Understanding the Relationship LNP Structure, Cholesterol Trafficking, and InVivo Delivery

了解 LNP 结构、胆固醇运输和体内递送之间的关系

• 批准号: 10753191
• 负责人: James Dahlman
• 金额: $ 37.79万
• 依托单位: EMORY UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-09-01 至 2024-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10753191
• 关键词: Understanding; Relationship; LNP; Structure; Cholesterol

Project Summary Scientists can create thousands of chemically distinct nanoparticles using a growing number of high throughput chemistries, but it is still difficult to test more than a few nanoparticles in vivo. The goal of this work is to substantially improve how lipid nanoparticles (LNPs) deliver nucleic acid therapies by performing a systematic high throughput in vivo LNP study. This goal will be achieved using cutting edge DNA barcoded nanoparticles; deliverer mediated by 300 different nanoparticles can be measured in a single mouse. 4,320 chemically distinct nanoparticles will be tested in vitro and in vivo, focusing on 2 fundamental questions. First, how does nanoparticle structure affect cell targeting in vivo? Nanoparticle chemical and physical traits affect delivery in vitro. However, the extent to which the same LNP traits influence delivery in animals (in vivo) is unclear. A recently developed bioinformatics pipeline will be used to (i) systematically analyze how LNP structure affects in in vivo delivery in macrophages, endothelial cells, and hepatocytes, both in vitro and in vivo. The same data will be used to (ii) quantify the precision with which in vitro drug delivery predicts in vivo drug delivery. Second, how do clinically relevant physiological changes affect delivery in vivo? LNPs are similar to lipoproteins, which are natural lipid-containing nanostructures. Lipoproteins are actively trafficked to endothelial cells, macrophages, and hepatocytes in vivo. Given that lipoprotein trafficking changes in patients with high cholesterol, taking statins, and patients with many other conditions, LNP transport may also change. The top 600 in vivo LNPs from the 4,320 LNP in vivo screen will be administered to genetic mouse models of aberrant lipid transport in order to (iii) investigate how genetic alterations in cholesterol trafficking affect in vivo delivery. This work will make 5 significant contributions to nanotechnology. First, the extent to which LNP chemical traits influence delivery directly in vivo will be tested; relationships between nanoparticle structure and delivery are studied in vitro. Second, the precision with which in vitro nanoparticle delivery predicts in vivo delivery will be quantified. This could increase the efficiency with which clinical nanoparticles are discovered. Third, the effect of clinically relevant physiological changes on LNP delivery will be examined. Nanoparticles can interact with cholesterol trafficking pathways; these interactions are likely to change with disease and can affect nanoparticle targeting / safety. Fourth, the feasibility of studying thousands of LNPs in vivo will be demonstrated. Fifth, open source protocols for nanoparticle barcoding will be established and disseminated. These results will provide crucial insight into the ways LNP chemical traits and specific genes alter LNP delivery, informing the design of LNPs that deliver nucleic acid cargos (e.g., siRNA, mRNA, CRISPR-Cas9) for numerous therapeutic applications.

项目摘要 科学家可以使用越来越多的高通量来制造数千种化学上截然不同的纳米颗粒 化学，但仍然很难在体内测试几个以上的纳米颗粒。这项工作的目标是 通过进行系统的研究，显著改善脂质纳米粒(LNPs)提供核酸治疗的方式 高通量体内LNP研究。这一目标将使用尖端的DNA条形码纳米颗粒来实现； 由300种不同纳米颗粒介导的递送器可以在一只小鼠身上测量。4320种不同的化学成分 纳米粒子将在体外和体内进行测试，重点是两个基本问题。首先，如何 纳米颗粒结构会影响体内的细胞靶向吗？纳米颗粒的化学和物理特性影响传递 在试管中。然而，相同的LNP特性在多大程度上影响动物体内的传递尚不清楚。一个 最近开发的生物信息学流水线将用于(I)系统地分析LNP结构如何影响 体内释放的巨噬细胞，内皮细胞和肝细胞，在体外和体内。相同的数据将 用于(Ii)量化体外给药预测体内给药的精确度。第二，如何 临床相关的生理变化会影响活体分娩吗？LNPs类似于脂蛋白，它 是天然的含脂纳米结构。脂蛋白被活跃地运输到内皮细胞、巨噬细胞、 和体内的肝细胞。鉴于高胆固醇患者脂蛋白转运的变化，服用他汀类药物， 与其他许多情况下的患者相比，LNP的转运也可能发生变化。体内前600个LNPs来自 将4,320个LNP体内筛选应用于遗传性脂质转运异常的小鼠模型，以期(III) 研究胆固醇转运中的基因改变如何影响体内传递。这项工作将使5 对纳米技术的重大贡献。首先，LNP的化学特性对交付的影响程度 直接在体内进行测试；在体外研究纳米颗粒结构和传递之间的关系。 第二，纳米颗粒体外释放预测体内释放的精确度将被量化。这 可以提高临床纳米粒子的发现效率。第三，临床疗效。 将检查有关LNP递送的生理变化。纳米颗粒可以与胆固醇相互作用 运输途径；这些相互作用可能会随着疾病而改变，并可能影响纳米颗粒的靶向/ 安全。第四，将论证在体内研究数千个LNPs的可行性。第五，开源 将建立和传播纳米颗粒条形码的协议。这些结果将提供至关重要的 洞察LNP化学特性和特定基因改变LNP传递的方式，为LNP的设计提供信息 为多种治疗应用提供核酸货物(例如，siRNA、mRNA、CRISPR-Cas9)。