Targeted delivery of microRNA-loaded microvesicle for cancer therapy

描述（由申请人提供）：一种新的治疗方式是使用寡核苷酸将肿瘤抑制mirna的水平恢复到正常细胞或组织的水平。治疗性寡核苷酸的成功靶向和递送一直是其临床发展的主要瓶颈。微泡是内吞起源的膜泡的一种小的同质亚型，天然含有多种细胞生化物质，包括miRNA。该方案将开发一种靶向治疗性mirna的微囊药物递送系统，用于肝癌细胞和肿瘤。该系统的一个独特之处在于核酸货物由产生微泡的细胞合成，从而减轻了对合成寡核苷酸的需求。微泡将被设计成表达一种靶向肽，将其导向癌细胞。修饰后的pre-miR-199a经过工程设计，包含了TAR RNA发夹的环区，将被插入到靶蛋白基因的内含子中，一旦剪接和加工，将通过结合靶蛋白上的c端HIV Tat肽被引导到微泡中。这个项目将使用在硅和生化方法

DESCRIPTION (provided by applicant): A new therapeutic modality is to use oligonucleotides to restore the levels of tumor suppressive miRNAs to that of normal cells or tissues. Successful targeting and delivery of therapeutic oligonucleotides has been a major bottleneck in their clinical development. Microvesicles are a small homogenous subtype of membrane vesicles of endocytic origin and naturally contain a variety of cellular biochemicals including miRNA. This proposal will develop an microvesicle drug delivery system to target therapeutic miRNAs to hepatocellular carcinoma cells and tumors. A unique feature of this system is that the nucleic acid cargo is synthesized by the cells that produce the microvesicles, thus alleviating the need for synthetic oligonucleotides. Microvesicles will be engineered to express a targeting peptide that will direct it to cancer cells. The modified pre-miR-199a, engineered to include the loop region of the TAR RNA hairpin, will be inserted into an intron of the targeting protein gene and once spliced and processed, will be directed to the microvesicle by binding to C-terminus HIV Tat peptide on the targeting protein. This project will use in silico and biochemical approaches to determine the optimal pre-miRNA sequences for peptide binding, correct processing and biological activity. The therapeutic activity and targeting ability of the microvesicle delivery system will be evaluated in vitro and in an orthotopic model of hepatocellular carcinoma. Pharmacodynamic and pharmacokinetic evaluations will provide detailed knowledge on dose, toxicity, efficacy, route of administration and biodistribution. Proposed here is a highly innovatie approach to synthesize and deliver therapeutic nucleic acids to cancer cells. This system does not rely upon the use of synthetic oligonucleotides or artificial drug delivery systems. Future manifestations of this technology can be applied to deliver other nucleic acid drugs such as shRNA and may be widely applicable to treat many diseases.