With the advent of efficient, easy-to-use genome editing by CRISPR–Cas9, editing human embryos is now possible, providing tremendous opportunities to study gene function and cell fate in early human development. The technique can also be used to modify the human germline. Unresolved questions about pre-implantation human development could be addressed by basic research using CRISPR–Cas9. In this Perspective, we discuss advances in human genome editing and consider ethical questions and potential clinical implications of this technology.
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Cong, L. et al. Multiplex genome engineering using CRISPR/Cas systems. Science 339, 819–823 (2013).
Jinek, M. et al. RNA-programmed genome editing in human cells. eLife 2, e00471 (2013).
Mali, P. et al. RNA-guided human genome engineering via Cas9. Science 339, 823–826 (2013).
Adli, M. The CRISPR tool kit for genome editing and beyond. Nat. Commun. 9, 1911 (2018).
Carroll, D. Genome engineering with zinc-finger nucleases. Genetics 188, 773–782 (2011).
Joung, J. K. & Sander, J. D. TALENs: a widely applicable technology for targeted genome editing. Nat. Rev. Mol. Cell Biol. 14, 49–55 (2013).
Silva, G. et al. Meganucleases and other tools for targeted genome engineering: perspectives and challenges for gene therapy. Curr. Gene Ther. 11, 11–27 (2011).
Gasiunas, G., Barrangou, R., Horvath, P. & Siksnys, V. Cas9-crRNA ribonucleoprotein complex mediates specific DNA cleavage for adaptive immunity in bacteria. Proc. Natl Acad. Sci. USA 109, E2579–E2586 (2012).
Jinek, M. et al. A programmable dual-RNA-guided DNA endonuclease in adaptive bacterial immunity. Science 337, 816–821 (2012).
Liang, P. et al. CRISPR/Cas9-mediated gene editing in human tripronuclear zygotes. Protein Cell 6, 363–372 (2015).
Kang, X. et al. Introducing precise genetic modifications into human 3PN embryos by CRISPR/Cas-mediated genome editing. J. Assist. Reprod. Genet. 33, 581–588 (2016).
Tang, L. et al. CRISPR/Cas9-mediated gene editing in human zygotes using Cas9 protein. Mol. Genet. Genomics 292, 525–533 (2017).
Joergensen, M. W. et al. Altered cleavage patterns in human tripronuclear embryos and their association to fertilization method: a time-lapse study. J. Assist. Reprod. Genet. 31, 435–442 (2014).
Lammers, J., Splingart, C., Barrière, P. & Fréour, T. Morphokinetic parameters of ICSI tripronucleated embryos observed using time lapse. Reprod. Biomed. Online 28, 658–660 (2014).
Yao, G. et al. Developmental potential of clinically discarded human embryos and associated chromosomal analysis. Sci. Rep. 6, 23995 (2016).
Ma, H. et al. Correction of a pathogenic gene mutation in human embryos. Nature 548, 413–419 (2017).
Safari, F., Farajnia, S., Ghasemi, Y. & Zarghami, N. New developments in CRISPR technology: improvements in specificity and efficiency. Curr. Pharm. Biotechnol. 18, 1038–1054 (2017).
Fogarty, N. M. E. et al. Genome editing reveals a role for OCT4 in human embryogenesis. Nature 550, 67–73 (2017).
Frum, T. et al. Oct4 cell-autonomously promotes primitive endoderm development in the mouse blastocyst. Dev. Cell 25, 610–622 (2013).
Nichols, J. et al. Formation of pluripotent stem cells in the mammalian embryo depends on the POU transcription factor Oct4. Cell 95, 379–391 (1998).
Daigneault, B. W., Rajput, S., Smith, G. W. & Ross, P. J. Embryonic POU5F1 is required for expanded bovine blastocyst formation. Sci. Rep. 8, 7753 (2018).
Davis, M. I. et al. Comprehensive analysis of kinase inhibitor selectivity. Nat. Biotechnol. 29, 1046–1051 (2011).
