Study of genome editing therapy for Sly disease based on different genome editing mechanisms

2022 Fiscal Year Final Research Report

• Project/Area Number: 20K08195
• Research Category: Grant-in-Aid for Scientific Research (C)
• Allocation Type: Multi-year Fund
• Section: 一般
• Review Section: Basic Section 52050:Embryonic medicine and pediatrics-related
• Research Institution: Jikei University School of Medicine
• Principal Investigator: Kanegae Yumi 東京慈恵会医科大学, 医学部, 教授 (80251453)
• Project Period (FY): 2020-04-01 – 2023-03-31
• Keywords: ゲノム編集 / アデノウイルスベクター / Sly病

Outline of Final Research Achievements

Genome editing is expected to be the ultimate gene therapy but remaining problems that sufficient therapeutic effects cannot be obtained due to low genome editing efficiency. Therefore, the HITI method and the PITCh method have been reported.
At first, I succeeded in constructing a "cell-specific high-efficient and short-term Cas9 expression system". It was also confirmed that editing by the HR method can be performed efficiently in the model mouse GUSB gene. Considering future gene therapy, I also developed a system that safely inserts the gene expression unit into the AAVS1 region. In the study using HuH-7 cells, it was shown that the HR method was able to insert the target expression unit 2 times more efficiently than the PITCh method and 17 times more efficiently than the HITI method. In the future, I plan to compare the insertion-efficiency with these methods using induces liver cells from iPS cells. I believe that this research will be useful for genome editing therapy.

Free Research Field

分子ウイルス学

Academic Significance and Societal Importance of the Research Achievements

本研究では、従来のゲノム編集治療法で問題であった修復効率を上昇するためのHITI法やPITCh法によるゲノム修復が増殖期の細胞ではHR法よりは劣っていたものの可能であることを明らかにした。今後はiPS細胞から誘導する肝臓細胞を用いて体細胞の殆どをしめる静止期細胞での修復効率について検討を進めていく。また、レンチウイルスベクターには劣るものの、AdVを用いてAAVS1などのセーフ・ハーバー領域にCAGプロモーターなど強力なプロモーターを用いて治療用遺伝子を高効率に導入可能であったことから、今後のex vivo遺伝子治療の安全性の向上に寄与できる結果が得られたと考える。