2018 Fiscal Year Research-status Report
DOCK11ノックダウンによりcccDNA排除の新規抗HBV治療応用への基礎研究
| Project/Area Number |
18K07966
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| Research Institution | Kanazawa University |
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Principal Investigator |
李 影奕 金沢大学, 医学系, 博士研究員 (70401940)
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| Co-Investigator(Kenkyū-buntansha) |
本多 政夫 金沢大学, 保健学系, 教授 (00272980)
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| Project Period (FY) |
2018-04-01 – 2021-03-31
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| Keywords | Hepatitis B virus / infection / cccDNA / Dock11 |
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| Outline of Annual Research Achievements |
Hepatitis B virus (HBV) infection remains a major health problem worldwide. One major hurdle to treat HBV infection is the persistence of viral covalently closed circular DNA (cccDNA) that is not directly targeted by current anti-HBV therapeutics. Thus, it is imperative to understand the molecular mechanism that control cccDNA biogenesis, homeostasis and decay.
Until now, we identified DOCK11, a cccDNA maintenance gene, using our established cell culture model. The purpose of this project is to understand the roles of DOCK11 in cccDNA maintenance and subsequently clarify the possibility of DOCK11 as a target molecule for treating chronic HBV infection.
In this year, we have performed in vitro experiment and realized that HBV DNA could entry to the nucleus by Dock11 regulating X gene.
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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
The project is going smoothly according to the experimental plan. The biological effects and mechanism of DOCK11 regarding HBV DNA maintenance were investigated and HBV Rc-DNA entering to the nucleus by Dock11 regulating X gene were clarified in vitro.
1) The overexpression and knockdown DOCK11 plasmids were constructed in lentiviral vector. 2) Stably DOCK11-expressing HepG2-NTCP-C4 cells were established in Dox-inducing system. 3) Stably DOCK11-knocked down HepG2-NTCP-C4 cells and Huh7 cells were established using CRISPR/Cas9 system. 4) The level of HBV DNA and cccDNA were increased or decreased by overexpression of Dock11 or silencing Dock11 by real-time PCR, Southern blotting, and Western blotting. 5) We demonstrated that HBV Rc-DNA entry to the nucleus by Dock11 regulating X gene.
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| Strategy for Future Research Activity |
We will perform the experiments according to the research plan.
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| Causes of Carryover |
There was a some difference in delivery prices for the experimental consumables.
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[Presentation] mTORC2-Akt phosphorylation is associsted with Notch signaling activation by nucleotide analogues but not nucleoside analogues for chronic HBV infection2018
Author(s)
Kazunori Kawaguchi, Zijing Wang, Masao Honda, takayoshi shirasaki, Hikari Okada, Ying-Yi Li, Kazuhisa Murai, Kouki Nio, Tetsuro Shimakami, Taro Yamashita, Shinichi Hashimoto, Kazuyuki kuroki, Seishi Murakami, Shuichi Kaneko
Organizer
2018 international HBV meeting
Int'l Joint Research
