2021 Fiscal Year Research-status Report
Ligand-free hepatocyte-targeting of nanomedicines by selective stealth coating of liver reticuloendothelial system scavenger cells
| Project/Area Number |
21K18062
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| Research Institution | Kawasaki Institute of Industrial Promotion Innovation Center of NanoMedicine |
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Principal Investigator |
ディリサラ アンジャネユル 公益財団法人川崎市産業振興財団(ナノ医療イノベーションセンター), ナノ医療イノベーションセンター, 研究員 (70794353)
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| Project Period (FY) |
2021-04-01 – 2023-03-31
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| Keywords | Liver stealth coating / Hepatocyte targeting / Gene delivery |
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| Outline of Annual Research Achievements |
The luciferase-encoding mRNA encapsulated cationic (lipid to mRNA ratio 5:1, charge +52 mV, size 155 nm) and anionic (lipid to mRNA ratio 1:2, charge -30 mV, size 221 nm) mRNA-lipoplexes were prepared by different coating amounts of the mRNA onto liposomes composed of DOTMA and DOPE. Prior stealth coating of the liver sinusoidal RES wall with two-arm-PEG-Oligo(L-Lysine) substantially enhanced the luciferase expression (14.1-fold increase) of anionic liposome in the liver compared to without liver coating. Similarly, PEG coating to the liver scavenger cell wall augmented the luciferase expression in the liver up to 3.6-fold compared to without coating. Based on these results, the applicant will choose anionic mRNA-lipoplex for future liver-directed therapeutic applications.
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| Current Status of Research Progress |
Current Status of Research Progress
2: Research has progressed on the whole more than it was originally planned.
Reason
We have optimized the best-in-class mRNA-lipoplex carrier for mRNA introduction into the liver using the mRNA-encoding reporter protein, luciferase, using a bioluminescence assay.
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| Strategy for Future Research Activity |
Rather than developing new carriers for mRNA delivery, the applicant will focus on relocating the existed mRNA carriers from the liver scavenger cells, liver sinusoidal endothelial cells, and Kupffer cells to the parenchymal hepatocytes without integrating complicated structural functionalities such as introducing surface ligands, charge and size modulation. In the future, such hepatocyte-selective protein expression will be exploited to develop a cure for chronic hepatitis B by destroying the virus's persistent covalently closed circular (ccc)DNA using a gene-editing tool, clustered regularly interspersed short palindromic repeats (CRISPR)/CRISPR-associated protein 9.
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| Causes of Carryover |
The applicant will purchase fluorescent dyes for labeling the mRNA-encapsulated liposome and two-arm-PEG-Oligo(L-Lysine) for tracking their distribution in the body using real-time in vivo confocal microscopy. The applicant will also purchase the experimental mice for evaluating organ- and cell-specific delivery of mRNA-liposomes with and without liver stealth coating using flow cytometry and histological examination.
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