Gene therapy of alcoholic liver disease

酒精性肝病的基因治疗

• 批准号: 9297754
• 负责人: DAVID Q WANG
• 金额: $ 24.01万
• 依托单位: ALBERT EINSTEIN COLLEGE OF MEDICINE, INC
• 依托单位国家: 美国
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-08-20 至 2019-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9297754
• 关键词: Alcohol consumption; Alcoholic Liver Diseases; Alcoholic beverage heavy drinker; Alcoholic liver damage; Anabolism; Animals; Apolipoproteins; Apoptosis; Atherosclerosis; Biological Markers; Blood Circulation; Capsid; Carnitine O-Palmitoyltransferase; Cessation of life; Chronic; Cicatrix; Clinical; Data; Dependovirus; Diabetes Mellitus; Dose; Energy Metabolism; Ethanol; Family; Fatty Acids; Fatty Liver; Fibrosis; Gene Transfer; Genes; Genome; Goals; Hepatic; Hepatocyte; Homeostasis; Human; Immune response; Impairment; Inflammation; Inflammatory Response; Injury; Knock-out; Knockout Mice; LacZ Genes; Liver; Liver Fibrosis; Liver diseases; Lysophosphatidylcholines; Mediating; Methods; Mission; Modeling; Molecular; Mus; National Institute on Alcohol Abuse and Alcoholism; Obesity; Pathogenesis; Plasma; Play; Predisposition; Prevention; Process; Regulation; Research; Research Personnel; Risk Factors; Role; Steatohepatitis; Testing; Therapeutic; Transgenic Organisms; Triglyceride Metabolism; Triglycerides; United States National Institutes of Health; Very low density lipoprotein; Viral Vector; Wild Type Mouse; Work; alcohol effect; base; cytotoxic; fascinate; fatty acid metabolism; feeding; gene therapy; human study; innovation; insight; liver development; member; neglect; novel; overexpression; oxidation; prevent; problem drinker; therapeutic development; therapy development

Abstract Abnormalities in triglyceride (TG) metabolism are a critical risk factor for fatty liver disease, atherosclerosis, obesity and diabetes, all of which are major clinical problems worldwide. ApoAV is a new member of the apolipoprotein family and is synthesized exclusively in the liver. A fact that is often neglected is that the concentration of apoAV in human plasma is extremely low (114-258 ng/mL), 1,000 to 10,000-fold lower than that of apoB100 and apoAI, respectively. It has fascinated investigators for many years how a very low circulating level of apoAV can exert such a profound effect on plasma TG homeostasis in humans and animals. Because ~20% of apoAV is secreted into the circulation, its concentration is significantly higher in the liver than in plasma. Our preliminary data show that apoAV plays a critical role in regulating hepatic TG metabolism, and ethanol disrupts hepatic TG homeostasis and exacerbates the accumulation of excess TG in the liver, leading to more severe alcoholic liver disease (ALD) in apoAV knockout (KO) mice than in wild-type (WT) mice. We further found that ethanol inhibits expression of apoAV in a dose- dependent manner in the mouse primary hepatocytes. Chronic and binge ethanol feeding (i.e., the NIAAA model) for 4 weeks leads to a significant and rapid development of liver steatosis, gradually evolving from simple steatosis to alcoholic steatohepatitis and then to liver fibrosis in apoAV KO, but not WT mice. More importantly, ethanol significantly increases hepatic concentrations of lysophosphatidylcholine (lysoPC), a major fatty acid metabolite, by enhancing its biosynthesis in apoAV KO mice. A recent human study has found that lysoPC is a new and important biomarker for excess alcohol intake; however, its vital role in the pathogenesis of ALD is still unknown. By contrast, transgenic expression of the human APOAV gene in mice protects against ethanol-induced liver injury. Our results demonstrate that the absence of apoAV is an important risk factor for ALD and apoAV is a novel target for the prevention and the treatment of ALD. Thus, our preliminary results support the hypothesis that adeno-associated-virus 2/8 (AAV2/8)-mediated gene transfer of the human APOAV protects against ALD in ethanol-fed mice. Of the commonly used viral vectors, AAV produces the lowest immune response and is non-pathogenic even in the wild-type state, as well as is the most suitable viral vector for therapeutic applications. AAV2/8 (i.e., AAV-8 pseudotypes in which AAV-2 genomes are packaged into AAV-8 capsids) displays highly efficient liver gene transfer. Therefore, AAV2/8 is a promising method for gene therapy of ALD. We will test the hypothesis in two specific aims: First, we will investigate whether overexpression of APOAV protects against ethanol-induced liver steatosis in mice transduced with AAV2/8- APOAV. Second, we will explore whether AAV2/8-APOAV treatment prevents ethanol-triggered progression from simple steatosis to alcoholic steatohepatitis through lysoPC in mice. The proposed studies are innovative because this project may provide an efficacious strategy for the gene therapy of ALD.

