GAPDH controls extracellular vesicle biogenesis and enhances therapeutic potential of EVs in silencing the Huntingtin gene in mice via siRNA delivery
GAPDH controls extracellular vesicle biogenesis and enhances therapeutic potential of EVs in silencing the Huntingtin gene in mice via siRNA delivery
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GAPDH 控制细胞外囊泡的生物合成，并通过 siRNA 传递增强 EV 沉默小鼠亨廷顿蛋白基因的治疗潜力
DOI：
10.1101/2020.01.09.899880
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发表时间：
2020
期刊：
影响因子：
--
通讯作者：
Dar G
中科院分区：
文献类型：
--
作者：
Dar G
细胞外囊泡（EVs）是一种生物纳米粒子，在细胞间通讯和病理生理中具有重要作用。它们在细胞之间转移生物分子的能力引发了对电动汽车进行生物工程的努力，使其成为药物输送载体。然而，需要更好地了解EV的生物发生机制和功能，以释放其巨大的治疗潜力。在这里，我们展示了GAPDH（一种糖酵解酶）在EV组装和分泌中的新作用，并利用这些发现开发了一种基于GAPDH的方法，将治疗性sirna装载到EV上，以靶向药物递送到大脑。在一系列实验中，我们观察到高水平的GAPDH结合到evsvia磷脂酰丝氨酸结合基序G58的外表面，并发现GAPDH的四聚体性质促进了广泛的EV聚集。在aDrosophilaEV生物发生模型中的研究表明，GAPDH对于内体腔室腔内囊泡的正常产生是绝对必需的，并促进细胞内外的囊泡聚集。将gapdh衍生的G58肽与dsrna结合基序融合，可以高效地将基于rna的药物（如siRNA）装载到ev表面。这种囊泡在全身注射后，在体外有效地将siRNA传递到靶细胞，并进入亨廷顿病小鼠模型的大脑，导致大脑多个解剖区域的亨廷顿蛋白基因沉默，并调节疾病的表型特征。综上所述，我们的研究证明了GAPDH在EV生物发生中的新作用，并且可以有效地利用EV上游离GAPDH结合位点的存在，从而大大增强EV介导的药物输送到大脑的治疗潜力。
Extracellular vesicles (EVs) are biological nanoparticles with important roles in intercellular communication and pathophysiology. Their capacity to transfer biomolecules between cells has sparked efforts to bioengineer EVs as drug delivery vehicles. However, a better understanding of EV biogenesis mechanisms and function is required to unleash their considerable therapeutic potential. Here we demonstrate a novel role for GAPDH, a glycolytic enzyme, in EV assembly and secretion, and we exploit these findings to develop a GAPDH-based methodology to load therapeutic siRNAs onto EVs for targeted drug delivery to the brain. In a series of experiments, we observe high levels of GAPDH binding to the outer surface of EVsviaa phosphatidylserine binding motif, designated as G58, and discover that the tetrameric nature of GAPDH promotes extensive EV aggregation. Studies in aDrosophilaEV biogenesis model demonstrate that GAPDH is absolutely required for normal generation of intraluminal vesicles in endosomal compartments and promotes vesicle clustering both inside and outside the cell. Fusing a GAPDH-derived G58 peptide to dsRNA-binding motifs permits highly efficient loading of RNA-based drugs such as siRNA onto the surface of EVs. Such vesicles efficiently deliver siRNA to target cellsin vitroand into the brain of a Huntington’s disease mouse model after systemic injection, resulting in silencing of the huntingtin gene in multiple anatomical regions of the brain and modulation of phenotypic features of disease. Taken together, our study demonstrates a novel role for GAPDH in EV biogenesis, and that the presence of free GAPDH binding sites on EVs can be effectively exploited to substantially enhance the therapeutic potential of EV-mediated drug delivery to the brain.
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影响因子：
12.4
作者：
Godinho, Bruno M. D. C.;Henninger, Nils;Khvorova, Anastasia
通讯作者：
Khvorova, Anastasia
负责膜融合的甘油醛-3-磷酸脱氢酶的磷脂酰丝氨酸结合位点的定位。
影响因子：
2.7
作者：
M. Kaneda;K. Takeuchi;K. Inoue;M. Umeda
通讯作者：
M. Umeda
影响因子：
12.4
作者：
Nakase, Ikuhiko;Hirose, Hisaaki;Futaki, Shiroh
通讯作者：
Futaki, Shiroh
影响因子：
--
作者：
R. Parton;A. Vallés;I. Dobbie;Ilan Davis
通讯作者：
Ilan Davis
影响因子：
4.8
作者：
Nakagawa, T;Hirano, Y;Horigome, T
通讯作者：
Horigome, T