AAV-delivered HIV inhibitors for SHIV therapy

项目总结 在美国，超过10%的新感染病例来自受污染的针头共用 注射毒品者(PWID)。重要的是，只有大约50%的男性和57%的女性 组对病毒血症有抑制作用。尽管缺乏对HIV-1进行消毒的疗法，但抗逆转录病毒药物疗法 (抗逆转录病毒药物)有效抑制HIV-1感染者的病毒复制。艺术的一个局限是 这些小分子药物不能消除病毒库。广谱中和抗体 (BNAbs)可以补充ART，并用于减少病毒库。多项研究表明 单剂bNAb可以减少HIV-1感染者的病毒血症，并导致病毒反弹 随着bNAb浓度的降低而发生。腺相关病毒(AAV)载体可以提供 能够维持病毒浓度的bNAbs长期表达的手段 通过肌肉接种抑制。然而，我们和其他人已经表明， 抗bNAbs的抗药抗体(ADA)限制了它们的整体表达。在这里，我们展示了我们已经取得了 显著的进步克服了这种宿主免疫反应。首先是通过利用免疫检查点 来调节免疫系统的途径。我们观察到艾滋病毒-1的浓度增加了21倍 恒河猴与编码AAV载体共接种时的bNab 10-1074 恒河猴PD-L1。第二，我们开发了一种新型的HIV-1进入抑制剂eCD4-Ig，它 当从AAV载体表达时，恒河猴往往更能耐受。与bNAbs不同，我们 已经证明编码eCD4-Ig的AAV载体可以在猕猴体内表达这种抑制物一年多 而且随着时间的推移，ADA对eCD4-Ig的反应会减少。因为eCD4-Ig中和了所有的HIV-1病毒， 艾滋病毒-2和SIV是分离的，很难逃脱，如果将其纳入治疗策略，它可能是有用的。 我们的初步研究表明，低浓度AAV表达的eCD4-Ig可以抑制SHV感染 恒河猴感染了两年，但仍可检测到病毒血症。这项建议结合了我们的AAV 确定AAV表达的抑制剂是否可以抑制AAV的单一策略的进展 希沃克病毒的感染和减少了病毒库。在目标1中，我们将评估 SHIV感染恒河猴ART解除后AAV携带的eCD4-Ig和10-1074的联合检测 猕猴。在目标2中，我们将确定是否通过AAV抑制已建立的SHV感染- 与ART相比，所提供的eCD4-Ig和10-1074产生了数量上不同的病毒库。在……里面 目的3，通过研制一种不可逆的“杀伤开关”来提高AAV基因治疗的安全性。 不表达转基因。

PROJECT SUMMARY More 10% of the new infections in the United States were from contaminated needle sharing among people who inject drugs (PWID). Importantly, only about 50% of men and 57% of women of the PWID group had suppressed viremia. Despite the lack of a sterilizing HIV-1 cure, antiretroviral drug therapies (ART) effectively suppress viral replication in people living with HIV-1. One limitation of ART is that these small molecule drugs cannot eliminate the viral reservoir. Broadly neutralizing antibodies (bNAbs) could supplement ART and be used to reduce the viral reservoir. Several studies have shown that a single dose of a bNAb can decrease viremia in HIV-1 infected individuals, with viral rebound occurring as the bNAb concentration decreases. Adeno-associated virus (AAV) vectors may provide the means for long-term expression of bNAbs at concentrations capable of maintaining viral suppression via intramuscular inoculations. However, we and others have shown that the emergence of anti-drug antibodies (ADA) to bNAbs limits their overall expression. Here, we show that we have made significant strides overcome this host immune response. First is through utilizing immune checkpoints that regulate immune system pathways. We observed a 21-fold increase in concentrations of the HIV-1 bNAb 10-1074 in rhesus macaques when macaques were co-inoculated with an AAV vector encoding rhesus macaque PD-L1. Second, we have developed a novel, HIV-1 entry inhibitor, eCD4-Ig, which tends to be more tolerated in rhesus macaques when expressed from AAV vectors. Unlike bNAbs, we have shown that AAV vectors encoding eCD4-Ig can express the inhibitor in macaques for over a year and the ADA response against eCD4-Ig decreases over time. Because eCD4-Ig neutralizes all HIV-1, HIV-2, and SIV isolates and is difficult to escape, it may be useful when included in a therapy strategy. Our pilot studies show that low concentrations AAV-expressed eCD4-Ig can suppress SHIV infection in rhesus macaques for two years, yet viremia is still detectable. This proposal combines our AAV advancements into a single strategy to determine whether AAV-expressed inhibitors can suppress a SHIV infection and reduce the viral reservoir. In Aim 1, we will assess the therapeutic efficacy of the combination of AAV-delivered eCD4-Ig and 10-1074 after ART is lifted in SHIV-infected rhesus macaques. In Aim 2, we will determine whether suppressing an established SHIV infection with AAV- delivered eCD4-Ig and 10-1074 results in a quantitatively different viral reservoir compared to ART. In Aim 3, we will improve the safety of AAV gene therapy by developing an irreversible “kill-switch” to turn off transgene expression.