Exosome based therapeutics in Huntington's disease

描述（由申请人提供）：UH 2和UH 3的目的是研究外泌体如何递送siRNA穿过血脑屏障进入神经元和其他脑细胞。直接靶点是突变的亨廷顿蛋白mRNA。亨廷顿氏病（HD）是由CAG三核苷酸重复序列增加到36个引起的;由于首先在纹状体和皮质中发生神经变性，然后转移到其他大脑结构，因此需要在高级护理设施中多年。HD患者存在认知障碍、抑郁和异常运动。大多数HD患者在30至40岁时出现;少数患者在幼年时发病。合理的治疗是减少突变亨廷顿蛋白mRNA的表达;这种治疗可以通过siRNA、反义寡核苷酸（阿索）和腺相关病毒（AAV）与针对亨廷顿蛋白mRNA的shRNAmir在HD小鼠模型中完成。然而，递送仍然是治疗的实际实施的陷阱。siRNA和阿索需要长期输注。在非人灵长类动物中，给予脊髓液的阿索不能到达纹状体，siRNA在脑中的扩散受到限制。尽管AAV-shRNA很有前途，但它需要多次注射到大脑区域，而且shRNAmir是不受调节的。HD治疗剂中的空白可以通过通常由细胞挤出的微囊泡、外泌体来填补。在其表面上具有狂犬病病毒糖蛋白（RVG）的外来体可以被注射到血液中，穿过血脑屏障，并进入神经元和神经胶质。RVG-外来体可以携带siRNA货物。外泌体进入血液循环后，会在神经元中沉积存款siRNA，从而参与RNA干扰。我们的目的是开发外泌体作为HD的治疗剂。UH 2检查了携带针对亨廷顿蛋白mRNA的siRNA的RVG-外泌体穿过血脑屏障进入神经元的能力。将研究脑中的定位和RNAi依赖性敲低。将寻求超功能siRNA。由于外来体由细胞的细胞质制成，因此外来体mRNA、miRNA和植入的siRNA将通过深度测序来鉴定。将研究免疫反应性和免疫中和，因为外来体具有潜在的抗原，如RVG，并且需要经常施用。UH 3通过研究HD小鼠模型中神经病理学和异常运动的逆转或预防，进一步建立了基于外泌体的治疗方法。将确保外泌体的给药。HD发病机制，siRNA开发，RNA鉴定和测量，RNAi机制和外泌体生产以及大脑递送方面的专家团队将进行研究。利用外泌体进行脑递送有望形成一种可行的治疗方法，以减少HD患者中突变亨廷顿蛋白的表达。患有其他遗传性神经变性的患者将受益。

DESCRIPTION (provided by applicant): The goal of this UH2 and UH3 is to study how exosomes can deliver siRNAs across the blood brain barrier to enter neurons and other brain cells. The immediate target is the mutant huntingtin mRNA. Huntington's disease (HD) is caused by an increase in the CAG trinucleotide repeats to ¿ 36 in series; it necessitates years in a high level nursing facility because of neurodegeneration first in striatum and cortex and then to other brain structures. HD patients have cognitive impairment, depression and aberrant movements. Most HD patient present by 30 to 40 years of age; a few have a juvenile onset. A rational treatment is to decrease expression of mutant huntingtin mRNA; this therapeutic can be accomplished in HD mouse models by siRNA, antisense oligonucleotides (ASO) and adeno-associated virus (AAV) with shRNAmir directed against huntingtin mRNA. However, delivery remains a pitfall to practical implementation of the therapeutics. siRNA and ASO require long-term infusion. In non-human primates, ASO administered to spinal fluid does not reach the striatum and spread of siRNA is limited in brain. Although promising, AAV-shRNA requires several injections into brain areas and the shRNAmir is unregulated. A gap in HD therapeutics can be filled by microvesicles normally extruded by cells, exosomes. Exosomes with rabies virus glycoprotein (RVG) on their surface can be injected into the blood, cross the blood brain barrier, and enter neurons and glia. RVG-exosomes can carry siRNA cargo. Delivered into the blood circulation, the exosomes deposit siRNA in neurons to engage in RNA interference. Our purpose is to develop exosomes as a therapeutic in HD. The UH2 examines the ability of RVG-exosomes carrying siRNA against huntingtin mRNA to cross the blood brain barrier to enter neurons. Localization in brain and RNAi dependent knock down will be studied. Hyper-functional siRNAs will be sought. Because exosomes are made from cytoplasm of cells, exosome mRNA, miRNA, and implaced siRNA will be identified by deep sequencing. Immune reactivity and immune-neutralization will be studied, since exosomes have potential antigens, like RVG, and will need to be administrated often. The UH3 further establishes exosome-based therapeutics, by study of reversal or prevention of neuropathology and aberrant movement in HD mouse models. Dosing of exosomes will be secured. A team of experts in HD pathogenesis, siRNA development, RNA identification and measurement, RNAi mechanisms and exosome production and brain delivery will carry out the studies. Harnessing exosomes for brain delivery is expected to form a viable therapeutic to reduce expression of mutant huntingtin in patients with HD. Patients with other genetically- based neurodegeneration will benefit.