Dual antiretroviral loaded nanoparticles for long-acting suppressive HIV therapy

项目摘要/摘要 尽管联合抗逆转录病毒疗法(CART)取得了巨大进展，但艾滋病毒仍然是一个全球威胁。 慢性炎症和免疫激活增加了CART中非艾滋病并存的风险 治疗艾滋病患者。这种情况是由多种因素造成的，包括抗逆转录病毒(ARV)毒性和 次级淋巴组织(SLT)中的病毒持久性与ARV在这些组织中的低渗透率有关 水库遗址。此外，每日服用抗逆转录病毒药物(ARV)仍然是一种负担，导致患者 可导致SLT病毒反弹的坚持。在这个项目中，我们建议开发一种长效的 由脂质包裹聚乳酸纳米粒(GM3-PLA-NPs)组成的“隐形”纳米制剂 使用非核苷类逆转录酶抑制剂利培韦林(RPV)和整合酶抑制剂卡替格列韦 (驾驶室)。这些GM3-PLA-NPs具有创新的靶向和设计特征，将增加药物进入和 在SLT中的滞留，除了调节炎症反应。事实上，这种膜结合了 单唾液酸二己糖神经节苷脂(GM3)可与唾液酸凝集素受体CD169特异性结合 髓系细胞(巨噬细胞和树突状细胞)，导致非酸性膜的形成 内陷，可防止NP降解，并导致长期建立细胞药库。 这些NP收集室(NPC)与先前定义的病毒相同，包含 CD169捕获HIV-1时形成的间隔(VCC)。由于VCC也参与了髓系细胞- 通过病毒学突触介导HIV传播到CD4+T细胞(转染性)，我假设 NPCCs可能促进ARV通过巨噬细胞向旁观者T细胞释放 突触连接。我们的初步数据表明，这些GM3-PLA-NPs是成功的 在CD169+巨噬细胞的NPCCs中保存一段时间，使这些细胞的抗病毒效力延长一年 体外培养一个月。随后，观察到在旁观者Jurkat T细胞中持续的病毒抑制。 与NP暴露的CD169+巨噬细胞共同培养。此外，我们的发现表明，GM3脂膜 赋予“隐形”属性，以抑制由加载的ARV和 纳米粒子芯。因此，在目标1中，我们将研究CD169+的细胞外药物释放机制 巨噬细胞转化为CD4+T细胞。在目标2中，我们将评估GM3的长期毒性和隐身机制- 中国人民解放军-NPs。最后，我们将在体内通过评估空间分布和隐身来证实这些结果 GM3-PLA-NPs在SLT中的表型。这些研究将阐明一种安全和 有效地向SLT提供长效治疗药物，可能有利于艾滋病毒的预防和 治疗以及其他疾病，如转移性癌症。

PROJECT SUMMARY/ABSTRACT HIV remains a global threat despite the tremendous progress of combinatorial antiretroviral therapy (cART). Chronic inflammation and immune activation support an increased risk of non-AIDS co-morbidities in cART- treated HIV patients. This condition is attributed to various factors including antiretroviral (ARV) toxicity and viral persistence in secondary lymphoid tissues (SLTs) which has been linked to low ARV penetration in these reservoir sites. Furthermore, uptake of daily antiretrovirals (ARVs) remains a burden resulting in poor adherence that can lead to viral rebound from the SLTS. In this project, we propose to develop a long-acting “stealth” nanoformulation composed of lipid wrapped poly-lactic acid nanoparticles (GM3-PLA-NPs) loaded with a non-nucleoside reverse transcriptase inhibitor, Rilpivirine (RPV) and integrase inhibitor, Cabotegravir (CAB). These GM3-PLA-NPs have innovative targeting and design features that will increase drug entry and retention in SLTs, in addition to modulating inflammatory responses. Indeed, the membrane incorporated monosialo di-hexosylganglioside (GM3) enables specific binding to the sialic acid lectin receptor CD169 on myeloid cells (macrophages and dendritic cells), leading to the formation of non-acidic membrane invaginations which can prevent NP degradation, and lead to long-term establishment of cellular drug depots. These NP collecting compartments (NPCCs) are identical to the previously defined virus containing compartments (VCCs) formed upon capture of HIV-1 by CD169. Since VCCs are also involved in myeloid cell- mediated HIV transmission to CD4+ T cells (trans-infection) through the virological synapse, I hypothesize that NPCCs could potentially promote sustained ARV release to bystander T cells through macrophage – T cell synaptic junctions. Our preliminary data have demonstrated that these GM3-PLA-NPs are successfully preserved in NPCCs of CD169+ macrophages over time which extended anti-viral potency in these cells for a month in vitro. Subsequently, sustained viral inhibition in bystander Jurkat T cells was observed upon co- culture with NP-exposed CD169+ macrophages. Additionally, our findings suggest that the GM3 lipid coating confers “stealth” properties to suppress non-specific immune activation induced by loaded ARVs and nanoparticle core. Thus, in aim 1, we will investigate the extracellular drug release mechanism from CD169+ macrophages to CD4+ T cells. In aim 2, we will assess the long-term toxicity and stealth mechanism of GM3- PLA-NPs. Finally, we will confirm these results in vivo by assessing the spatial distribution and stealth phenotype of GM3-PLA-NPs in SLTs in aim 3. These studies will elucidate a new approach for safe and effective delivery of long-acting therapeutics to SLTs which could be beneficial for HIV prevention and treatment as well as other diseases such as metastatic cancer.