Optimized Cholesterol-siRNA Chemistry Improves Productive Loading onto Extracellular Vesicles
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Authors
Haraszti, Reka AMiller, Rachael
Didiot, Marie C.
Biscans, Annabelle
Alterman, Julia F.
Hassler, Matthew R.
Roux, Loic
Echeverria, Dimas
Sapp, Ellen
DiFiglia, Marian
Aronin, Neil
Khvorova, Anastasia
UMass Chan Affiliations
Department of MedicineProgram in Molecular Medicine
RNA Therapeutics Institute
Document Type
Journal ArticlePublication Date
2018-03-01Keywords
RNA therapychemical modification
exosomes
extracellular vesicles
nanovesicles
oligonucleotides
siRNA
UMCCTS funding
Biochemistry
Cell Biology
Genetics and Genomics
Medicinal-Pharmaceutical Chemistry
Molecular Biology
Nucleic Acids, Nucleotides, and Nucleosides
Therapeutics
Translational Medical Research
Metadata
Show full item recordAbstract
Extracellular vesicles are promising delivery vesicles for therapeutic RNAs. Small interfering RNA (siRNA) conjugation to cholesterol enables efficient and reproducible loading of extracellular vesicles with the therapeutic cargo. siRNAs are typically chemically modified to fit an application. However, siRNA chemical modification pattern has not been specifically optimized for extracellular vesicle-mediated delivery. Here we used cholesterol-conjugated, hydrophobically modified asymmetric siRNAs (hsiRNAs) to evaluate the effect of backbone, 5'-phosphate, and linker chemical modifications on productive hsiRNA loading onto extracellular vesicles. hsiRNAs with a combination of 5'-(E)-vinylphosphonate and alternating 2'-fluoro and 2'-O-methyl backbone modifications outperformed previously used partially modified siRNAs in extracellular vesicle-mediated Huntingtin silencing in neurons. Between two commercially available linkers (triethyl glycol [TEG] and 2-aminobutyl-1-3-propanediol [C7]) widely used to attach cholesterol to siRNAs, TEG is preferred compared to C7 for productive exosomal loading. Destabilization of the linker completely abolished silencing activity of loaded extracellular vesicles. The loading of cholesterol-conjugated siRNAs was saturated at approximately 3,000 siRNA copies per extracellular vesicle. Overloading impaired the silencing activity of extracellular vesicles. The data reported here provide an optimization scheme for the successful use of hydrophobic modification as a strategy for productive loading of RNA cargo onto extracellular vesicles.Source
Mol Ther. 2018 Aug 1;26(8):1973-1982. doi: 10.1016/j.ymthe.2018.05.024. Epub 2018 Jun 21. Link to article on publisher's site
DOI
10.1016/j.ymthe.2018.05.024Permanent Link to this Item
http://hdl.handle.net/20.500.14038/50341PubMed ID
29937418Related Resources
ae974a485f413a2113503eed53cd6c53
10.1016/j.ymthe.2018.05.024