2022 Fiscal Year Final Research Report
Generation of a persistent therapeutic transgene expression system by engineered latency associated transcript expression cassette of herpes simplex virus
| Project/Area Number |
19K22505
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| Research Category |
Grant-in-Aid for Challenging Research (Exploratory)
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| Allocation Type | Multi-year Fund |
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| Review Section |
Medium-sized Section 47:Pharmaceutical sciences and related fields
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| Research Institution | Nippon Medical School |
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Principal Investigator |
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| Project Period (FY) |
2019-06-28 – 2023-03-31
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| Keywords | ヘルペスウイルス / CRISPRa / インシュレーター |
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| Outline of Final Research Achievements |
In this study, we generated a persistent therapeutic gene expression system by modifying the latency associated transcript (LAT) locus in a non-toxic herpes virus (HSV) vector that we have recently developed. In this novel expression system, a complex of artificial transcriptional activator dCas9 and single guide RNA (sgRNA) to target the LAT locus were continuously recruited to the LAT locus of non-toxic HSV, which allowed the region to be kept in a transcriptionally active state persistent. Moreover, we found that the transcription in the LAT locus can be further activated by using multiple sgRNAs. Our study showed that the engineered expression system generated in this study can be useful for persistent and stable therapeutic transgene expression from non-toxic HSV vectors.
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| Free Research Field |
遺伝子治療学
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| Academic Significance and Societal Importance of the Research Achievements |
従来、sgRNAの発現にはPol IIIプロモーターが用いられ、転写調節因子dCas9とは別々に発現される。この場合、これらの発現系はそれぞれ転写機構が異なるため、人工的に転写調節を同時に行うことは困難である。一方、今回提案する発現系では、miRNA発現系を巧みに利用してPol IIプロモーターによりsgRNAを効率的に発現できる。以上により、本発現系を構成する全ての因子を一括して発現制御することを可能とし、自己プロモーターの恒常的な転写活性化を実現できる。本発現系を用いた遺伝子治療が完成すれば、恒久的な治療効果が期待され、ベクター使用量及びベクター由来免疫応答を最小限に抑えることができる。
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