Mechanisms of in vivo protection from HIV infection

描述（由申请人提供）：我们建议开发和评估两种旨在增强HIV感染者免疫反应的治疗方法，作为“概念验证”。首先，我们将评估增强HIV-1感染者的HIV-1特异性免疫的有效性，通过用编码各种HIV-1特异性TCR的慢病毒载体的混合物转导他们的CD 34+造血干细胞（HSC）或CD 8 + T淋巴细胞，这些TCR限制于受体的I类HLA，在自体移植后诱导分化为有效的HIV-1特异性CTL。具体而言，我们将研究是否可以通过将编码源自有效HIV特异性CTL的TCR的α和β链的基因递送到CD 34 + HSC或CD 8 + T淋巴细胞中来产生功能性HIV特异性CD 8 + CTL。为此，我们将在体外和体内的功能，人类HIV特异性CTL，克隆出TCR α和β链的cDNA从最有效的HIV特异性CD 8 + CTL克隆，并将它们插入到新的慢病毒载体。然后，我们将确定这些载体将克隆的TCR基因引入人造血前体细胞或CD 8+淋巴细胞的能力，并将其编程以分化为HIV特异性CD 8 + CTL，这些CTL显示出体外和体内识别和消除HIV感染细胞的能力。其次，我们将研究最近发现的与HIV免疫的相关性，即阻断PD-1/PD-L1抑制途径恢复“耗尽的”LCMV特异性CTL的功能活性。我们建议通过密切相关和协调的目标来实现这一目标：1）评价编码源自有效的人HIV-1特异性CTL的TCR α和β链的慢病毒载体在转导人CD 8+淋巴细胞后产生功能性人HIV特异性CTL的能力，2）为了确定表达HIV特异性TCR α和β链的慢病毒载体是否可以编程人CD 34 + HSC以分化成功能性HIV特异性CTL，和3）研究通过用Fc-PD 1或Ig-PD 1融合蛋白处理阻断PD 1/PD-L1抑制途径是否增加HIV特异性CTL的体外和体内功能。这些方法通过重定向免疫应答以产生识别免疫保护性表位的CTL，并通过阻断PD-1（由活化的CTL表达的抑制性分子）的活化，增强HIV特异性CTL的功能，从而提供了用于增加HIV特异性免疫的新模式。

DESCRIPTION (provided by applicant): We propose to develop and evaluate as a "proof-of-concept" two therapeutic approaches designed to augment the immune response of HIV-infected individuals. First, we will evaluate the effectiveness of augmenting HIV-1-specific immunity in HIV-1-infected individuals by transducing their CD34+ hematopoietic stem cells (HSC) or CD8+ T lymphocytes with a cocktail of lentiviral vectors encoding various HIV-1-specific TCRs restricted to the recipient's Class I HLA that will induce differentiation into effective HIV-1-specific CTLs after autologous transplantation. Specifically we will examine whether functional HIV-specific CD8+ CTLs can be generated by delivering genes encoding the alpha and beta chain of TCRs derived from potent HIV-specific CTLs into CD34+ HSC or CD8+ T lymphocytes. To this end we will characterize the in vitro and in vivo function of human HIV-specific CTLs, clone out the TCR alpha and beta chain cDNA from the most potent HIV-specific CD8+ CTL clones and insert them into novel lentiviral vectors. We will then determine the capacity of these vectors to introduce the cloned TCR genes into human hematopoietic precursor cells or CD8+ lymphocytes and program them to differentiate into HIV-specific CD8+ CTLs that display the in vitro and in vivo capacity to recognize and eliminate HIV-infected cells. Second, we will examine the relevance to HIV immunity of the recent finding that blocking the PD-1/PD-L1 inhibitory pathway restores functional activity to "exhausted" LCMV-specific CTLs. We propose to achieve this goal via closely related and coordinated aims: 1 )To evaluate the capacity of lentiviral vectors encoding TCR alpha and beta chains derived from potent human HIV-1-specific CTLs to generate functional human HIV-specific CTL after transduction of human CD8+ lymphocytes, 2) To determine whether lentiviral vectors expressing HIV- specific TCR alpha and beta chains can program human CD34+ HSC to differentiate into functional HIV- specific CTLs, and 3) To investigate whether blockade of the PD1/PD-L1 inhibitory pathway by treatment with Fc-PD1 or lg-PD1 fusion proteins increases the in vitro and in vivo function of HIV-specific CTLs. These approaches provide new modalities for increasing HIV-specific immunity by redirecting the immune response to produce CTLs that recognize immunoprotective epitopes and by blocking activation of PD-1, an inhibitory molecule expressed by activated CTL, enhances the function of HIV-specific CTLs.