Endonuclease-mediated disruption of latent HSV as curative therapy

核酸内切酶介导的潜伏 HSV 破坏作为治疗方法

基本信息

批准号：
10182099
负责人：
KEITH R JEROME
金额：
$ 44万
依托单位：
FRED HUTCHINSON CANCER RESEARCH CENTER
依托单位国家：
美国
项目类别：
财政年份：
2018
资助国家：
美国
起止时间：
2018-06-12 至 2023-05-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10182099
关键词：
Address Afferent Neurons Animal Model Animals Antiviral Agents Antiviral Therapy Area Autonomic ganglion CRISPR/Cas technology Cell Death Cells Chronic Clinic Clinical DNA DNA Integration Data Dependovirus Development Disease Enzymes Evaluation Excision Frequencies Ganglia Gene Delivery Genes Genome Genomics Goals HIV Health behavior Hepatitis B Virus Herpesvirus Type 3 Homing Human Genome Human Papillomavirus Immunochemistry Individual Infiltration Inflammatory Injections Kinetics Laboratories Latent virus infection phase Lead Measures Mediating Morbidity - disease rate Mus Mutagenesis Mutate Mutation Neurons Pathway interactions Phenotype Primary Infection Process Recurrence Recurrent disease Reporter Reproducibility Route Safety Sampling Sensory Ganglia Serotyping Shock Simplexvirus Site Source Staphylococcus aureus System Technology Testing Therapeutic Therapeutic Effect Time Treatment Efficacy Vaccines Vibrissae Viral Viral Genome Viral Load result Viral Pathogenesis Viral reservoir Virus Virus Diseases Virus Integration Virus Latency Virus Replication Work clinical application clinical development curative treatments endonuclease genome-wide analysis genotoxicity improved in vivo innovation insertion/deletion mutation mouse genome mouse model next generation sequencing novel therapeutic intervention nuclease preclinical development promoter reactivation from latency screening targeted nucleases therapeutic development transgene delivery transgene expression transmission process vector viral DNA virology

项目摘要

Project Summary Herpes simplex virus (HSV) infections remain a common, serious problem associated with significant morbidity. After primary infection, HSV establishes latency, which is not eliminated by current antiviral therapy. Latent virus is the source for viral reactivation and the recurrence of clinical disease. Despite much effort, a vaccine remains elusive. Therefore, there is a need for a novel therapeutic approach that would cure latent HSV infection. We have been developing a new curative strategy to latent HSV infection, in which an HSV-targeted endonuclease induces mutagenesis of essential HSV genes, disabling viral genomes and rendering the virus incapable of replication or reactivation from latency. Our hypothesis is that recent advances in the field of gene-editing technologies and in vivo gene delivery offer the opportunity to improve our current anti-viral approach and reach therapeutic efficacy. Here, we propose to evaluate Staphylococcus aureus (Sa)CRIPSR/Cas9 and homing endonucleases (HEs) for their ability to disable HSV in latently infected neurons by targeted viral genome disruption, thus eliminating the source of viral pathogenesis in an animal model of HSV infection. The goal of this project is to maximize the efficacy and safety of our approach to eliminate latent HSV infection in vivo, using a murine model of HSV latent infection. Our results will also be applicable in efforts to cure varicella zoster virus, another alphaherpesvirus that like HSV establishes latency in sensory neurons. Furthermore, the data generated will be highly relevant to the development of cures for other chronic or latent viral infections such as hepatitis B virus, HIV, or human papillomavirus. In SA1: Address the remaining barriers to effective in vivo HSV gene editing, we will compare 1) new AAV serotypes to our current AAV serotypes for transgene delivery after administration via different routes, and 2) the gene editing abilities of CRISPR/Cas9 vs. HEs. In SA2: Optimize the efficacy of in vivo gene editing and determine the impact on viral pathogenesis, we will evaluate whether the simultaneous targeting of two HSV sites provides superior efficacy over targeting a single site, and the efficacy of gene editing necessary to impact viral pathogenesis. In SA3: Evaluate the safety of in vivo gene editing in our mouse model, we will use our mouse model of latent HSV infection to evaluate tolerability, safety, and genotoxicity of nuclease exposure. This project is expected to demonstrate the feasibility of our therapeutic approach directed towards the elimination of HSV pathogenesis in vivo, and to provide critical information for the development of a larger scale animal study necessary to bring this new therapeutic approach to the clinic.

项目概要单纯疱疹病毒 (HSV) 感染仍然是一个常见且严重的问题，与重大疾病相关发病率。原发感染后，HSV 会建立潜伏期，目前的抗病毒治疗无法消除这种潜伏期。潜伏病毒是病毒再激活和临床疾病复发的根源。尽管付出了很多努力，疫苗仍然难以捉摸。因此，需要一种新的治疗方法来治愈潜伏性HSV感染。我们一直在开发一种针对潜伏 HSV 感染的新治疗策略，其中 HSV 靶向核酸内切酶诱导 HSV 必需基因突变，使病毒基因组失活，使病毒无法复制或从潜伏期重新激活。我们的假设是，最近的进展在基因编辑技术和体内基因传递领域，提供了改善我们现有技术的机会抗病毒方法并达到治疗效果。在这里，我们建议评估金黄色葡萄球菌 (Sa)CRIPSR/Cas9 和归巢核酸内切酶 (HE) 能够在潜伏感染中禁用 HSV 通过有针对性的病毒基因组破坏来破坏神经元，从而消除动物体内病毒发病机制的来源 HSV感染模型。该项目的目标是最大限度地提高我们方法的有效性和安全性使用HSV潜伏感染的小鼠模型消除体内潜伏的HSV感染。我们的结果也将是适用于治疗水痘带状疱疹病毒，这是另一种与 HSV 类似的 α 疱疹病毒，在感觉神经元。此外，生成的数据将与其他治疗方法的开发高度相关。慢性或潜伏的病毒感染，例如乙型肝炎病毒、艾滋病毒或人乳头瘤病毒。在 SA1：解决有效体内 HSV 基因编辑的剩余障碍中，我们将比较 1) 新将 AAV 血清型改为我们当前的 AAV 血清型，通过不同途径给药后进行转基因递送， 2) CRISPR/Cas9 与 HE 的基因编辑能力。 SA2中：优化体内基因功效编辑并确定对病毒发病机制的影响，我们将评估是否同时靶向两个 HSV 位点比靶向单个位点具有更优异的功效，并且基因的功效影响病毒发病机制所必需的编辑。在 SA3 中：评估我们体内基因编辑的安全性小鼠模型，我们将使用潜伏 HSV 感染的小鼠模型来评估耐受性、安全性和核酸酶暴露的遗传毒性。该项目预计将证明我们针对以下疾病的治疗方法的可行性消除HSV体内发病机制，并为更大范围的开发提供关键信息将这种新的治疗方法引入临床所必需的大规模动物研究。