Development of Gamma delta CAR-T cells to target CNS HIV reservoir

项目摘要 HIV在储库组织中的持续存在仍然是实现治疗性治愈的主要障碍， 抗逆转录病毒疗法（ART）。根除中枢神经系统（CNS）中的HIV感染细胞特别重要， 这是一个挑战，因为它是一个免疫豁免的部位，并且很难获得ART药物。而认知 在现代艾滋病毒治疗的时代，艾滋病毒感染者的损害大大减少， 30%的ART抑制的HIV患者仍然表现出HIV相关的神经认知障碍（HAND）， 这表明，即使是低水平的艾滋病毒在大脑中持续存在，也会导致神经系统损伤。因此，功能 迫切需要能够安全地根除CNS储库中复制病毒的治疗策略。新兴 有证据表明，骨髓细胞，包括脑巨噬细胞（Mf），是HIV持续存在的避难所， CNS和HIV感染的巨噬细胞比CD 4 T细胞对CTL更具抗性。这个目标 该项目旨在开发一种通用的γ δ CAR-T平台，具有双重HIVenv/Mf靶向HIV- 中枢神经系统中受感染的骨髓细胞我们的中心假设是，将Mf特异性CAR添加到抗HIV γδ中， CAR-T细胞将能够有效靶向CNS储库中的HIV感染细胞。我们在恒河猴中的数据 猕猴显示γδ CD 4-CAR-T细胞对HIV感染的单核细胞的体外杀伤增强， 颗粒酶B的细胞毒性，已被证明是关键的CTL杀伤HIV感染的原发性 巨噬细胞基于γδ T细胞独特的功能特性，包括（i）充分证明的CTL活性， 在癌症免疫治疗中，（ii）迁移到神经炎症部位的能力，和（iii）先天性抗HIV/SIV CTL 功能;我们假设双重γδ CAR-T细胞将诱导更有效的髓系HIV靶向， 水库在目标1中，我们将开发用于产生基于逆转录病毒和慢病毒载体的γδ CD 4的方法。 CAR-T细胞对SHIV感染的原代Mf和小胶质细胞的靶向增强。在目标2中，我们将测试 病毒血症感染期间优化的γδ CAR-T细胞在CNS中迁移和植入的体内潜力 新的嗜巨噬细胞TF SHIV.D CNS发病机制模型。所提出的研究是有意义的 因为它的成功完成将导致开发出一种通用的γδ CAR-T细胞模型， 在SHIV.D感染的临床相关模型中靶向组织巨噬细胞储库，包括CNS 恒河猴

PROJECT SUMMARY HIV persistence in reservoir tissues remains a major barrier to achieving a therapeutic cure despite effective antiretroviral therapy (ART). Eradicating HIV-infected cells in the central nervous system (CNS) is particularly challenging since it is a site of immune privilege and is poorly accessible to ART drugs. While cognitive impairment among people living with HIV has significantly reduced in the era of modern HIV treatment, about 30% of ART-suppressed HIV patients still display conditions of HIV-associated neurocognitive disorder (HAND), suggesting that even low levels of HIV persisting in the brain can cause neurological damage. Thus, functional cure strategies that can safely eradicate replicating virus the CNS reservoir are urgently needed. Emerging evidence suggests that myeloid cells, including brain macrophages (Mf), are sanctuaries of HIV persistence in the CNS, and that HIV-infected macrophages are more resistant to CTLs than CD4 T cells. The goal of this project is to develop a universal gamma delta (γδ) CAR-T platform with dual HIVenv/Mf-targeting for HIV- infected myeloid cells in the CNS reservoir. Our central hypothesis is that adding Mf specific CAR to anti-HIV γδ CAR-T cells will enable efficient targeting of HIV-infected cells in the CNS reservoir. Our data in rhesus macaques show enhanced in vitro killing of HIV-infected monocytic cells by γδ CD4-CAR-T cells and greater Granzyme B cytotoxicity, which has been shown to be critical for CTL killing of HIV-infected primary macrophages. Based on the unique functional properties of γδ T cells, including (i) well-documented CTL activity in immunotherapy of cancer, (ii) ability to migrate to sites of neuroinflammation, and (iii) innate anti-HIV/SIV CTL functions; we hypothesize that the dual γδ CAR-T cells will induce more effective targeting of myeloid HIV reservoirs. In Aim 1, we will develop approaches for generating retroviral and lentiviral vector-based γδ CD4- CAR-T cells toward enhanced targeting of SHIV-infected primary Mf and microglial cells. In Aim 2, we will test the in vivo potential of the optimized γδ CAR-T cell to migrate and engraft in the CNS during viremic infection with the novel macrophage-tropic TF SHIV.D model of CNS pathogenesis. The proposed research is significant because its successful completion will lead to the development of a universal γδ CAR-T cell model with enhanced targeting of tissue macrophage reservoirs including the CNS in a clinically relevant model of SHIV.D infected rhesus macaques.