Silica Nanocapsule-Mediated Nonviral Delivery of CRISPR Base Editor mRNA and Allele Specific sgRNA for Gene Correction in Leber Congenital Amaurosis

二氧化硅纳米胶囊介导的 CRISPR 碱基编辑器 mRNA 和等位基因特异性 sgRNA 非病毒传递用于 Leber 先天性黑蒙的基因校正

• 批准号: 10668166
• 负责人: SHAOQIN GONG
• 金额: $ 186.61万
• 依托单位: UNIVERSITY OF WISCONSIN-MADISON
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-05-16 至 2028-04-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10668166
• 关键词: 3-Dimensional; Acceleration; Alleles; Animals; Biological Products; Biomanufacturing; Blindness; Cells; Cellular Assay; Chemicals; Chemistry; Childhood; Clinical; Clustered Regularly Interspaced Short Palindromic Repeats; Collaborations; Data; Development; Dose; Drug Packaging; Engineering; Eye; FDA approved; Formulation; Future; Genes; Good Manufacturing Process; Guide RNA; Human; Immune; In Vitro; Inherited; Investigational Drugs; Investigational New Drug Application; Kir7.1 channel; Knowledge; Lead; Leber' s amaurosis; Ligands; Mediating; Messenger RNA; Methods; Modality; Mus; Particle Size; Patients; Pharmaceutical Preparations; Point Mutation; Prevalence; Process; Production; Property; Quality Control; Rattus; Research Contracts; Research Methodology; Retina; Retinal Diseases; Running; Safety; Silicon Dioxide; Specificity; Structure of retinal pigment epithelium; Surface; Technology Transfer; Testing; Therapeutic Effect; Toxic effect; Toxicology; Translations; Tretinoin; Validation; Vision; base editing; base editor; biological systems; biomaterial compatibility; cGMP production; cell type; clinical translation; functional restoration; gene correction; gene therapy; genome editing; genotoxicity; in vivo; induced pluripotent stem cell; inward rectifier potassium channel; lead candidate; manufacture; manufacturing facility; manufacturing scale-up; meetings; mouse model; mutation correction; nanocapsule; nonhuman primate; novel therapeutics; pre-Investigational New Drug meeting; pre-clinical; preclinical study; process optimization; programs; rare condition; safety study; scale up; small molecule; subretinal injection; success; therapeutic genome editing

PROJECT SUMMARY/ABSTRACT - LEAD TRAILBLAZER PROJECT 1 Leber Congenital Amaurosis (LCA) is a rare but severe form of pediatric blindness. One subtype, LCA16, is caused by several single-point mutations in the KCNJ13 gene, which encodes the inwardly-rectifying potassium channel Kir7.1 in the retina. There is no FDA-approved treatment for ultra-rare conditions such as LCA16. The objective of Lead Project 1 is to develop a new LCA16 gene therapy utilizing a CRISPR base editor (BE) delivered to the retinal pigment epithelium (RPE) via a nonviral silica nanocapsule (SNC). The SNCs possess many desirable properties, including high delivery efficiency, versatile surface chemistry for ligand conjugation, small particle sizes, good biocompatibility, and scalable production. In preliminary studies, we show that the SNC can transiently deliver a wide range of biologics, including a BE to RPE cells in mice and LCA16 patient-derived induced pluripotent stem cell (iPSC)-RPE. Within five years, we seek to optimize and validate a lead candidate, SNC-101, progress to scaled-up and CGMP production of the product to enable non-human primate (NHP) studies and file an Investigational New Drug (IND) application to the FDA for base editing therapy of LCA16. In Aim 1, we will generate a preclinical validation package with a lead SNC formulation for W53* KCNJ13 correction. We will first optimize the amount of ATRA targeting ligand (RPE cell-specific) and the modified KCNJ13 sgRNA in vitro via the Human Cell Assays Core and in vivo in a W53* LCA16 mouse model. We will then develop a scale-up production process for the optimized SNC (i.e., SNC101) for preclinical studies. We plan to complete one INTERACT meeting at the end of this aim. In Aim 2, we will determine the gene correction efficiency, dosing, and toxicity in mouse models and NHPs. We will comprehensively evaluate the immune, structural, and functional consequences of subretinal delivery of SNC-101 through our W53* LCA16 mouse model. Using our Large Animal Core, we will also perform dose-escalation safety studies in NHPs. Alongside the species comparison, the study will ascertain storage stability, dosing, and toxicity profiles in detail, an important milestone to support an IND with the FDA. In Aim 3, we will conduct IND enabling studies of SNC-101 for W53* KCNJ13 correction. A pre-IND meeting package will be submitted to FDA CBER with the help of our Regulatory Core. Waisman Biomanufacturing will develop a full-scale engineering run to be used in GLP toxicology study in rats and NHPs. Concurrent with the toxicology study, one entire CGMP batch will be manufactured. We will submit an IND application to the FDA in the final year. To date, no nonviral genome editing therapeutic has reached an IND in the eye. Success here with SNC-101 would pave the way for this new therapeutic modality. Finally, the knowledge gained along this development path will accelerate the translation of other nonviral genome editing leads, including SNC-201 and TAGE-101, in the Follower Projects of the CRISPR Vision Program.

