Ionic liquid-assisted drug delivery to brain reservoirs for treatment of neuroHIV

项目摘要/摘要 尽管抗逆转录病毒疗法(ARTS)取得了成功，但它们不能从 身体，特别是中枢神经系统(CNS)的那些。此外，阿片类药物上调外排转运体 进一步促进抗逆转录病毒药物在中枢神经系统的亚治疗浓度。我们使用了生物相容的离子液体 (ILS)，一种由不对称阳离子和阴离子组成的熔融盐，可以将纳米颗粒与 不同的细胞类型。使用这一策略，我们已经开发出对红细胞具有平衡亲和力的IL和 小胶质细胞促进纳米颗粒在红细胞上搭便车，将它们运送到大脑，细胞- 一旦小胶质细胞被送到中央隔室，就有选择性的靶向。在老鼠身上的初步数据表明 大约48%的注射纳米颗粒在6小时内在大脑中积累，比目前的情况有了很大的改善 纳米颗粒输送策略。初步分析表明，90%以上的CNS纳米颗粒是 与小胶质细胞有关。我们进一步证明了将ART(阿巴卡韦)装载到纳米粒中的能力 对感染HIV的人外周血单核细胞仍有抗病毒作用 (PBMCs)。我们假设我们可以进一步改进IL配方的可调轮廓，以优化货物 递送到大脑和靶向其他类型的细胞(包括星形胶质细胞)。我们预计这批货物 尽管阿片类药物促进了中枢神经系统细胞黏附/转运体的改变，但该策略将是安全有效的 暴露/依赖。为此，我们将(目标1)产生至少5个新的ILS，除了我们目前的领先地位， 具有不同的细胞类型亲和力。我们将确认ILS对猿猴和人类血液的偏好 作为潜在货物承运人的零部件。(目标2)我们将评估安全性(亚急性、急性、亚慢性、 最多5种新型ILS在阿片类药物依赖或阿片类药物依赖的大鼠体内的生物分布。 ILS将装载预制的加扰Cas9载体和EGFP报告程序，以确认交付的能力 CRISPR-CAS9构建了潜在的HIV治愈策略。具有CNS有利生物分布的安全ILS将是 用CART(阿巴卡韦、多洛替格韦和拉米夫定)装载并评估对SIV感染或HIV感染的疗效。 分别感染猴或人的PBMCs，或人的小胶质细胞。所有的细胞都将是阿片-幼稚或阿片- 暴露了。对小胶质细胞和星形胶质细胞具有选择性的ILS将被优先考虑。(目标3)白线(基于中枢神经系统 分布和小胶质细胞/星形胶质细胞的选择性)将评估体内的安全性、生物分布和急性 SIV恒河猴模型的疗效。在猕猴体内，IL辅助的纳米颗粒将被装载到纳米金中， 除CART外，还可通过X射线确认生物分布的时间进程。将通过以下途径监测SIV感染 脑脊液和抽血。完整的血细胞计数、化学和细胞因子分析将在血浆中进行 和/或脑脊液。ART的分布将通过LC/MS确认。Gross将对器官进行组织病理学检查 此外，还将对中枢神经系统组织进行小胶质细胞增多症、星形胶质细胞增多症和亚致死性神经元损伤的评估。伊尔- 辅助纳米粒可实现安全、细胞特异性、中枢神经系统药物/货物递送的目标。

PROJECT SUMMARY/ABSTRACT Despite the success of antiretroviral therapeutics (ARTs), they cannot eradicate HIV from reservoirs within the body, particularly those in the central nervous system (CNS). Moreover, opioids upregulate efflux transporters further promoting subtherapeutic concentrations of ART in the CNS. We have used biocompatible ionic liquids (ILs), molten salts comprised of asymmetric cations and anions, that can `tune' the affinity of nanoparticles to different cell types. Using this strategy, we have developed an IL with a balanced affinity for erythrocytes and microglia which promotes nanoparticle `hitchhiking' on erythrocytes to deliver them to the brain, and cells- selective targeting of microglia once delivered to the central compartment. Preliminary data in rats demonstrate ~48% of injected nanoparticles accumulating in the brain within 6 hours, a vast improvement over current nanoparticle delivery strategies. Preliminary analyses indicated that over 90% of CNS nanoparticles were associated with microglia. We have further demonstrated the capacity to load ART (abacavir) into nanoparticles which retained antiviremic efficacy when administered to HIV-infected human peripheral blood mononuclear cells (PBMCs). We hypothesize that we can further improve the tunable profile of our IL formulation to optimize cargo delivery to the brain and target additional cell types (including astrocytes). We anticipate that this cargo delivery strategy will be safe and efficacious in spite of CNS cell adhesion/transporter changes promoted by opioid exposure/dependence. To this end, we will (Aim 1) generate at least 5 novel ILs, in addition to our current lead, with varied cell-type affinity. We will confirm the preference that ILs confer to simian and human blood components as potential cargo carriers. (Aim 2) We will assess the safety (subacute, acute, subchronic, reproductive, mutagenic) and biodistribution of up to 5 novel ILs in rats that are opioid-naïve or opioid-dependent. ILs will be loaded with a premade scramble Cas9 vector with an eGFP reporter to confirm the capacity to deliver CRISPR-Cas9 constructs for potential HIV cure strategies. Safe ILs with a CNS-favorable biodistribution will be loaded with cART (abacavir, dolutegravir, and lamivudine) and assessed for efficacy in SIV-infected or HIV- infected simian or human PBMCs, respectively, or human microglia. All cells will be opioid-naïve or opioid- exposed. ILs with selectivity for microglia and astrocytes will be prioritized. (Aim 3) IL leads (based on CNS distribution and microglial/astrocytic selectivity) will be assessed for in vivo safety, biodistribution, and acute efficacy in a rhesus macaque model of SIV. In macaques, IL-assisted nanoparticles will be loaded with nanogold, in addition to cART, to confirm the time-course of biodistribution via X-ray. SIV infection will be monitored via CSF and blood draws. Complete blood count, chemistry, and cytokine profiling will be conducted on plasma and/or CSF. ART distribution will be confirmed via LC/MS. Gross histopathology will be conducted on organs and CNS tissues will be additionally assessed for microgliosis, astrogliosis, and sublethal neuronal damage. IL- assisted nanoparticles may realize the goal of achieving safe, cell-specific, CNS drug/cargo delivery.