De Souza, A. T. et al. Transcriptional and phenotypic comparisons of Ppara knockout and siRNA knockdown mice. Nucleic Acids Res. 34, 4486–4494 (2006).
Blij, S., Frum, T., Akyol, A., Fearon, E. & Ralston, A. Maternal Cdx2 is dispensable for mouse development. Development 139, 3969–3972 (2012).
Kok, F. O. et al. Reverse genetic screening reveals poor correlation between morpholino-induced and mutant phenotypes in zebrafish. Dev. Cell 32, 97–108 (2015).
Jackson, A. L. et al. Expression profiling reveals off-target gene regulation by RNAi. Nat. Biotechnol. 21, 635–637 (2003).
Jackson, A. L. & Linsley, P. S. Noise amidst the silence: off-target effects of siRNAs? Trends Genet. 20, 521–524 (2004).
Scacheri, P. C. et al. Short interfering RNAs can induce unexpected and divergent changes in the levels of untargeted proteins in mammalian cells. Proc. Natl Acad. Sci. USA 101, 1892–1897 (2004).
Robu, M. E. et al. p53 activation by knockdown technologies. PLoS Genet. 3, e78 (2007).
Rossi, A. et al. Genetic compensation induced by deleterious mutations but not gene knockdowns. Nature 524, 230–233 (2015).
Clift, D., So, C., McEwan, W. A., James, L. C. & Schuh, M. Acute and rapid degradation of endogenous proteins by Trim-Away. Nat. Protoc. 13, 2149–2175 (2018).
Hardy, K., Handyside, A. H. & Winston, R. M. The human blastocyst: cell number, death and allocation during late preimplantation development in vitro. Development 107, 597–604 (1989).
Boomsma, C. M. et al. Endometrial secretion analysis identifies a cytokine profile predictive of pregnancy in IVF. Hum. Reprod. 24, 1427–1435 (2009).
Braude, P., Bolton, V. & Moore, S. Human gene expression first occurs between the four- and eight-cell stages of preimplantation development. Nature 332, 459–461 (1988).
Vassena, R. et al. Waves of early transcriptional activation and pluripotency program initiation during human preimplantation development. Development 138, 3699–3709 (2011).
Artley, J. K., Braude, P. R. & Johnson, M. H. Gene activity and cleavage arrest in human pre-embryos. Hum. Reprod. 7, 1014–1021 (1992).
Jarvis, G. E. Early embryo mortality in natural human reproduction: What the data say. F1000Res. 5, 2765 (2016).
Koot, Y. E., Teklenburg, G., Salker, M. S., Brosens, J. J. & Macklon, N. S. Molecular aspects of implantation failure. Biochim. Biophys. Acta 1822, 1943–1950 (2012).
Hertig, A. T., Rock, J., Adams, E. C. & Menkin, M. C. Thirty-four fertilized human ova, good, bad and indifferent, recovered from 210 women of known fertility; a study of biologic wastage in early human pregnancy. Pediatrics 23, 202–211 (1959).
Lee, A. & Kiessling, A. A. Early human embryos are naturally aneuploid-can that be corrected? J. Assist. Reprod. Genet. 34, 15–21 (2017).
Scott, R. T. Jr. et al. Blastocyst biopsy with comprehensive chromosome screening and fresh embryo transfer significantly increases in vitro fertilization implantation and delivery rates: a randomized controlled trial. Fertil. Steril. 100, 697–703 (2013).
Mantikou, E., Wong, K. M., Repping, S. & Mastenbroek, S. Molecular origin of mitotic aneuploidies in preimplantation embryos. Biochim. Biophys. Acta 1822, 1921–1930 (2012).
Delhanty, J. D. & Handyside, A. H. The origin of genetic defects in the human and their detection in the preimplantation embryo. Hum. Reprod. Update 1, 201–215 (1995).
Maurer, M. et al. Chromosomal aneuploidies and early embryonic developmental arrest. Int. J. Fertil. Steril. 9, 346–353 (2015).
