Novel nanoparticles for siRNA delivery

用于 siRNA 递送的新型纳米颗粒

基本信息

批准号：
8212466
负责人：
Leaf Huang
金额：
$ 23.86万
依托单位：
UNIV OF NORTH CAROLINA CHAPEL HILL
依托单位国家：
美国
项目类别：
财政年份：
2010
资助国家：
美国
起止时间：
2010-04-01 至 2015-01-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8212466
关键词：
Acids Address Animal Model Binding Bioavailable Blood Blood Circulation Caliber Case Study Cell Nucleus Cells Charge Complex Confocal Microscopy Cultured Cells Cultured Tumor Cells Cytoplasm DNA Data Dimerization Distal Dose Drug Formulations Drug Kinetics Drug or chemical Tissue Distribution Employee Strikes Encapsulated Endosomes Fluorescence Fluorinated Hydrocarbons Gene Targeting Gene Transfer High temperature of physical object Human Hyaluronic Acid Image Incubated Inflammatory Injection of therapeutic agent Label Ligands Lipid Bilayers Lipids Liposomes Liver Lung Neoplasms Lytic Malignant Neoplasms Malignant neoplasm of lung Metastatic Neoplasm to the Lung Methods Modeling Modification Molecular Motivation Mus Names Neoplasm Metastasis Non-Viral Vector Nucleic Acids Organelles Osmotic Pressure Penetration Phospholipids Physical condensation Polyethylene Glycols Polylysine Polysaccharides Protamines Research Research Personnel Rupture Serum Serum Proteins Slice Small Interfering RNA Solutions Solvents Spleen Staining method Stains Structure Surface Swelling Temperature Testing Therapeutic Time Tissues Toxic effect Transfection Xenograft Model aqueous base calcium phosphate cancer cell design disulfide bond endosome membrane gene therapy clinical trial gene transfer vector improved inorganic phosphate interest intravenous injection meetings monomer nanoparticle neoplastic cell neovasculature non-viral gene therapy novel particle physical property plasmid DNA polycation pressure public health relevance success targeted delivery trafficking tumor tumor xenograft uptake vector zeta potential

项目摘要

DESCRIPTION (provided by applicant): In the recent years, we have successfully modified the Liposome/Protamine/DNA (LPD) nanoparticle formulation for targeted delivery of siRNA to the human lung cancer cells in a xenograft model. This core/shell, self-assembled nanoparticles contained a compact protamine/DNA/siRNA core which is wrapped around by two cationic lipid bilayers. When the nanoparticles were incubated with a polyethylene glycol (PEG)-phospholipid conjugate (DSPE-PEG), the outer bilayer was stripped off, but the inner bilayer survived with a high degree of PEGylation. The densely packed surface PEG formed a brush protection layer to shield the cationic charges of the nanoparticles and reduced opsonization by serum proteins. The result was a very low degree of uptake by the liver and the spleen, and a very high level of tumor uptake, up to 60-80% injected dose per g tissue. If a targeting ligand, anisamide, was tethered to the distal end of PEG, the targeted nanoparticles efficiently delivered siRNA to silence a target gene in the entire tumor. However, upon a closer look at the delivered siRNA, most of the dose was sequestered in the endosomes and was not bioavailable. This is because the formulation did not possess any endosomal lytic activity. Thus, we propose to replace the core of LPD with Ca phosphate (CaP) which dissolves at the acidic endosome pH. We expect that dissolved CaP will significantly increase the osmotic pressure of the endosome and induce swelling and rupture of the organelle, resulting in the release of the encapsulated siRNA. The proposed improvement of the non-viral vector will combine high level of tumor uptake and efficient endosome release of the siRNA cargo. The other aim of the project will deal with tumors with not-so-leaky neovasculature. We have successfully prepared small bioactive lipoplex (SBL) by mixing siRNA and a cationic lipid, DOTAP, in a fluorohydrocarbon solvent at an elevated temperature and pressure. The resulting nanoparticles were small (30-50 nm), and active in transfection. We will further modify the surface of SBL with PEGylation and ligand tethering. We expect that the new nanoparticle will deliver siRNA to tumor cells in which the neovasculature is not so leaky. PUBLIC HEALTH RELEVANCE: The project will address two pressing issues in delivering siRNA to the tumor. The first is to improve the release of siRNA from the endosome to the cytoplasm. The other is to transport siRNA to tumors with not-so-leaky vasculature. If successful, the project will significantly advance siRNA as cancer therapeutics.

描述（由申请人提供）：近年来，我们成功地修改了脂质体/精胺/DNA（LPD）纳米颗粒制剂，用于在异种移植模型中靶向siRNA向人类肺癌细胞的靶向递送。这种芯/壳体，自组装的纳米颗粒包含一个紧凑的精蛋白/DNA/siRNA核，该核心由两个阳离子脂质双层包裹。当纳米颗粒与聚乙烯乙二醇（PEG） - 磷脂结合物（DSPE-PEG）孵育时，将外部双层剥离，但内部双层具有高度的pegylation。密集填充的表面钉形成了刷子保护层，以遮盖纳米颗粒的阳离子电荷，并通过血清蛋白减少调子化。结果是肝脏和脾脏的摄取程度很低，肿瘤吸收水平很高，每克组织注射剂量高达60-80％。如果将靶向配体（Anisamide）束缚在PEG的远端，则靶向纳米颗粒有效地传递了siRNA，以使整个肿瘤中的靶基因静音。但是，在仔细观察传递的siRNA时，大部分剂量被隔离在内体中，并且不可生物利用。这是因为该公式没有任何内体裂解活性。因此，我们建议用溶解在酸性内体pH的Ca磷酸盐（CAP）代替LPD的核心。我们预计溶解的盖将显着增加内体的渗透压并诱导细胞器的肿胀和破裂，从而释放封装的siRNA。提出的非病毒载体的改善将结合siRNA货物的高水平肿瘤吸收和有效的内体释放。该项目的另一个目的将处理不太晕实的新生血管造成的肿瘤。我们通过将siRNA和阳离子脂质（dotap）混合在氟氢碳酸碳溶剂中以升高的温度和压力中混合，成功制备了小型生物活性脂质（SBL）。所得的纳米颗粒很小（30-50 nm），并在转染中活跃。我们将通过卵形和配体绑扎进一步修改SBL的表面。我们预计新的纳米颗粒会将siRNA输送到肿瘤细胞中，其中新生血管造影不太漏水。公共卫生相关性：该项目将解决将siRNA运送到肿瘤方面的两个紧迫问题。首先是改善siRNA从内体到细胞质的释放。另一种是将siRNA运送到肿瘤的脉络膜不太晕实的肿瘤。如果成功，该项目将大大推动siRNA作为癌症治疗剂。