2019 Fiscal Year Annual Research Report
Fine-tuning of polymer amphiphile for high performance of messenger RNA delivery
| Project/Area Number |
18K18378
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| Research Institution | The University of Tokyo |
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Principal Investigator |
Kim HyunJin 東京大学, 大学院医学系研究科(医学部), 特任助教 (10755002)
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| Project Period (FY) |
2018-04-01 – 2020-03-31
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| Keywords | cationic polymer / mRNA transfection / amphiphile / CRISPR-Cas technology |
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| Outline of Annual Research Achievements |
This Wakate project synthesized new amphiphilic cationic polymers for high efficient local delivery of in vitro transcribed mRNA (IVT mRNA) and applied to gene editing via CRISPR-Cas9 technology. The best polymer was selected from 7 different amphiphilic polymers, which were prepared by ring-opening polymerization and subsequent aminolysis. A series of polymers formed 100-150 nm sized nanoparticles with firefly or gaussian luciferase IVT mRNA. These nanoparticles were transfected into cultured mouse myoblast cells and neuronal cancer cells. The best polymer exhibited 5 orders of magnitude higher than the other polymer with the least efficacy. We found that there was the optimal hydrophobicity of the polymers between nanoparticle stability and mRNA release property.
For gene editing, we used genetically engineered mouse model (Ai9 mouse). When gene editing occurs, the cells in the Ai9 mouse emit red fluorescence. The best polymer formed nanoparticles with Cas9 mRNA and single guide RNA, respectively. The nanoparticles were locally injected into brain cerebroventricular area, nose, and skeletal muscle of Ai9 mouse. Gene editing in each tissue was successfully observed by using confocal microscope. These data indicate that the precise hydrophobicity control in the cationic polymer considerably enhances performance of IVT mRNA delivery efficacy. The new polymer has a high potential for gene editing therapy via Cas9 mRNA delivery, which may overcome disadvantages of Adeno-associated virus (AAV)-mediated CRISPR-Cas9 technology.
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