Targeted cell-depleting immunotherapy for TB and HIV

项目摘要 结核分枝杆菌和MDSC已经成为抑制结核肉芽肿中效应免疫的关键抑制细胞。 此外，TdR与HIV潜伏期的维持有关。我们选择了两个 使用白喉毒素（DT）融合蛋白有效消耗THP和MDSC的生物制剂 技术：DT-IL 2靶向IL 2-R阳性（CD 25+）细胞（THBE），DT-IL 4靶向IL 4-R阳性（CD 124+） 细胞（MDSC）。我们的初步数据表明：（i）DT-IL 2和DT-IL 4降低Treg的丰度， DT-IL 2和DT-IL 4分别降低小鼠肺和脾中M. tb到 在急性鼠TB感染模型中增殖，（iii）具有有效抗MDSC活性的药物他喹莫德， 降低了M. tb在急性TB感染模型中增殖并伴随MDSC的减少， 和（iv）将DT-IL 2给予来自控制HIV的患者的CD 4 + T细胞再激活病毒复制。 这些数据支持了以下假设：（1）TGFAP和MDSC在M. TB 遏制，和（2）Treg细胞在HIV潜伏期维持中发挥作用。一个核心的科学前提是 应用是进一步确定TcB和MDSC在TB和HIV感染中的免疫抑制作用 将揭示重要的免疫发病机制，可用于开发改善的 针对这两种疾病的宿主导向疗法（HDTs）。在本提案中，我们寻求通过以下方式解决这一前提： 以下目标。目的1：使用靶向CD 25+细胞去除来确定Treg细胞在TB肉芽肿中的作用 DT-IL2我们将（a）评估细胞（C57 BL/6，WT，其为sst 1 R）和 使用单细胞RNA-Seq检测CD 25+细胞耗竭和未耗竭的坏死性（C57 BL/6 sst 1 S）鼠肉芽肿 （B）评估靶向CD 25+细胞耗竭在慢性炎症中的功效， 用和不用抗TB药物治疗的鼠TB，和（c）评估Treg细胞对该过程的贡献 的空洞（肉芽肿破裂），使用一种新的模型，兔空洞结核病。目标2：确定 使用DT-IL 4靶向CD 124+细胞耗竭的TB肉芽肿中的MDSC。我们将（a）评估细胞 使用SSRS进行CD 124+细胞去除和未去除的细胞和坏死性鼠肉芽肿的组成 与传统方法一样，（B）评估靶向CD 124+细胞耗竭在慢性鼠TB中的功效 （c）评估双重CD 25+和CD 124+细胞耗竭在TB患者中的影响， 使用SSRS以及传统方法，用B6 sst 1 R和sst 1 S小鼠的肉芽肿，和（d）评估 双重CD 25+和CD 124+细胞去除在有和没有抗TB药物治疗的慢性鼠TB中的功效。 目的3：利用（i）BCG疫苗接种和（ii）HIV进行CD 25+细胞耗竭的翻译应用 重新激活我们将（a）评估靶向CD 25+细胞耗竭作为疫苗佐剂的有效性， BCG疫苗接种和（B）评估靶向CD 25+细胞耗竭对PBMC中HIV再活化的影响 艾滋病病毒感染得到控制的患者。

PROJECT SUMMARY Tregs and MDSCs have emerged as key suppressor cells that inhibit effector immunity in the TB granuloma. In addition, Tregs have been implicated in the maintenance of HIV latency. We have generated two selective biologic agents for potent depletion of Tregs and and MDSCs using diphtheria toxin (DT) fusion protein technology: DT-IL2 targets IL2-R-positive (CD25+) cells (Tregs), and DT-IL4 targets IL4-R positive (CD124+) cells (MDSCs). Our preliminary data demonstrate that: (i) DT-IL2 and DT-IL4 reduce the abundance of Treg and MDSC cells, respectively, in mouse lungs and spleens, (ii) DT-IL2 and DT-IL4 reduces the ability of M. tb to proliferate in an acute murine TB infection model, (iii) the drug, tasquinimod which has potent anti-MDSC activity, reduced the ability of M. tb to proliferate in an acute TB infection model with a concomitant reduction in MDSCs, and (iv) administration of DT-IL2 to CD4+ T cells from patients with controlled HIV reactivates viral replication. These data support the hypotheses that (1) Tregs and MDSCs play important inhibitory roles in M. tb containment, and (2) that Treg cells play a role in HIV latency maintenance. A central scientific premise of this application is that further definition of the immunosuppressive roles of Tregs and MDSCs in TB and HIV infection will reveal important immunopathogenesis mechanisms that may be exploited towards developing improved host-directed therapies (HDTs) for both diseases. In this proposal we seek to address this premise by pursuing the following aims. Aim 1: define the role of Treg cells in the TB granuloma using targeted CD25+ cell depletion with DT-IL2. We will (a) evaluate the cellular composition of both cellular (C57BL/6, WT which are sst1R) and necrotic (C57BL/6 sst1S) murine granulomas with and without CD25+ cell depletion using single-cell RNA-Seq (SSRS) as well as traditional methods, (b) evaluate the efficacy of targeted CD25+ cell depletion in chronic murine TB with and without anti-TB drug therapy, and (c) evaluate the contribution of Treg cells to the process of cavitation (granuloma breakdown) using a novel model of rabbit cavitary TB. Aim 2: define the role of MDSCs in the TB granuloma using targeted CD124+ cell depletion with DT-IL4. We will (a) evaluate the cellular composition of both cellular and necrotic murine granulomas with and without CD124+ cell depletion using SSRS as well as traditional methods, (b) evaluate the efficacy of targeted CD124+ cell depletion in chronic murine TB with and without anti-TB drug therapy, (c) assess the impact of dual CD25+ and CD124+ cell depletion in the TB granuloma with B6 sst1R and sst1S mice using SSRS as well as traditional methods, and (d) evaluate the efficacy of dual CD25+ and CD124+ cell depletion in chronic murine TB with and without anti-TB drug therapy. Aim 3: conduct translational applications of CD25+ cell depletion with (i) BCG vaccination and (ii) HIV reactivation. We will (a) evaluate the efficacy of targeted CD25+ cell depletion as a vaccine adjuvant prior to BCG vaccination and (b) evaluate the impact of targeted CD25+ cell depletion on HIV reactivation in PBMCs from patients with controlled HIV infection.