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: 10172933
• 负责人: James Dahlman
• 金额: $ 37.18万
• 依托单位: GEORGIA INSTITUTE OF TECHNOLOGY
• 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/10172933
• 关键词: Acrylates; Affect; Amines; Animals; Apolipoprotein E; Bar Codes; Biocompatible Materials; Bioinformatics; CRISPR/Cas technology; Cell Culture Techniques; Cell Line; Cells; Chemical Structure; Chemicals; Chemistry; Cholesterol; Clinical; Custom; DNA; DNA delivery; Data; Disease; Drug Delivery Systems; Dyslipidemias; Endothelial Cells; Epoxy Compounds; Feasibility Studies; Genes; Genetic; Genetic Models; Goals; Heart; Hepatocyte; High Fat Diet; Human; Immune response; In Vitro; Individual; Knockout Mice; Kupffer Cells; Lead; Length; Lipids; Lipoproteins; Liver; Lung; Measures; Mediating; Messenger RNA; Mus; Mutation; Nanostructures; Nanotechnology; Nucleic Acids; Organism; Pathway interactions; Patients; Physiological; Property; Protocols documentation; Safety; Scientist; Small Interfering RNA; Spleen; Structure; Testing; Therapeutic; Tissues; Toxic effect; Wild Type Mouse; Work; bioinformatics pipeline; cell type; cholesterol trafficking; clinical effect; clinically relevant; deep sequencing; design; experimental study; improved; in vitro testing; in vivo; insight; iterative design; lipid nanoparticle; lipid transport; macrophage; mouse model; nanoparticle; nanoparticle delivery; open source; patient population; tertiary amine; trafficking; trait

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 科学家们可以使用越来越多的高通量技术制造出数千种化学性质不同的纳米颗粒。 化学，但仍然难以在体内测试超过几个纳米颗粒。这项工作的目标是 通过进行系统的核酸治疗， 高通量体内LNP研究。这一目标将通过使用尖端的DNA条形码纳米颗粒来实现; 可以在单个小鼠中测量由300种不同纳米颗粒介导的递送剂。4，320种化学成分不同 纳米颗粒将在体外和体内进行测试，重点是两个基本问题。首先， 纳米颗粒结构影响体内细胞靶向？纳米颗粒的化学和物理特性影响递送 体外然而，在何种程度上相同的LNP性状影响交付在动物（体内）是不清楚的。一 最近开发的生物信息学管道将用于（i）系统地分析LNP结构如何影响 在巨噬细胞、内皮细胞和肝细胞中的体内递送，包括体外和体内。同样的数据将 用于（ii）量化体外药物递送预测体内药物递送的精度。二是如何 临床相关生理变化是否影响体内给药？LNP类似于脂蛋白， 是天然的含脂质的纳米结构。脂蛋白被主动运输到内皮细胞，巨噬细胞， 和肝细胞。考虑到高胆固醇患者脂蛋白运输的变化，服用他汀类药物， 以及患有许多其他疾病的患者，LNP转运也可能发生变化。来自美国的前600个体内LNP 将4，320 LNP体内筛选施用至异常脂质转运的遗传小鼠模型，以（iii） 研究胆固醇运输的遗传改变如何影响体内递送。这项工作将使5 对纳米技术的重大贡献。首先，LNP化学性状影响分娩的程度 直接在体内进行测试;纳米颗粒结构和递送之间的关系在体外进行研究。 其次，将量化体外纳米颗粒递送预测体内递送的精度。这 可以提高发现临床纳米颗粒的效率。三、临床效果 将检查LNP递送的相关生理变化。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变化，并可能影响纳米颗粒靶向/靶向性。 安全为代价的第四，将证明在体内研究数千种LNP的可行性。第五，开源 将制定和传播纳米粒子条形码化的方案。这些结果将提供至关重要的 深入了解LNP化学性状和特定基因改变LNP递送的方式，为LNP的设计提供信息 其递送核酸货物（例如，siRNA、mRNA、CRISPR-Cas9）用于多种治疗应用。