Vanneste, E. et al. Chromosome instability is common in human cleavage-stage embryos. Nat. Med. 15, 577–583 (2009).
Hassold, T. et al. A cytogenetic study of 1000 spontaneous abortions. Ann. Hum. Genet. 44, 151–178 (1980).
Bazrgar, M. et al. DNA repair signalling pathway genes are overexpressed in poor-quality pre-implantation human embryos with complex aneuploidy. Eur. J. Obstet. Gynecol. Reprod. Biol. 175, 152–156 (2014).
Kiessling, A. A. et al. Evidence that human blastomere cleavage is under unique cell cycle control. J. Assist. Reprod. Genet. 26, 187–195 (2009).
Kiessling, A. A. et al. Genome-wide microarray evidence that 8-cell human blastomeres over-express cell cycle drivers and under-express checkpoints. J. Assist. Reprod. Genet. 27, 265–276 (2010).
Martinez, F. et al. Caspase activity in preimplantation human embryos is not associated with apoptosis. Hum. Reprod. 17, 1584–1590 (2002).
Kermi, C., Aze, A. & Maiorano, D. Preserving genome integrity during the early embryonic DNA replication cycles. Genes (Basel) 10, (E398 (2019).
Hardy, K. Apoptosis in the human embryo. Rev. Reprod. 4, 125–134 (1999).
Cubbon, A., Ivancic-Bace, I. & Bolt, E. L. CRISPR-Cas immunity, DNA repair and genome stability. Biosci. Rep. 38, BSR20180457 (2018).
Canny, M. D. et al. Inhibition of 53BP1 favors homology-dependent DNA repair and increases CRISPR-Cas9 genome-editing efficiency. Nat. Biotechnol. 36, 95–102 (2018).
Takayama, K. et al. Highly efficient biallelic genome editing of human ES/iPS cells using a CRISPR/Cas9 or TALEN system. Nucleic Acids Res. 45, 5198–5207 (2017).
Wilde, J.J. et al. Efficient zygotic genome editing via RAD51-enhanced interhomolog repair. Preprint at bioRxiv https://doi.org/10.1101/263699 (2018).
Balakier, H., MacLusky, N. J. & Casper, R. F. Characterization of the first cell cycle in human zygotes: implications for cryopreservation. Fertil. Steril. 59, 359–365 (1993).
Capmany, G., Taylor, A., Braude, P. R. & Bolton, V. N. The timing of pronuclear formation, DNA synthesis and cleavage in the human 1-cell embryo. Mol. Hum. Reprod. 2, 299–306 (1996).
Kim, S., Kim, D., Cho, S. W., Kim, J. & Kim, J. S. Highly efficient RNA-guided genome editing in human cells via delivery of purified Cas9 ribonucleoproteins. Genome Res. 24, 1012–1019 (2014).
Horlbeck, M. A. et al. Nucleosomes impede Cas9 access to DNA in vivo and in vitro. eLife 5, e12677 (2016).
Isaac, R. S. et al. Nucleosome breathing and remodeling constrain CRISPR-Cas9 function. eLife 5, e13450 (2016).
Gutschner, T., Haemmerle, M., Genovese, G., Draetta, G. F. & Chin, L. Post-translational regulation of Cas9 during G1 enhances homology-directed repair. Cell Rep. 14, 1555–1566 (2016).
Gu, B., Posfai, E. & Rossant, J. Efficient generation of targeted large insertions by microinjection into two-cell-stage mouse embryos. Nat. Biotechnol. 36, 632–637 (2018).
Sakaue-Sawano, A. et al. Visualizing spatiotemporal dynamics of multicellular cell-cycle progression. Cell 132, 487–498 (2008).
Sakaue-Sawano, A., Kobayashi, T., Ohtawa, K. & Miyawaki, A. Drug-induced cell cycle modulation leading to cell-cycle arrest, nuclear mis-segregation, or endoreplication. BMC Cell Biol. 12, 2 (2011).
