| Autor: |
Youdiil Ophinni, Sayaka Miki, Yoshitake Hayashi, Masanori Kameoka |
| Jazyk: |
angličtina |
| Rok vydání: |
2020 |
| Předmět: |
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| Zdroj: |
Viruses, Vol 12, Iss 11, p 1223 (2020) |
| Druh dokumentu: |
article |
| ISSN: |
1999-4915 |
| DOI: |
10.3390/v12111223 |
| Popis: |
HIV-1 cure strategy by means of proviral knock-out using CRISPR-Cas9 has been hampered by the emergence of viral resistance against the targeting guide RNA (gRNA). Here, we proposed multiple, concentrated gRNA attacks against HIV-1 regulatory genes to block viral escape. The T cell line were transduced with single and multiple gRNAs targeting HIV-1 tat and rev using lentiviral-based CRISPR-Cas9, followed by replicative HIV-1NL4-3 challenge in vitro. Viral p24 rebound was observed for almost all gRNAs, but multiplexing three tat-targeting gRNAs maintained p24 suppression and cell viability, indicating the inhibition of viral escape. Multiplexed tat gRNAs inhibited acute viral replication in the 2nd round of infection, abolished cell-associated transmission to unprotected T cells, and maintained protection through 45 days, post-infection (dpi) after a higher dose of HIV-1 infection. Finally, we describe here for the first time the assembly of all-in-one lentiviral vectors containing three and six gRNAs targeting tat and rev. A single-vector tat-targeting construct shows non-inferiority to the tat-targeting multi-vector in low-dose HIV-1 infection. We conclude that Cas9-induced, DNA repair-mediated mutations in tat are sufficiently deleterious and deplete HIV-1 fitness, and multiplexed disruption of tat further limits the possibility of an escape mutant arising, thus elevating the potential of CRISPR-Cas9 to achieve a long-term HIV-1 cure. |
| Databáze: |
Directory of Open Access Journals |
| Externí odkaz: |
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