Developing Nanopieces, a Platform RNAi Delivery Technology for Treatment of Multiple Diseases

发展中的纳米粒子 7.项目摘要/摘要 开发新型RNAi药物的瓶颈是缺乏高效无毒的药物 体内非肝组织的RNA递送。输送带负电荷的核酸尤其具有挑战性。 在难以到达的组织中，包括关节软骨中，转化为无血管的、致密的和带负电荷的基质。纳米De 治疗公司(NanoDe)是一家致力于开发RNAi递送的发展阶段公司 治疗，从而创造出更有效的药物。NanoDe建立在一种新型的RNA递送平台上 名为NanopiecesTM的技术，这是一种从小分子JBAK衍生的新型仿生纳米材料， Janus-含胺或赖氨酸(K)的碱。通过模仿其生物相容的Janus-Base单位的自我相互作用 DNA碱基对，JBAK与带正电荷的胺或赖氨酸形成非共价纳米管(NT 浮出水面。JBAK NT进一步与siRNA组装形成JBAK NP，从而包裹带负电荷 SiRNA转化为带正电荷的NP。我们的初步数据表明：1)NPs可以穿透富含基质的物质 常规载体无法获得的组织，并高效地在细胞内释放siRNA疗法 修改其他无法治疗的疾病；2)NP递送的RNAi治疗取得了成功 在多种疾病的治疗中，包括类风湿性关节炎，一种罕见的实体肿瘤，称为软骨肉瘤， 和创伤后骨关节炎(PTOA)的动物模型；3)NP具有优良的 生物相容性和生物降解性，这对维持体内最低毒性至关重要。这样做的目的是 第一阶段的应用是通过将PTOA作为一种“用例”疾病来进一步开发NP交付技术。这个 中心假设是NP的最佳正电荷是使其成为可能的关键参数之一 渗透到带负电的软骨基质，内吞到软骨细胞，释放siRNA到 抑制疾病基因的表达，从而对PTOA取得显著的治疗效果。这一假说将 测试有两个目的：1)确定NP的最佳装药量，以使其能够渗透和保留 软骨组织内，转染软骨细胞，抑制基质蛋白水解酶ADAMTS-5基因 分别在软骨细胞中表达；2)确定NP的最适电荷，使其能够在软骨细胞内表达。 关节给药和关节内的长半衰期，并在PTOA动物身上取得显著的治疗结果 模特。拟议的研究具有创新性，因为：1)与传统的交付方式有本质的区别 作为载体，Nanopiess是一种非共价载体，具有独特的优势，例如在维度上的多功能性 和表面电荷，与细胞外基质的亲和力，良好的生物降解性和生物相容性。2)对于 我们首次确定了成功传递siRNA所需的NP的重要技术参数，包括 基质穿透、细胞转染和基因敲除的最佳载体表面电荷。3) 技术突破为治疗多种疾病提供了一种RNAi的治疗方法 包括PTOA。

STTR_Developing Nanopieces 7. Project Summary/Abstract The bottleneck of developing new RNAi (RNA Interference) drugs is the lack of highly efficient and non-toxic RNA delivery to non-liver tissues in vivo. It is especially challenging to deliver negatively-charged nucleic acid into avascular, dense, and negatively-charged matrix in hard-to-reach tissues including joint cartilage. NanoDe Therapeutics, Inc. (NanoDe) is a development stage company dedicated to developing delivery of RNAi therapeutics, thereby creating more effective drugs. NanoDe is founded on a novel platform RNA delivery technology termed NanopiecesTM, a novel biomimetic nanomaterial derived from a small molecule JBAK, Janus-Base with Amine or lysine (K). Through self-interaction of its biocompatible Janus-Base units mimicking DNA base pairs, JBAK forms non-covalent nanotubes (NT) with positively charged amine or lysine on the surface. JBAK NT further assembles with siRNA to form JBAK NP, thereby encapsulating negatively charged siRNA into positively charged NP. Our preliminary data has shown that 1) NPs can penetrate matrix-rich tissues that conventional vehicles cannot, and release siRNA therapeutics intracelluarly in high efficiency to modify otherwise untreatable diseases; 2) NP delivered RNAi therapeutics has achieved successful outcomes in the treatment of multiple diseases including rheumatoid arthritis, a rare solid tumor called chondrosarcoma, and post-traumatic osteoarthritis (PTOA) in animal models respectively; and 3) NP has excellent biocompatibility and biodegradability, which are critical for maintaining minimal toxicity in vivo. The goal of this Phase I application is to further develop NP delivery technology by using PTOA as a “use case” disease. The central hypothesis is that the optimal positive charge of NP is one of the critical parameters to enable its penetration into negatively charged cartilage matrix, endocytosis into chondrocytes, and release siRNA to inhibit disease gene expression, thereby achieving significant therapeutic effects on PTOA. This hypothesis will be tested with the two aims: 1) Determining the optimal charge of NP to enable its penetration and retention within cartilage tissue, transfection into chondrocytes, and inhibiting matrix proteinase ADAMTS-5 gene expression in chondrocytes, respectively; and 2) Determining the optimal charge of NP to enable its intra- articular delivery and long half-life within joint, and achieve significant therapeutic outcomes in a PTOA animal model. The proposed research is innovative because: 1) Essentially different from conventional delivery vehicles, Nanopiece is a non-covalent vehicle presenting unique advantages, such as versatility in dimensions and surface charge, affinity to extracellular matrix, excellent biodegradability and biocompatibility. 2) For the first time, we identify important technology parameters of NP required for successful siRNA delivery including the optimal vehicle surface charge for matrix penetration, cell transfection, and gene knockdown. 3) The technology breakthrough enlightens a therapeutic approach to deliver RNAi for treatment of multiple diseases including PTOA.