Zhou, Y. et al. Painting a specific chromosome with CRISPR/Cas9 for live-cell imaging. Cell Res. 27, 298–301 (2017).
Reichmann, J., Eguren, M., Lin, Y., Schneider, I. & Ellenberg, J. Live imaging of cell division in preimplantation mouse embryos using inverted light-sheet microscopy. Methods Cell Biol. 145, 279–292 (2018).
Abe, T. et al. Visualization of cell cycle in mouse embryos with Fucci2 reporter directed by Rosa26 promoter. Development 140, 237–246 (2013).
Suzuki, T., Asami, M. & Perry, A. C. Asymmetric parental genome engineering by Cas9 during mouse meiotic exit. Sci. Rep. 4, 7621 (2014).
Egli, D. et al. Inter-homologue repair in fertilized human eggs? Nature 560, E5–E7 (2018).
Adikusuma, F. et al. Large deletions induced by Cas9 cleavage. Nature 560, E8–E9 (2018).
Reichmann, J. et al. Dual-spindle formation in zygotes keeps parental genomes apart in early mammalian embryos. Science 361, 189–193 (2018).
Ma, H. et al. Ma et al. reply. Nature 560, E10–E23 (2018).
Kosicki, M., Tomberg, K. & Bradley, A. Repair of double-strand breaks induced by CRISPR-Cas9 leads to large deletions and complex rearrangements. Nat. Biotechnol. 36, 765–771 (2018).
Komor, A. C., Kim, Y. B., Packer, M. S., Zuris, J. A. & Liu, D. R. Programmable editing of a target base in genomic DNA without double-stranded DNA cleavage. Nature 533, 420–424 (2016).
Rees, H. A. & Liu, D. R. Base editing: precision chemistry on the genome and transcriptome of living cells. Nat. Rev. Genet. 19, 770–788 (2018).
Li, G. et al. Highly efficient and precise base editing in discarded human tripronuclear embryos. Protein Cell 8, 776–779 (2017).
Liang, P. et al. Correction of β-thalassemia mutant by base editor in human embryos. Protein Cell 8, 811–822 (2017).
Zhou, C. et al. Highly efficient base editing in human tripronuclear zygotes. Protein Cell 8, 772–775 (2017).
Zeng, Y. et al. Correction of the Marfan syndrome pathogenic FBN1 mutation by base editing in human cells and heterozygous embryos. Mol. Ther. 26, 2631–2637 (2018).
Jin, S. et al. Cytosine, but not adenine, baseeditors induce genome-wide off-target mutations in rice. Science 364, 292–295 (2019).
Zuo, E. et al. Cytosine baseeditor generates substantial off-target single-nucleotide variants in mouse embryos. Science 364, (289–292 (2019).
Yang, G. et al. Base-editing-mediated R17H substitution in histone H3 reveals methylation-dependent regulation of Yap signaling and early mouse embryo development. Cell Rep. 26, 302–312.e4 (2019).
Anzalone, A. V. et al. Search-and-replace genome editing without double-strand breaks or donor DNA. Nature https://doi.org/10.1038/s41586-019-1711-4 (2019).
McArthur, S. J. et al. Blastocyst trophectoderm biopsy and preimplantation genetic diagnosis for familial monogenic disorders and chromosomal translocations. Prenat. Diagn. 28, 434–442 (2008).
Adler, A. et al. Blastocyst culture selects for euploid embryos: comparison of blastomere and trophectoderm biopsies. Reprod. Biomed. Online 28, 485–491 (2014).
Vilarino, M. et al. Mosaicism diminishes the value of pre-implantation embryo biopsies for detecting CRISPR/Cas9 induced mutations in sheep. Transgenic Res. 27, 525–537 (2018).
Thornhill, A. R. & Snow, K. Molecular diagnostics in preimplantation genetic diagnosis. J. Mol. Diagn. 4, 11–29 (2002).
Blais, J. et al. Risk of misdiagnosis due to allele dropout and false-positive PCR artifacts in molecular diagnostics: analysis of 30,769 genotypes. J. Mol. Diagn. 17, 505–514 (2015).
