CRISPR-Cas Editing as a Genetic Cure for Autosomal Dominant Polycystic Kidney Disease

CRISPR-Cas 编辑作为常染色体显性多囊肾病的基因治疗

• 批准号: 10822502
• 负责人: Demetrios S Maxim
• 金额: $ 32.5万
• 依托单位: NEPHROGEN INC.
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-09-19 至 2024-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10822502
• 关键词: 3' Untranslated Regions; 3-Dimensional; Address; Affect; Age; American; Amyloidosis; Arteries; Attenuated; Autosomal Dominant Polycystic Kidney; Bilateral; Biological Assay; Capsid; Catheterization; Cell Culture Techniques; Cell Separation; Cells; Chronic Disease; Clinical; Clinical Trials; Clustered Regularly Interspaced Short Palindromic Repeats; Collaborations; Cyst; DNA; DNA delivery; Dependovirus; Development; Dialysis procedure; Disease; Disease Progression; End stage renal failure; Epithelial Cells; FDA approved; Family; Fluids and Secretions; Fluorescence-Activated Cell Sorting; Gene Expression; Genes; Genetic; Goals; Grant; Guide RNA; Human; Immunofluorescence Immunologic; In Vitro; Injections; Investigational New Drug Application; Kidney; Kidney Diseases; Kidney Failure; Lead; Life; Ligation; Mediating; Mendelian disorder; Methods; Modeling; Modification; Mus; Mutation; Operative Surgical Procedures; PKD1 gene; PKD2 gene; PKD2 protein; Patients; Persons; Phase; Phase I Clinical Trials; Phenotype; Polyuria; Prealbumin; Procedures; Proliferating; Publishing; Renal Replacement Therapy; Renal function; Reporter; Research Personnel; Sickle Cell Anemia; Small Business Technology Transfer Research; Structure of renal vein; System; Technology; Testing; Therapeutic; Transplantation; Treatment Efficacy; Tubular formation; Universities; Variant; Vasopressins; Viral Vector; alternative treatment; antagonist; base; base editing; curative treatments; delivery vehicle; design; early onset; functional restoration; gene product; gene therapy; in vivo; innovation; insertion/deletion mutation; lead candidate; lead optimization; liver injury; minimally invasive; mouse model; next generation sequencing; novel; polycystic kidney disease 1 protein; pre-clinical; preclinical study; prevent; primary endpoint; prime editing; promoter; receptor; side effect; small molecule; standard of care; success; therapeutic genome editing; tolvaptan; trafficking; transduction efficiency; translational approach

PROJECT SUMMARY Autosomal dominant polycystic kidney disease (ADPKD) is the most common monogenic disorder in the world and affects 650,000 Americans. ADPKD is caused by mutations in Pkd1 or Pkd2 that trigger proliferation and fluid secretion by renal tubular epithelial cells into cysts, which bilaterally enlarge the kidney and lead to progressive loss of kidney function. There is no cure for ADPKD with most patients requiring renal replacement therapy in the form of dialysis or transplantation by age 55 years to sustain life. Gene therapy to restore functional levels of the polycystin-1 (Pkd1) or polycystin-2 (Pkd2) gene products offers a promising, innovative therapeutic approach for ADPKD that could serve as an outright cure. However, the development of ADPKD gene therapy has been hindered by the lack of an efficient DNA delivery vehicle to renal tubular epithelial cells and by a one-size-fits-all gene editing solution. In collaboration with our academic partner, Stanford University, we developed several adeno-associated virus (AAV) capsid variants that efficiently transduce the kidney in vivo. The goal of this project is to perform lead optimization for these novel AAV capsids and use them to establish feasibility for a curative ADPKD gene therapy with CRISPR-Cas editing. Our studies will focus on Pkd1 as a proof-of-concept because Pkd1 accounts for 85% of all clinical ADPKD cases and Pkd1 mutations are associated with an earlier onset of kidney failure (more severe disease) than Pkd2. In Specific Aim #1, we will use base or prime editing to inactivate the miR-17 motif within the 3’ UTR of Pkd1 in both human and mouse ADPKD renal tubular epithelial cells in vitro. Preclinical studies have shown that removing or suppressing this miR-17 motif is a mutation-agnostic approach that reverses and attenuates ADPKD progression. We will evaluate several CRISPR editors, guide RNAs, and delivery vehicles to achieve optimal editing efficiency. We will use our published 3-D cell culture model of cystogenesis to evaluate the phenotypic and gene expression changes induced by CRISPR editing. In Specific Aim #2, we will further optimize in vivo delivery of our AAV capsid variants by testing various surgical modifications to the intra-renal artery injection method and testing new promoters. This delivery method is an attractive translational approach because the vessels can be accessed via conventional catheterization to allow for minimally invasive delivery in humans without requiring surgery. We will use the optimized delivery method identified from these studies to deliver the most efficient editing therapeutics from Aim #1 to ADPKD mouse models. At the conclusion of these studies, we expect to demonstrate feasibility for our ADPKD gene therapy approach and identify a lead candidate to advance for preclinical IND-enabling studies (Investigational New Drug application). Our Phase II STTR proposal will focus on completing these IND-enabling studies to enable us to file our IND application and initiate clinical trials. This potentially, curative therapeutic approach for ADPKD addresses a large unmet need given the lack of curative treatment options and has the potential to fundamentally alter the standard of care.

