Development of Allogeneic CAR T Cell Therapy for a Functional Cure of HIV Infection

摘要：抗逆转录病毒疗法(ART)显著降低了HIV相关的发病率和死亡率(1)。 然而，这并不是一种实际的治愈方法，因为仅通过抗逆转录病毒疗法根除艾滋病毒估计需要60多年的时间。 治疗(1，2)。大量研究支持HIV特异性T细胞反应是有效靶向的关键 以及消除作为慢性感染源的艾滋病毒感染细胞(3-10)。不幸的是，病毒逃逸 而功能有限的病毒特异性效应器CD8T细胞的存在会破坏这些细胞的效力 慢性感染者的反应(11-17)。因此，人们对发展……越来越感兴趣 靶向和消除HIV感染细胞以实现病毒感染的新免疫治疗方法 在缺乏艺术的情况下进行压制，这是一种“功能疗法”。 嵌合抗原受体(CAR)T细胞免疫疗法已显示出巨大的前景 血癌(18-20岁)，现在也证明了减轻恒河猴艾滋病毒/SIV感染的潜力 (21，22)和人源化小鼠(23-28)。我们最近发现，HIV特异性的基于CD4的双重CAR T细胞共同 表达独立的4-1BB和CD28共刺激结构域抑制HIV复制并降低病毒负担 在人源化的小鼠中(23)。然而，目前使用自体T细胞衍生CAR T细胞产品的局限性 (TCPs)，包括耗时和昂贵的制造，患者来源的T细胞不足或功能失调， 以及TCP在患者之间的异质性，是其广泛应用于人类疾病的障碍。 然而，从健康的人类捐赠者中提取的同种异体TCP的开发可能会提供一种现成的 克服这些障碍的治疗选择，以及加快CAR T细胞疗法的使用(29-38)。 不幸的是，输注后受者免疫系统的消除仍然是一个主要障碍(39-41)。 在这里，我们建议利用我们在CAR T细胞生物学(21，23，25，26，42，43)，碱基编辑方面的专业知识 (44-49)，以及人源化的HIV感染小鼠模型(50-58)，以开发同种异体CAR T细胞疗法 对抗艾滋病毒。基于我们的初步数据，我们对CAR T细胞进行了高效的多重基数编辑，我们 假设碱基编辑方法和最佳同种异体供体的识别都将使 抗同种异体排斥反应的基于CD4的CAR-TCP的研制 在没有抗逆转录病毒药物的情况下抑制艾滋病毒。为了验证这一假设，我们提出了以下具体目标： 目的1：确定同种异体T细胞的基因修饰是否可以在体内增强其功能 坚持不懈。 目的2：鉴定与增强相关的同种异体HIV特异性CD4CAR T细胞的特征 持久性和抗病毒疗效。 目的3：比较同种异体和自体HIV特异性CD4CAR T细胞在体内的HIV疗效。 整合了AIM 1和AIM 2‘S的签名，改进了同种异体功能。

Abstract: Antiretroviral therapy (ART) dramatically reduces HIV-associated morbidity and mortality (1). However, it is not a practical cure as eradication of HIV through ART alone is estimated to require over 60 years of treatment (1, 2). Numerous studies support that HIV-specific T cell responses are critical for efficient targeting and elimination of HIV infected cells that are the source of chronic infection (3-10). Unfortunately, viral escape and a limited presence of functional virus-specific effector CD8+ T cells undermine the potency of these responses in chronically infected individuals (11-17). As such, there is growing interest in the development of novel immunotherapeutic approaches to target and eliminate HIV-infected cells to achieve viral suppression in the absence of ART, a “functional cure”. Chimeric antigen receptor (CAR) T cell immunotherapies have demonstrated great promise against blood cancers (18-20), and now also demonstrate the potential to mitigate HIV/SIV infection in rhesus macaques (21, 22) and humanized mice (23-28). We recently showed that HIV-specific Dual CD4-based CAR T cells co- expressing independent 4-1BB and CD28 costimulatory domains restrict HIV replication and reduce viral burden in humanized mice (23). However, current limitations of using autologous T cells to derive CAR T cell products (TCPs), including time-consuming and costly manufacturing, insufficient or dysfunctional patient-derived T cells, and the inter-patient heterogeneity of TCPs, are barriers to their widespread application to human diseases. Development of allogeneic TCPs derived from healthy human donors, could, however, provide an ‘off-the-shelf’ treatment option to overcome these hurdles, as well as accelerate the use of CAR T cell therapies (29-38). Unfortunately, post-infusion elimination by the recipient’s immune system remains a major hurdle (39-41). Here we propose to leverage our expertise in CAR T cell biology (21, 23, 25, 26, 42, 43), base editing (44-49), and a humanized mouse model of HIV infection (50-58) to develop an allogeneic CAR T cell therapy against HIV. Building on our preliminary data applying efficient multiplex base editing to CAR T cells, we hypothesize that both base editing approaches and identification of an optimal allogenic donor will enable the development of an allorejection-resistant CD4-based CAR TCP with enhanced efficacy to eliminate HIV-infected cells and suppress HIV in the absence of ART. To test this hypothesis, we propose the following specific aims: Aim 1: Determine whether genetic modifications to allogeneic T cells can augment their in vivo persistence. Aim 2: Identify characteristics of allogeneic HIV-specific CD4CAR T cells that associate with enhanced persistence and antiviral efficacy. Aim 3: Compare the in vivo HIV efficacy of allogeneic versus autologous HIV-specific CD4CAR T cells, incorporating Aim 1 and 2’s signatures of improved allogeneic functionality.