Novel adeno-associated virus vector production system development

新型腺相关病毒载体生产系统的开发

基本信息

批准号：
8311963
负责人：
WEIDONG XIAO
金额：
$ 45.07万
依托单位：
TEMPLE UNIV OF THE COMMONWEALTH
依托单位国家：
美国
项目类别：
财政年份：
2012
资助国家：
美国
起止时间：
2012-02-15 至 2017-01-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8311963
关键词：
Abbreviations Adenovirus Vector Adenoviruses Adopted Animal Experiments Baculoviruses Biomedical Engineering Bioreactors Cell Culture Techniques Cell Line Cell Nucleus Cells Centrifugation Clinical Clinical Trials Cyclic GMP DNA Data Dependovirus Development Disease Dose Engineering Face Factor IX Future Gene Expression Gene Transduction Agent Generations Genes Genetic Recombination Genome Goals HeLa S3 Helper Viruses Hemophilia A Hemophilia B Hereditary Disease Herpesviridae Human Inverted Terminal Repeat Leber&apos s amaurosis Mediating Mus Patients Plasmids Procedures Process Production Protocols documentation Quality of life RPE65 protein Recombinant adeno-associated virus (rAAV)Recombinants Reporting Research Research Project Grants Role Safety Serotyping Single Stranded DNA Virus Suspension substance Suspensions System Systems Development TCF3 gene Technology Therapeutic Transfection Transgenes Vaccinia Vaccinia virus Vaccinium Viral Virion Virus Replication Vision adeno-associated viral vector base bioprocess cost effective flexibility follow-up gene therapy human subject improved innovation large scale production manufacturing process meetings novel particle promoter response tool transgene expression vector

Abbreviations Adenovirus Vector Adenoviruses Adopted Animal Experiments Baculoviruses Biomedical Engineering Bioreactors Cell Culture Techniques Cell Line Cell Nucleus Cells Centrifugation Clinical Clinical Trials Cyclic GMP DNA Data Dependovirus Development Disease Dose Engineering Face Factor IX Future Gene Expression Gene Transduction Agent Generations Genes Genetic Recombination Genome Goals HeLa S3 Helper Viruses Hemophilia A Hemophilia B Hereditary Disease Herpesviridae Human Inverted Terminal Repeat Leber&apos s amaurosis Mediating Mus Patients Plasmids Procedures Process Production Protocols documentation Quality of life RPE65 protein Recombinant adeno-associated virus (rAAV)Recombinants Reporting Research Research Project Grants Role Safety Serotyping Single Stranded DNA Virus Suspension substance Suspensions System Systems Development TCF3 gene Technology Therapeutic Transfection Transgenes Vaccinia Vaccinia virus Vaccinium Viral Virion Virus Replication Vision adeno-associated viral vector base bioprocess cost effective flexibility follow-up gene therapy human subject improved innovation large scale production manufacturing process meetings novel particle promoter response tool transgene expression vector

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项目摘要

DESCRIPTION (provided by applicant): AAV is a single stranded DNA virus which has shown great promise as a gene therapy vector. Recombinant AAV vectors delivering RPE65 gene have been demonstrated to cure Leber's congenital amaurosis (LCA) disease and patients are able to regain normal vision after receiving the vectors. In hemophilia B clinical trial, patient receiving AAV vectors were able to express therapeutic level of factor IX gene expression. Systemic delivery of rAAV into human subjects would require a lot more vectors. We anticipate the demand for high quality rAAV vector for human clinical trials will exceed the current capability in our field. To face this challenge, we propose to improve rAAV vector production utilizing a new concept to eliminate wild type replication rcAAV particles and engineer scalable manufacturing processes. We would also investigate the complimentary helper function of adenovirus over cytoplasmic carrier helper in facilitating rAAV replication and packaging. In order to further improve the quality of recombinant AAV production for human gene therapy, we hypothesize that unwanted replication competent AAV particle formation could be eliminated by sequestering the helper function and vector sequences into different cellular compartments. The production and process system will eliminate plasmid transfection and other procedures which are not feasible for large scale production. Hence, our three specific aims are: 1. To study enhancement of AAV encapsidation by adenovirus in a cytoplasmic helper packaging system; 2. To develop a rAAV production system utilizing a cytoplasmic rep/cap carrier; 3. To engineer a protocol for scalable production of rAAV vectors free of contaminants. Completion of these specific aims will greatly enhance the way of rAAV vectors to be used in the field of human gene therapy. PUBLIC HEALTH RELEVANCE: The completion of this project will improve AAV production technology and allow a high quality rAAV vector to be produced economically in large scale. It has an immediate impact on human gene therapy and has the potential to improve the quality of life of patients with genetic diseases, especially for hemophilia which requires systemic delivery of vectors.

描述（由申请人提供）：AAV是一种链的DNA病毒，它作为基因治疗载体表现出了很大的希望。已证明输送RPE65基因的重组AAV载体可以治愈Leber的先天性症（LCA）疾病，并且患者在接受载体后能够恢复正常视力。在血友病B临床试验中，接受AAV载体的患者能够表达因子IX基因表达的治疗水平。将RAAV的系统交付给人类受试者将需要更多的向量。我们预计，对人类临床试验的高质量RAAV载体的需求将超过我们领域的当前能力。为了面对这一挑战，我们建议利用一种新概念来改善RAAV矢量生产，以消除野生型复制RCAAV颗粒和工程师可扩展的制造过程。我们还将研究腺病毒在促进RAAV复制和包装方面的腺病毒的免费辅助功能。为了进一步提高重组AAV产生的人类基因治疗的质量，我们假设可以通过将辅助功能和矢量序列隔离为不同的细胞隔室来消除不必要的复制胜任的AAV颗粒形成。生产和过程系统将消除质粒转染和其他对于大规模生产而言不可行的过程。因此，我们的三个具体目标是： 1。研究腺病毒在细胞质辅助包装系统中对AAV封装的增强； 2。使用细胞质rep/cap载体开发RAAV生产系统； 3。设计一种可扩展生产RAAV载体的协议，没有污染物。这些特定目标的完成将大大增强RAAV向量用于人类基因疗法领域的方式。公共卫生相关性：该项目的完成将改善AAV生产技术，并使高质量的RAAV媒介在经济上大规模生产。它对人类基因疗法有直接的影响，并有可能改善遗传疾病患者的生活质量，尤其是对于需要全身性载体的血友病。