项目摘要 常染色体显性遗传性多囊肾病（ADPKD）是世界上最常见的单基因遗传病 影响了65万美国人ADPKD是由Pkd 1或Pkd 2突变引起的，这些突变触发增殖， 肾小管上皮细胞分泌液体进入囊肿，使肾脏双侧扩大， 肾功能逐渐丧失ADPKD无法治愈，大多数患者需要肾脏替代 在55岁之前接受透析或移植治疗以维持生命。基因治疗恢复 多囊蛋白-1（Pkd 1）或多囊蛋白-2（Pkd 2）基因产物的功能水平提供了一个有前途的，创新的 ADPKD的治疗方法，可以作为彻底治愈。然而，ADPKD的发展 基因治疗由于缺乏有效的将DNA递送到肾小管上皮细胞的载体而受阻 和一种通用的基因编辑解决方案。与我们的学术伙伴斯坦福大学合作， 我们开发了几种腺相关病毒（腺相关病毒）衣壳变体，可以有效地转导肾脏， vivo.本项目的目标是对这些新型AAV衣壳进行先导优化，并使用它们来 建立了利用CRISPR-Cas编辑的治愈性ADPKD基因疗法的可行性。我们的研究将集中在 Pkd 1作为概念验证，因为Pkd 1占所有临床ADPKD病例的85%，Pkd 1突变 与比Pkd 2更早发生肾衰竭（更严重的疾病）相关。具体目标#1，我们 将使用碱基或引物编辑来在人和哺乳动物中Pkd 1的3' UTR内重复miR-17基序， 小鼠ADPKD肾小管上皮细胞的体外培养。临床前研究表明，去除或 抑制该miR-17基序是一种逆转和减弱ADPKD的突变不可知方法 进展我们将评估几种CRISPR编辑器，指导RNA和递送载体，以实现最佳的 编辑效率。我们将使用我们发表的3-D细胞培养模型来评估表型 和由CRISPR编辑诱导的基因表达变化。在具体目标#2中，我们将进一步优化体内 通过测试对肾内动脉注射的各种手术修饰来递送我们的AAV衣壳变体 方法和测试新的启动子。这种递送方法是有吸引力的转化方法，因为 可以通过传统的导管插入术进入血管 而不需要手术我们将使用从这些研究中确定的优化交付方法来交付 从Aim #1到ADPKD小鼠模型的最有效的编辑疗法。在这些研究结束时， 我们希望证明我们的ADPKD基因治疗方法的可行性，并确定一个领先的候选人， 推进临床前IND使能研究（研究性新药申请）。我们的第二阶段STTR 提案将侧重于完成这些IND使能研究，使我们能够提交IND申请， 启动临床试验。这种潜在的ADPKD治愈性治疗方法解决了大量未满足的需求 由于缺乏治愈性治疗选择，并有可能从根本上改变护理标准。