Chakrabarti, A. M. et al. Target-specific precision of CRISPR-mediated genome editing. Mol. Cell 73, 699–713.e6 (2019).
Allen, F. et al. Predicting the mutations generated by repair of Cas9-induced double-strand breaks. Nat. Biotechnol. 37, 64–72 (2018).
Shen, M. W. et al. Predictable and precise template-free CRISPR editing of pathogenic variants. Nature 563, 646–651 (2018).
Kim, D. et al. Digenome-seq: genome-wide profiling of CRISPR-Cas9 off-target effects in human cells. Nat. Methods 12, 237–243 (2015).
Tsai, S. Q. et al. GUIDE-seq enables genome-wide profiling of off-target cleavage by CRISPR-Cas nucleases. Nat. Biotechnol. 33, 187–197 (2015).
Tsai, S. Q. et al. CIRCLE-seq: a highly sensitive in vitro screen for genome-wide CRISPR-Cas9 nuclease off-targets. Nat. Methods 14, 607–614 (2017).
Cameron, P. et al. Mapping the genomic landscape of CRISPR-Cas9 cleavage. Nat. Methods 14, 600–606 (2017).
Yan, W. X. et al. BLISS is a versatile and quantitative method for genome-wide profiling of DNA double-strand breaks. Nat. Commun. 8, 15058 (2017).
Wienert, B. et al. Unbiased detection of CRISPR off-targets in vivo using DISCOVER-Seq. Science 364, 286–289 (2019).
Kim, D. & Kim, J. S. DIG-seq: a genome-wide CRISPR off-target profiling method using chromatin DNA. Genome Res. 28, 1894–1900 (2018).
Smith, C. et al. Whole-genome sequencing analysis reveals high specificity of CRISPR/Cas9 and TALEN-based genome editing in human iPSCs. Cell Stem Cell 15, 12–13 (2014).
Veres, A. et al. Low incidence of off-target mutations in individual CRISPR-Cas9 and TALEN targeted human stem cell clones detected by whole-genome sequencing. Cell Stem Cell 15, 27–30 (2014).
Baltimore, D. et al. Biotechnology. A prudent path forward for genomic engineering and germline gene modification. Science 348, 36–38 (2015).
Lanphier, E., Urnov, F., Haecker, S. E., Werner, M. & Smolenski, J. Don’t edit the human germ line. Nature 519, 410–411 (2015).
Savulescu, J., Pugh, J., Douglas, T. & Gyngell, C. The moral imperative to continue gene editing research on human embryos. Protein Cell 6, 476–479 (2015).
National Academies of Sciences Engineering and Medicine. Human Genome Editing: Science, Ethics, and Governance. (The National Academies Press, Washington, DC, 2017).
Nuffield Council on Bioethics. Genome Editing and Human Reproduction: social and ethical issues. (Nuffield Council on Bioethics, London, 2018).
De Rycke, M. et al. ESHRE PGD Consortium data collection XIV-XV: cycles from January 2011 to December 2012 with pregnancy follow-up to October 2013. Hum. Reprod. 32, 1974–1994 (2017).
Jacobs, H. S. & Agrawal, R. Complications of ovarian stimulation. Baillieres Clin. Obstet. Gynaecol. 12, 565–579 (1998).
Baron, K. T., Babagbemi, K. T., Arleo, E. K., Asrani, A. V. & Troiano, R. N. Emergent complications of assisted reproduction: expecting the unexpected. Radiographics 33, 229–244 (2013).
Steffann, J., Jouannet, P., Bonnefont, J. P., Chneiweiss, H. & Frydman, N. Could failure in preimplantation genetic diagnosis justify editing the human embryo genome? Cell Stem Cell 22, 481–482 (2018).
Natsume, T., Kiyomitsu, T., Saga, Y. & Kanemaki, M. T. Rapid protein depletion in human cells by auxin-inducible degron tagging with short homology donors. Cell Rep. 15, 210–218 (2016).