项目摘要/摘要 - 领先开拓者项目1 Leber先天性瘤（LCA）是一种罕见但严重的小儿失明形式。一种亚型，LCA16，是 由KCNJ13基因中的几个单点突变引起 视网膜中的Kir7.1通道。对于LCA16等超稀有条件，没有FDA批准的治疗方法。这 铅项目1的目的是利用CRISPR基本编辑器（BE）开发一种新的LCA16基因疗法 通过非病毒二氧化硅纳米胶囊（SNC）传递到视网膜色素上皮（RPE）。 SNC拥有 许多理想的特性，包括高递送效率，用于配体共轭的多功能表面化学， 小粒径，良好的生物相容性和可扩展的产生。在初步研究中，我们证明了SNC 可以瞬时提供广泛的生物制剂，包括小鼠和LCA16患者衍生的RPE细胞 诱导多能干细胞（IPSC）-RPE。在五年之内，我们寻求优化和验证主要候选人， SNC-101，产品的扩展和CGMP产生，以实现非人类灵长类动物（NHP） 研究并向FDA提出了研究新药（IND），以用于LCA16的基础编辑疗法。在 AIM 1，我们将生成一个具有W53* KCNJ13校正的铅SNC公式的临床前验证包。 我们将首先优化靶向配体的ATRA量（RPE特异性）和修饰的KCNJ13 SGRNA 通过人体细胞分析在W53* LCA16小鼠模型中进行体外体外。然后，我们将开发一个 临床前研究的优化SNC（即SNC101）的扩展生产过程。我们计划完成 此目标结束时的一次互动会议。在AIM 2中，我们将确定基因校正效率，给药， 和小鼠模型和NHP中的毒性。我们将全面评估免疫，结构和 通过我们的W53* LCA16小鼠模型将视网膜下递送的功能后果。使用我们的 大动物核心，我们还将在NHP中进行剂量降低安全性研究。与物种旁边 比较，该研究将详细确定储存稳定性，给药和毒性概况，这是一个重要的里程碑 用FDA支持IND。在AIM 3中，我们将对W53* KCNJ13的SNC-101进行IND允许研究 更正。在我们的监管核心的帮助下，将向FDA CBER提交预定的会议计划。 Waisman生物制造将开发全面的工程，用于在大鼠的GLP毒理学研究中使用 和NHP。与毒理学研究同时，将制造整个CGMP批次。我们将提交 最后一年对FDA的IND申请。迄今为止，尚无非病毒基因组编辑治疗尚未达到 在眼中。 SNC-101的成功将为这种新的治疗方式铺平道路。最后， 沿着这种发展路径获得的知识将加速其他非病毒基因组编辑的翻译 在CRISPR Vision计划的追随者项目中，包括SNC-201和Tage-101在内的潜在客户。