Third generation liver-directed dual-function AAT vectors

第三代肝脏定向双功能 AAT 载体

基本信息

批准号：
9071193
负责人：
Christian Mueller
金额：
$ 50.25万
依托单位：
UNIV OF MASSACHUSETTS MED SCH WORCESTER
依托单位国家：
美国
项目类别：
财政年份：
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/9071193
关键词：
Accounting Algorithms Alleles American Amino Acid Substitution Animal Model Animals Biodistribution Body Size Capsid Cells Clinical Clinical Trials Collaborations DNA cassette Data Dose Elements Engineering Enzymes Epitopes European Generations Genes Genome Goals Hepatocyte Hepatotoxicity Human Immune response Infusion procedures Intravenous Intravenous infusion procedures Investigational New Drug Application Knockout Mice Lead Liver Liver diseases Lung diseases Mediating MicroRNAs Modeling Monitor Monkeys Mus Muscle Mutation Patients Peripheral Phase Phase I Clinical Trials Physiological Plasma Polymers Population Positioning Attribute Primates Procedures Production Protease Inhibitor Proteins Publishing Pulmonary Emphysema RNA Interference Recombinants Regimen Regulatory T-Lymphocyte Resources Respiratory physiology Rodent Model Route Safety Serum Stress Testing Therapeutic Time Tissues Toxic effect Transgenic Mice Translating Viral Virus Receptors Western World abstracting adeno-associated viral vector alpha 1-Antitrypsin alpha 1-Antitrypsin Deficiency base c-myc Genes design gain of function gene therapy intravenous administration intravenous injection knock-down liver biopsy lysylglutamic acid miRNA expression profiling mutant nonhuman primate novel strategies prevent programs research study response secondary outcome therapeutic protein time use transgene expression vector

项目摘要

Project Summary/Abstract Alpha-1 antitrypsin (AAT) is the major circulating serum antiprotease, with normal plasma levels ranging from 11 to 30 microMolar. Approximately 4% of the North American and Northern European populations possess at least one copy of a mutant allele, known as Pi*Z, which results from a single amino acid substitution of Lys for Glu at position 342. In the homozygous state (Pi*ZZ), this mutation leads to severe deficiency of AAT and can result in liver disease which is due to a toxic gain-of-function of the Z-AAT mutant protein. Accumulation of Z- AAT polymers within hepatocytes consistently results in a state of serum AAT deficiency due to inefficient secretion. Two hundred thousand is a conservative estimate of the number of ZZ patients in the western world; however, there is currently no cure for the lung or liver disease. Protein replacement is given by intravenous infusion, with the approved dosing regimen being weekly infusions, and the only alternative dosing with proven equivalence being every-other-week infusions. Thus, gene augmentation with normal (PiM) AAT delivered via recombinant adeno- associated viral (rAAV) vectors has been developed as a potential alternative for “sustained release” of the therapeutic protein. We believe that in order to safely augment AAT from Z-AAT patient livers it may be necessary to silence the endogenous Z-AAT to prevent overwhelming the already stressed hepatocyte. Preliminary data in PiZ transgenic mice suggest that expressing rAAV-derived AAT without first silencing the endogenous Z-AAT can be hepatotoxic. Thus, this project aims to clinically translate a therapeutic strategy that combines traditional gene therapy with RNAi therapeutics in a single rAAV vector approach. We have termed these as dual-function rAAV vectors, and we have engineered them as a “liver- sparing” approach to target the burdened livers of PiZ AATD patients. Proof-of-concept studies in PiZ mice, confirmed that the dual function vector reduced misfolded AAT and concomitantly augmented serum levels of the normal protein. We hypothesize that endogenous Z-AAT silencing and M-AAT augmentation is feasible and a safe therapeutic approach for treating alpha-1 antitrypsin deficiency. We will test this hypothesis with the following three aims. In aim 1 we will optimize the dual function expression cassette for maximum knockdown and augmentation efficiency. These vectors will then be modified with non- human primate sequences (NHP) for aim 2, where we will assess efficacy of the dual function vector in non- human primates. In aim 3 we will perform a formal GLP pharm/tox and biodistribution study with the clinical version of the vector to prepare for an investigational new drug application with the FDA. Finally aim 4 will investigate the safety of peripheral intravenous delivery of these vectors in patients with alpha-1 antitrypsin deficiency.

项目概要/摘要 Alpha-1 抗胰蛋白酶 (AAT) 是主要的循环血清抗蛋白酶，正常血浆水平范围为大约 4% 的北美和北欧人口拥有 11 至 30 微摩尔。至少有一个突变等位基因的拷贝，称为 Pi*Z，它是由 Lys 的单个氨基酸取代而产生的 342 位的 Glu。在纯合状态 (Pi*ZZ) 下，该突变导致 AAT 严重缺乏，并可由于 Z-AAT 突变蛋白的毒性功能获得而导致肝脏疾病。肝细胞内的 AAT 聚合物由于效率低下，始终导致血清 AAT 缺乏状态二十万是西方世界 ZZ 患者数量的保守估计；然而，目前尚无法通过静脉注射来治疗肺部或肝脏疾病。输注，批准的给药方案是每周输注，并且是经证实的唯一替代剂量相当于每隔一周输注一次，通过正常（PiM）AAT 进行基因增强。重组腺相关病毒（rAAV）载体已被开发为潜在的替代品我们相信，为了安全地增强 Z-AAT 的 AAT。患者肝脏可能需要沉默内源性 Z-AAT，以防止压倒已经存在的 Z-AAT。 PiZ 转基因小鼠的应激肝细胞的初步数据表明表达 rAAV 衍生的 AAT。如果不首先沉默内源性 Z-AAT 可能具有肝毒性，因此，该项目旨在临床转化。在单个 rAAV 载体中将传统基因疗法与 RNAi 疗法相结合的治疗策略我们将这些称为双功能 rAAV 载体，并将它们设计为“肝病毒载体”。 Sparing”方法针对 PiZ AATD 患者的肝脏负担进行了 PiZ 小鼠的概念验证研究。证实双功能载体减少了错误折叠的 AAT，并同时增加了 AAT 的血清水平我们努力确定内源性 Z-AAT 沉默和 M-AAT 增强是否可行。以及治疗 α-1 抗胰蛋白酶缺乏症的安全治疗方法。我们将通过以下三个目标来检验这个假设。在目标 1 中，我们将优化双函数表达式。然后用非修饰盒来修饰这些载体以获得最大的击倒和增强效率。目标 2 的人类灵长类动物序列（NHP），我们将评估双功能载体在非在目标 3 中，我们将针对临床进行正式的 GLP 药物/毒性和生物分布研究。载体的版本，为 FDA 的研究性新药申请做准备最后目标 4 将。研究这些载体在接受 α-1 抗胰蛋白酶治疗的患者中进行外周静脉注射的安全性不足。