腺相关病毒在脂代谢研究和降脂基因治疗中的应用

The Application of Adeno-asscociated Virus in Lipid Metabolism Research and Lipid-lowering Gene Therapy

动脉粥样硬化性心脑血管疾病是当前全球死亡率最高的疾病,而脂代谢紊乱是动脉粥样硬化性心脑血管疾病的主要病因,其既可以导致心肌梗死、脑卒中、急性胰腺炎等急性病,也可以导致慢性肾脏疾病。近年来基因治疗技术的快速发展,为脂代谢机制研究提供了有效的手段,特别是使先天性脂代谢异常患者的治愈成为可能。腺相关病毒(adeno associated virus,AAV)宿主范围广、安全性高、免疫原性低、表达长期稳定,是当前单基因遗传病基因治疗首选的递送工具。以AAV为载体的多种降脂基因治疗药物目前已经得到临床应用或正在进行临床试验。本文综述了AAV载体在脂质代谢机制研究的新进展及其在降脂基因治疗中的应用和前景。

Cardiovascular and cerebrovascular diseases,usually result from atherosclerosis,has the highest mortality rate globally.Lipid metabolism disorder is the main cause of atherosclerotic cardiovascular and cerebrovascular diseases,which not only lead to acute diseases such as myocardial infarction,stroke,acute pancreatitis,but also chronic kidney disease.In recent years,the advancement of gene therapy technologies has provided novel means for lipid metabolism study,and has also made it possible to cure patients with congenital lipid metabolism abnormalities.Adeno-associatd virus has a wide host range,high safety,low immunogenicity,and especially the ability of long-term stable expression in vivo,making it the preferred delivery tool for gene therapy of monogenic genetic diseases.Alipogene triprivec,also known as Glybera,was approved by the European Medicines Agency in 2012.It is the first gene therapy drug that uses recombinant AAV1 vector to directly deliver a highly active LPL protein S447X mutant to muscle cells for the treatment of patients with hereditary LPL deficiency.To enhance the targeted transduction efficiency of AAV carriers,recombinant AAV8.TBG.hLDLR utilizes the tissue tropsim of AAV8 to liver,meanwhile utilizes a liver specific thyroxine binding globulin promoter to control gene transcription,thereby achieving liver cell specific high expression of human low-density lipoprotein receptors(LDLR).In patients with familial hypercholesterolemia,AAV8.TBG.hLDLR treatment effectively lower the level of plasma LDL for a long time,thus preventing the occurrence of atherosclerosis.Proprotein convert subunit kexin 9(PCSK9)is secreted by liver cells.PCSK9 binds and transports LDLR to lysosomes for degradation,preventing the circulation and regeneration of LDLR,leading to accelerated degradation of LDLR and finally resulting in the accumulation of low-density lipoprotein cholesterol in plasma.Using AAV to deliver Cas9 of Staphylococcus aureus and gRNA targeting the Pcsk9 gene can knock out Pcsk9 in mouse liver,leading to a long-term significant decrease in plasma cholesterol levels in mice.Hepatocyte specific angiopoietin related protein 3(Angptl3)is an endogenous inhibitor of LPL.Using the AAV9 mediated AncBE4max system and the dCas9 mediated single base gene editing system to introduce early termination codons,the knockout of Angptal3 in liver cells was achieved with an average knockout efficiency of 63.3%.After 2-4 weeks of administration in mice,the Angptl3 protein was completely undetectable in the peripheral blood,and serum triglycerides and total cholesterol decreased by 58%and 61%,respectively.Ring finger containing protein 130(RNF130)is an E3 ubiquitin ligase.Research has shown that overexpression of RNF130 using AAV2/8 leads to ubiquitination degradation and redistribution of LDLR on the cell membrane,significantly reducing LDLR expression on liver cells and increasing plasma LDLC levels,while knocking out Rnf130 gene using the AAV-CRISPR system results in the opposite effect.This AAV mediated RNF130 function study proves that RNF130 is a posttranslational regulatory protein of LDLR and plays an important role in the regulation of serum LDLC.As mentioned above,recently,various lipid-lowering gene therapy drugs carried by different serotypes of adeno-associated virus have been applied in clinic or are undergoing clinical trials,and adeno-associated virus has emerging to be an important tool for lipid metabolism research.This article reviews the new progress of adeno-associated virus vectors in lipid metabolism study and lipid-lowering gene therapy.