Department of Chemistry and Biochemistry, Brigham Young University, Provo, Utah, USA.
Laboratory of Viral Diseases, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, Maryland, USA.
J Virol. 2020 May 4;94(10). doi: 10.1128/JVI.00190-20.
The major obstacle to a cure for HIV infection is the persistence of replication-competent viral reservoirs during antiretroviral therapy. HIV-specific chimeric antigen receptor (CAR) T cells have been developed to target latently infected CD4 T cells that express virus either spontaneously or after intentional latency reversal. Whether HIV-specific CAR-T cells can recognize and eliminate the follicular dendritic cell (FDC) reservoir of HIV-bound immune complexes (ICs) is unknown. We created HIV-specific CAR-T cells using human peripheral blood mononuclear cells (PBMCs) and a CAR construct that enables the expression of CD4 (domains 1 and 2) and the carbohydrate recognition domain of mannose binding lectin (MBL) to target native HIV Env (CD4-MBL CAR). We assessed CAR-T cell cytotoxicity using a carboxyfluorescein succinimidyl ester (CFSE) release assay and evaluated CAR-T cell activation through interferon gamma (IFN-γ) production and CD107a membrane accumulation by flow cytometry. CD4-MBL CAR-T cells displayed potent lytic and functional responses to Env-expressing cell lines and HIV-infected CD4 T cells but were ineffective at targeting FDC bearing HIV-ICs. CD4-MBL CAR-T cells were unresponsive to cell-free HIV or concentrated, immobilized HIV-ICs in cell-free experiments. Blocking intercellular adhesion molecule-1 (ICAM-1) inhibited the cytolytic response of CD4-MBL CAR-T cells to Env-expressing cell lines and HIV-infected CD4 T cells, suggesting that factors such as adhesion molecules are necessary for the stabilization of the CAR-Env interaction to elicit a cytotoxic response. Thus, CD4-MBL CAR-T cells are unable to eliminate the FDC-associated HIV reservoir, and alternative strategies to eradicate this reservoir must be sought. Efforts to cure HIV infection have focused primarily on the elimination of latently infected CD4 T cells. Few studies have addressed the unique reservoir of infectious HIV that exists on follicular dendritic cells (FDCs), persists during antiretroviral therapy, and likely contributes to viral rebound upon cessation of antiretroviral therapy. We assessed the efficacy of a novel HIV-specific chimeric antigen receptor (CAR) T cell to target both HIV-infected CD4 T cells and the FDC reservoir Although CAR-T cells eliminated CD4 T cells that express HIV, they did not respond to or eliminate FDC bound to HIV. These findings reveal a fundamental limitation to CAR-T cell therapy to eradicate HIV.
治愈 HIV 感染的主要障碍是在抗逆转录病毒治疗期间复制能力强的病毒储库的持续存在。已经开发了 HIV 特异性嵌合抗原受体 (CAR) T 细胞，以靶向潜伏感染的 CD4 T 细胞，这些细胞要么自发表达病毒，要么在有意逆转潜伏后表达病毒。HIV 特异性 CAR-T 细胞是否能够识别和消除 HIV 结合免疫复合物 (IC) 的滤泡树突状细胞 (FDC) 储库尚不清楚。我们使用人外周血单核细胞 (PBMC) 和一种 CAR 构建体创建了 HIV 特异性 CAR-T 细胞，该构建体能使 CD4（域 1 和 2）和甘露糖结合凝集素 (MBL) 的碳水化合物识别域的表达靶向天然 HIV Env（CD4-MBL CAR）。我们使用羧基荧光素琥珀酰亚胺酯 (CFSE) 释放测定法评估了 CAR-T 细胞的细胞毒性，并通过流式细胞术评估了 CAR-T 细胞通过干扰素 γ (IFN-γ) 产生和 CD107a 膜积累的激活。CD4-MBL CAR-T 细胞对表达 Env 的细胞系和 HIV 感染的 CD4 T 细胞表现出强大的溶细胞和功能反应，但对携带 HIV-IC 的 FDC 无效。CD4-MBL CAR-T 细胞对无细胞 HIV 或无细胞实验中固定的浓缩 HIV-IC 无反应。阻断细胞间黏附分子-1 (ICAM-1) 抑制了 CD4-MBL CAR-T 细胞对表达 Env 的细胞系和 HIV 感染的 CD4 T 细胞的细胞溶解反应，表明细胞黏附分子等因素对于 CAR-Env 相互作用的稳定对于引发细胞毒性反应是必要的。因此，CD4-MBL CAR-T 细胞无法消除与 FDC 相关的 HIV 储库，必须寻求其他消除该储库的策略。治愈 HIV 感染的努力主要集中在消除潜伏感染的 CD4 T 细胞上。很少有研究涉及存在于滤泡树突状细胞 (FDC) 上的感染性 HIV 的独特储库，该储库在抗逆转录病毒治疗期间持续存在，并可能导致抗逆转录病毒治疗停止后病毒反弹。我们评估了一种新型 HIV 特异性嵌合抗原受体 (CAR) T 细胞靶向 HIV 感染的 CD4 T 细胞和 FDC 储库的功效。尽管 CAR-T 细胞消除了表达 HIV 的 CD4 T 细胞，但它们对与 HIV 结合的 FDC 无反应或不消除。这些发现揭示了 CAR-T 细胞疗法根除 HIV 的一个基本局限性。