抽象的 甘油三酯（TG）代谢异常是脂肪肝、动脉粥样硬化、 肥胖和糖尿病都是世界范围内的主要临床问题。 ApoAV 是新成员 载脂蛋白家族，仅在肝脏中合成。一个经常被忽视的事实是 人血浆中apoAV的浓度极低（114-258 ng/mL），比血浆低1,000至10,000倍 分别是apoB100和apoAI。多年来，调查人员一直着迷于如此低的 apoAV 的循环水平可以对人类和动物的血浆 TG 稳态产生如此深远的影响。 由于约 20% 的 apoAV 被分泌到循环系统中，因此其在肝脏中的浓度明显高于肝脏中的浓度。 在血浆中。我们的初步数据表明apoAV在调节肝脏TG中发挥着关键作用 代谢，乙醇会破坏肝脏 TG 稳态并加剧 肝脏中过量的 TG，导致 apoAV 敲除 (KO) 中更严重的酒精性肝病 (ALD) 小鼠比野生型（WT）小鼠。我们进一步发现乙醇在一定剂量下抑制apoAV的表达 小鼠原代肝细胞中的依赖性方式。长期和暴食乙醇喂养（即 NIAAA 模型）4周导致肝脏脂肪变性显着且快速发展，逐渐从 在 apoAV KO 小鼠中，单纯性脂肪变性发展为酒精性脂肪性肝炎，然后发展为肝纤维化，但 WT 小鼠则不然。更多的 重要的是，乙醇显着增加肝脏中溶血磷脂酰胆碱（lysoPC）的浓度，溶血磷脂酰胆碱是一种主要的 脂肪酸代谢物，通过增强 apoAV KO 小鼠的生物合成。最近的一项人类研究发现 lysoPC 是一种新的重要的酒精摄入过量生物标志物；然而，它在发病机制中的重要作用 ALD 的发病率仍未知。相比之下，人类 APOAV 基因在小鼠体内的转基因表达可以防止 乙醇引起的肝损伤。我们的结果表明，apoAV 的缺失是一个重要的风险因素。 ALD和apoAV是预防和治疗ALD的新靶点。因此，我们的初步结果 支持腺相关病毒 2/8 (AAV2/8) 介导的人类基因转移的假设 APOAV 可防止乙醇喂养的小鼠发生 ALD。在常用的病毒载体中，AAV 产生 免疫反应最低，即使在野生型状态下也是非致病性的，也是最合适的病毒 用于治疗应用的载体。 AAV2/8（即包装了 AAV-2 基因组的 AAV-8 假型） 转化为 AAV-8 衣壳）显示出高效的肝脏基因转移。因此，AAV2/8是一种有前途的方法 ALD 的基因治疗。我们将在两个具体目标上检验该假设：首先，我们将调查是否 APOAV 过表达可防止 AAV2/8- 转导小鼠乙醇诱导的肝脏脂肪变性 APOAV。其次，我们将探讨 AAV2/8-APOAV 治疗是否可以预防乙醇引发的进展 通过小鼠中的 lysoPC 从简单的脂肪变性到酒精性脂肪肝炎。拟议的研究具有创新性 因为该项目可能为ALD的基因治疗提供有效的策略。