GENE THERAPY FOR DYSLIPIDEMIA AND ATHEROSCLEROSIS

血脂异常和动脉粥样硬化的基因治疗

• 批准号: 7110300
• 负责人: Daniel James Rader
• 金额: $ 41.47万
• 依托单位: UNIVERSITY OF PENNSYLVANIA
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/7110300
• 关键词: Adenoviridae; RNA interference; adeno associated virus group; alkyltransferase; antiatherogenic agent; apolipoprotein B; apolipoprotein E; atherosclerosis; atherosclerotic plaque; biotechnology; blood lipoprotein biosynthesis; blood lipoprotein metabolism; diacylglycerols; familial hyperlipoproteinemia type II; gene therapy; high density lipoproteins; laboratory mouse; laboratory rabbit; lipoprotein disorder; low density lipoprotein; low density lipoprotein receptor; nonhuman therapy evaluation; pathologic process; transfection /expression vector; very low density lipoprotein

Dyslipidemia is a major risk factor for premature atherosclerotic cardiovascular disease (ASCVD). Although many patients with dyslipidemia can be treated effectively with existing drugs, others are not effectively treated and remain at exceptionally high risk of premature ASCVD; the classic example is homozygous familial hypercholesterolemia (FH). Therefore, understanding of the regulation of the secretion and catabolism of apoB-containing lipoproteins by the liver and the complex pathways of HDL metabolism is of major importance to the development of new therapies targeted toward these pathways. Liver-directed somatic gene transfer is a useful biological tool for addressing hypotheses regarding the physiological effects of expressing specific genes in the liver on apoB lipoprotein and HDL metabolism; furthermore, it could be a strategy for treating severe dyslipidemia. In this project, we will utilize liver-directed gene transfer using vectors based on novel adeno-associated virus (AAV) pseudotypes (as described in Project 1) to address questions related to the impact of specific gene products and their interactions on the regulation of hepatic secretion of apoB-containing lipoproteins. These projects have been designed to leverage the substantial advantages of vectors based on novel AAV pseudotypes, such as AAV2/8, including the stable and efficient transduction of hepatocytes after intravenous administration without apparent inflammation. AAV2/8-based vectors are a major advance over AAV2-based vectors with regard to levels of expression (more than an order of magnitude in our experience) and over adenoviral based vectors because of the lack of inflammation (compared to severe inflammation with adenoviral vectors) and the stable long-term expression (compared to transient expression with adenoviral vectors). Specific Aim 1. To determine the roles of hepatic DGAT1 and DGAT2 expression in affecting hepatic VLDL TG and apoB production through AAV-mediated gene transfer of cDNAs to achieve overexpression through AAV-mediated gene transfer of RNAi to achieve reduction of expression. To test the hypothesis that hepatic MTP expression interacts with hepatic DGAT expression in influencing hepatic VLDL production. Specific Aim 2. To test the hypothesis that AAV-mediated overexpression of the LDL receptor (LDLR) or VLDL receptor (VLDLR) increases presecretory apoB degradation, reducing the hepatic VLDL triglyceride and apoB production rate. To determine whether AAV-mediated LDLR and VLDLR overexpression can normalize VLDL production in mouse models of VLDL triglyceride and apoB overproduction. Specific Aim 3. To compare AAV-mediated liver-directed gene transfer of the LDLR and the VLDLR in LDLR/apobec-1 double knockout mice on progression and regression of pre-existing atherosclerosis. To compare AAV-mediated gene transfer of the LDL receptor and the VLDL receptor in WHHL rabbits on lipoprotein metabolism and atherosclerosis. Specific Aim 4: To use AAV-mediated liver-directed gene transfer of apoA-I to test whether long-term expression of apoA-I can prevent progression and induce regression of atherosclerosis and in murine and rabbit models of atherosclerosis. To test whether long-term stable expression of apoA-I is additive to long-term cholesterol reduction with regard to atherosclerosis progression and regression. The overall goals of this grant proposal are to use AAV-mediated liver-directed gene transfer: 1) to test specific hypotheses related to the role of DGAT1 and DGAT2 in the regulation of hepatic VLDL TG and apoB production in vivo through gene transfer-mediated overexpression and gene knockdown approaches; 2) to compare gene transfer of the LDLR and the VLDLR with regard to effects on VLDL production, on plasma lipoproteins, and on atherosclerosis in two different animal models of FH; and 3) to generate proof-of-principle that AAV-mediated gene transfer of apoA-I to liver can induce regression and prevent progression of atherosclerosis in mice and rabbits. This project will generate new biological information and will also generate proof of principle in relevant animal models of homozygous familial hypercholesterolemia and low HDL cholesterol that would support the concept of gene transfer as a potential therapeutic approach to these disorders.

