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The efficacy of CRISPR-Cas9-mediated induction of the CCR5delta32 mutation in the human embryo
Abstract
The editing of the CCR5 gene in the CD4+ T cell genome is an effective way of preventing HIV-1 proliferation. Very similar strategies can be used to protect the fetus of an HIV-infected female showing a weak response to antiretroviral therapy. Inducing the “natural” CCR5delta32 mutation in a zygote may guard the fetus against HIV infection both in utero and at birth. In this study, we optimize the CRISPR-Cas9 system to induce a homozygous 32-nt deletion similar to the naturally occurring CCR5delta32 allele in the human zygote at the S-phase. Edits were done in the abnormal tripronuclear zygotes unsuitable for IVF. Sixteen tripronuclear zygotes in the S-phase obtained from WT CCR5 donors were injected with an original CRISPR-Cas9 system designed by the authors. Upon injection, the zygotes were transferred into the Blastocyst (COOK) embryo culture medium and cultured for 5 days in a CO2 incubator until blastocysts were formed (approximately 250 cells). Eight zygotes that successfully developed into blastocysts were PCR-genotyped to analyze the efficacy of genome editing. Of 16 zygotes injected with CRISPR-Cas9, only 8 reached the blastocyst stage. PCR genotyping revealed the absence of the initial WT CCR5 variant in 5 of 8 blastocysts (100% CCR5delta32 homozygous). Two had about 3% and one about 20% of WT CCR5 mosaicism. This leads us to conclude that the efficacy of the proposed CRISPR-Cas9 system for the induction of the CCR5delta32 mutation in human embryos is very high producing more than 50% of completely modified embryos.
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“As in the experiments of He, he was planning to edit the CCR5 gene, and the preliminary work from his laboratory on non-viable embryos was published in the Bulletin of the Russian National Research Medical University [59].”
From CRISPR–Cas9: A History of Its Discovery and Ethical Considerations of Its Use in Genome Editing (2022) · cites this paper as [59]
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Genome editing reveals a role for OCT4 in human embryogenesis
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Research works printed but not linked: 14
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Cited by
Sorted by citations
CRISPR–Cas9: A History of Its Discovery and Ethical Considerations of Its Use in Genome Editing
Gostimskaya, I. (2022) · Biochemistry (Moscow) · doi:10.1134/s0006297922080090 · Cited by 224
Metadata
- Authors
- Kodyleva, T. A., Kirillova, A. O., Tyschik, E. A., Makarov, V. V., Khromov, A. V., Guschin, V. A., Abubakirov, A. N., Rebrikov, D. V., & Sukhikh, G. T.
- Affiliations
- Kulakov National Medical Research Center for Obstetrics, Gynecology and Perinatology, Moscow
- Lomonosov Moscow State University, Moscow
- Kulakov National Medical Research Center for Obstetrics, Gynecology and Perinatology, Moscow; Pirogov Russian National Research Medical University, Moscow
- Venue
- Bulletin of Russian State Medical University
- Published
- 2018-10-16
- Issue
- (4)2018
- Pages
- 70-74
- ISSN
- 2500-1094
- 2542-1204
- Cited by
- 3
- References
- 1
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- Kodyleva, T. A., Kirillova, A. O., Tyschik, E. A., Makarov, V. V., Khromov, A. V., Guschin, V. A., Abubakirov, A. N., Rebrikov, D. V., & Sukhikh, G. T. (2018). The efficacy of CRISPR-Cas9-mediated induction of the CCR5delta32 mutation in the human embryo. Bulletin of Russian State Medical University. https://doi.org/10.24075/brsmu.2018.052
