将加强对HIV复制的控制，从而在体内产生较低的HIV病毒载量和较高的CD4T细胞计数。本研究的具体目的是：1)通过用抗PD-1配体单抗和可溶性PD-1阻断PD-1通路，增强慢性HIV感染者PBMC中HIV特异性CD4T细胞的功能。2)验证HIV慢性感染RAG-Hu小鼠模型的有效性，并利用该模型研究阻断PD-1途径对HIV体内复制的影响。 与公共卫生的相关性：据估计，人类免疫缺陷病毒(艾滋病毒)感染了4000万人，并在2006年造成全世界290万人死亡。开发基于免疫的疗法以增加HIV特异性CD4T细胞的功能对于控制疾病至关重要。PD-1途径阻断疗法可以减缓疾病进展，并与治疗性疫苗接种结合使用，以增加疫苗产生的HIV特异性CD4T细胞反应的功能。

DESCRIPTION (provided by applicant): During chronic human immunodeficiency virus (HIV) infection, HIV-specific T cells fail to control viral replication, in part because they lose the ability to proliferate, produce cytokines, and lyse infected target cells. Functional restoration of HIV-specific T cells is therefore essential for limiting disease progression, and could also be used in combination with therapeutic vaccination to increase the functionality of vaccine generated HIV-specific T cell responses. Most attempts to restore activity in dysfunctional HIV-specific T cells by blocking or stimulating cell surface markers have not been successful. Recently, however, blocking signaling of death protein 1 (PD-1), an inhibitory receptor in the CD28 family, was shown to dramatically increase the frequency and function of virus-specific CD8+ T cells both in subjects with chronic HIV infection and in mice infected with lymphocytic choriomeningitis virus (LCMV), a model system for T cell dysfunction induced by chronic viral infection. Our preliminary data demonstrate that PD-1 is also specifically up-regulated on HIV-specific CD4+ T cells during chronic HIV infection and that blockade increases HIV-specific CD4+ T cell proliferation. However little is known about the effect of PD-1 blockade on HIV-specific CD4+ T cell cytokine production and maturation. PD-1 pathway blockage did result in a substantial decrease in LCMV viral load in chronically infected mice, but since there is no validated mouse model for chronic HIV infection, it has been difficult to determine whether the same would be true for HIV infection. We propose to use a two pronged in vitro and in vivo approach to determine if blockade of PD-1 ligation is a viable therapeutic approach for restoring or augmenting HIV-specific CD4+ T cell function in subjects with chronic HIV infection. We hypothesize that blockade of PD-1 binding in HIV-infected subject's peripheral blood mononuclear cells (PBMC) will specifically enhance HIV-specific CD4+ T cell function in vitro and that "cocktails" of mAbs which simultaneously block PD- 1 ligation and activate costimulatory molecules CD28, 4-1BB or OX40 will lead to even greater enhancement of function than PD-1 blockade alone. In addition we will examine the effects of blockade of PD-1 ligation on HIV viral load and CD4 count in vivo using a novel humanized Rag2-/-?c/- mouse (RAG-hu) model of HIV infection. Rag2-/-3c/- mice are reconstituted with human CD34 hematopoietic stems cells which differentiate into human immune cells capable of mounting immune responses. We have recently demonstrated that these mice are permissive to HIV infection. We hypothesize that blockade of PD-1 binding in HIV-infected RAG-hu mice will enhance control of HIV replication resulting in lower HIV viral loads and higher CD4+ T cell counts in vivo. The specific aims of this study are: 1)To enhance HIV-specific CD4+ T cell function in PBMC from chronically HIV-infected subjects by blocking the PD-1 pathway using anti PD-1 ligand mAbs and soluble PD-1. 2) To validate the RAG-hu mouse model of chronic HIV infection for the study of HIV-specific T cell responses and use it to investigate the effects of blocking the PD-1 pathway on HIV replication in vivo. PUBLIC HEALTH RELEVANCE: The human immunodeficiency virus (HIV) is estimated to have infected 40 million people and to be responsible for 2.9 million deaths worldwide in 2006. Development of immune-based therapies to increase the function of HIV-specific CD4+ T cells are vital for controlling the disease. PD-1 pathway blocking therapies could slow disease progression and be used in combination with therapeutic vaccination to increase the functionality of vaccine generated HIV-specific CD4+ T cell responses.