血脂异常是过早动脉粥样硬化心血管疾病（ASCVD）的主要危险因素。尽管许多血脂异常患者可以通过现有药物有效治疗，但其他患者没有有效治疗，并且仍然处于过早ASCVD的高风险。经典的例子是纯合家族性高胆固醇血症（FH）。所以， 了解肝脏对含APOB的脂蛋白的分泌和分解代谢的理解以及HDL代谢的复杂途径对于开发针对这些途径的新疗法至关重要。肝脏定向的体细胞基因转移是一种有用的生物学工具，用于解决有关生理效应的假设 在APOB脂蛋白和HDL代谢上表达肝脏中的特定基因；此外，这可能是治疗严重血脂异常的策略。在这个项目中，我们将使用基于新型腺相关病毒（AAV）假型（如项目1中所述）的媒介使用载体转移基因转移，以解决与特定基因产物及其相互作用对APOB抗APOB抗APOB抗抗蛋白质的肝分泌的影响有关的问题。这些项目旨在利用基于新型AAV伪型（例如AAV2/8）的矢量的实质优势，包括静脉内给药后稳定有效的肝细胞的稳定，有效的转导而没有明显的炎症。基于AAV2/8的矢量是基于AAV2的矢量的主要进步，因为缺乏炎症（与腺病毒载体严重的炎症相比，与稳定的长期表达相比，由于缺乏炎症（与腺病毒的强烈炎症相比 向量）。具体目的1。确定通过AAV介导的基因转移CDNA的基因转移肝DGAT1和DGAT2表达在影响肝VLDL TG和APOB产生中的作用，从而通过RNAI的AAV介导的基因转移实现过表达，以实现表达的减少。为了测试肝MTP表达在影响肝VLDL产生时与肝DGAT表达相互作用的假设。具体目的2。为了测试AAV介导的LDL受体（LDLR）或VLDL受体（VLDLR）的过表达的假设增加了预先分泌的APOB降解，从而降低了肝VLDL甘油三酸酯和APOB的产生速率。为了确定AAV介导的LDLR和VLDLR过表达是否可以使VLDL甘油三酸酯和APOB过量生产的小鼠模型中的VLDL产生标准化。具体目的3。要比较LDLR/APOBEC-1双基因敲除小鼠中LDLR和VLDLR的AAV介导的肝脏指导的基因转移与预先存在的动脉粥样硬化的进展和退化。比较WHHL兔中LDL受体和VLDL受体在脂蛋白代谢和动脉粥样硬化中的AAV介导的基因转移。特定目的4：使用AAV介导的肝脏指导的基因转移APOA-I来测试ApoA-I的长期表达是否可以防止动脉粥样硬化的进展和诱导动脉粥样硬化模型的回归。为了测试apoA-I的长期稳定表达是否在动脉粥样硬化的进展和消退方面是长期胆固醇还原的添加剂。该赠款提案的总体目标是使用AAV介导的肝脏指导的基因转移：1）测试与DGAT1和DGAT2在调节肝VLDL TG和APOB生产中通过基因转移介导的过表达和基因敲低方法在体内调节肝VLDL TG和APOB生产相关的特定假设； 2）比较对VLDL产生，血浆脂蛋白以及对vLDL的影响的基因转移和VLDLR的基因转移 FH的两个不同动物模型中的动脉粥样硬化； 3）为了产生原理证明，AAV介导的apoA-I基因转移至肝脏可以诱导小鼠和兔子动脉粥样硬化的回归并防止动脉粥样硬化的进展。该项目将产生新的生物学信息，并将在纯合家族性高胆固醇血症和低HDL胆固醇的相关动物模型中产生原理证明，这将支持基因转移的概念，作为这些疾病的潜在治疗方法。