Novel nanoparticles for siRNA delivery

用于 siRNA 递送的新型纳米粒子

基本信息

批准号：
8433254
负责人：
Leaf Huang
金额：
$ 27.64万
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/8433254
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.

描述（由申请人提供）：近年来，我们成功地改进了脂质体/鱼精蛋白/DNA（LPD）纳米颗粒制剂，用于在异种移植模型中将siRNA靶向递送至人肺癌细胞。这种核/壳自组装纳米粒子包含一个紧凑的鱼精蛋白/DNA/siRNA 核心，该核心被两个阳离子脂质双层包裹。当纳米颗粒与聚乙二醇 (PEG)-磷脂缀合物 (DSPE-PEG) 一起孵育时，外双层被剥离，但内双层以高度聚乙二醇化的形式保留下来。紧密堆积的表面 PEG 形成刷保护层，以屏蔽纳米颗粒的阳离子电荷并减少血清蛋白的调理作用。结果是肝脏和脾脏的吸收程度非常低，而肿瘤的吸收程度非常高，高达每克组织注射剂量的 60-80%。如果将靶向配体茴香酰胺连接到 PEG 的远端，则靶向纳米颗粒可以有效地递送 siRNA，以沉默整个肿瘤中的靶基因。然而，在仔细观察递送的 siRNA 后，发现大部分剂量被隔离在内体中并且没有生物利用度。这是因为该制剂不具有任何内体溶解活性。因此，我们建议用在酸性内体 pH 值下溶解的磷酸钙 (CaP) 取代 LPD 的核心。我们预计溶解的CaP将显着增加内体的渗透压并诱导细胞器的肿胀和破裂，从而导致封装的siRNA的释放。所提出的非病毒载体的改进将结合高水平的肿瘤摄取和siRNA货物的有效内体释放。该项目的另一个目标是治疗具有不那么渗漏的新血管系统的肿瘤。我们通过在高温高压下将 siRNA 和阳离子脂质 DOTAP 在氟代烃溶剂中混合，成功制备了小型生物活性脂质复合物 (SBL)。所得纳米粒子很小（30-50 nm），并且在转染中具有活性。我们将通过聚乙二醇化和配体束缚进一步修饰 SBL 的表面。我们期望新的纳米颗粒能够将 siRNA 递送至新血管系统不那么渗漏的肿瘤细胞中。