Sternberg, N. & Hamilton, D. Bacteriophage P1 site-specific recombination. I. Recombination between loxP sites. J. Mol. Biol. 150, 467–486 (1981).
Sauer, B. & Henderson, N. Site-specific DNA recombination in mammalian cells by the Cre recombinase of bacteriophage P1. Proc. Natl Acad. Sci. USA 85, 5166–5170 (1988).
Lander, E. S. et al. Adopt a moratorium on heritable genome editing. Nature 567, 165–168 (2019).
Cyranoski, D. Russian biologist plans more CRISPR-edited babies. Nature 570, 145–146 (2019).
Cyranoski, D. China set to introduce gene-editing regulation following CRISPR-baby furore. Nature https://doi.org/10.1038/d41586-019-01580-1 (2019).
Normile, D. China tightens rules on gene editing. Science 363, 1023 (2019).
Cyranoski, D. Japan set to allow gene editing in human embryos. Nature https://doi.org/10.1038/d41586-018-06847-7 (2018).
Chiba, N. Japanese government considering legally restricting genome editing technology. https://mainichi.jp/english/articles/20190412/p2a/00m/0na/004000c (2019).
Reardon, S. World Health Organization panel weighs in on CRISPR-babies debate. Nature 567, 444–445 (2019).
Dzau, V. J., McNutt, M. & Ramakrishnan, V. Academies’ action plan for germline editing. Nature 567, 175 (2019).
Ledford, H. CRISPR babies: when will the world be ready? Nature 570, 293–296 (2019).
Cree, L. & Loi, P. Mitochondrial replacement: from basic research to assisted reproductive technology portfolio tool-technicalities and possible risks. Mol. Hum. Reprod. 21, 3–10 (2015).
Claiborne, A. B., English, R. A., Kahn, J. P. & ETHICS, O. F. NEW TECHNOLOGIES. Finding an ethical path forward for mitochondrial replacement. Science 351, 668–670 (2016).
Human Fertilisation and Embryology Authority. Scientific Review of the Safety and Efficacy of Methods to Avoid Mitochondrial Disease Through Assisted Conception: 2016 update. (Human Fertilisation and Embryology Authority, London, 2016).
Hyslop, L. A. et al. Towards clinical application of pronuclear transfer to prevent mitochondrial DNA disease. Nature 534, 383–386 (2016).
Committee on the Ethical and Social Policy Considerations of Novel Techniques for Prevention of Maternal Transmission of Mitochondrial DNA Diseases; Board on Health Sciences Policy; Institute of Medicine; National Academies of Sciences, Engineering, and Medicine (eds Claiborne, A., English, R. & Kahn, J.) Mitochondrial replacement techniques: Ethical, social, and policy considerations. (National Academies Press, Washington DC, 2016).
Deglincerti, A. et al. Self-organization of the in vitro attached human embryo. Nature 533, 251–254 (2016).
Shahbazi, M. N. et al. Self-organization of the human embryo in the absence of maternal tissues. Nat. Cell Biol. 18, 700–708 (2016).
Macaulay, I. C. et al. G&T-seq: parallel sequencing of single-cell genomes and transcriptomes. Nat. Methods 12, 519–522 (2015).
de Silva, E. & Stumpf, M. P. HIV and the CCR5-Delta32 resistance allele. FEMS Microbiol. Lett. 241, 1–12 (2004).
Luzzatto, L. Sickle cell anaemia and malaria. Mediterr. J. Hematol. Infect. Dis. 4, e2012065 (2012).
Dzau, V. J., McNutt, M. & Bai, C. Wake-up call from Hong Kong. Science 362, 1215 (2018).
We thank members of the Niakan, J. Turner and R. Lovell-Badge laboratories for helpful discussions and comments on the Perspective. We are grateful to J. Brock in the Scientific Illustrations team for generating figures. Work in the Niakan laboratory is supported by the Francis Crick Institute, which receives its core funding from Cancer Research UK (FC001120), the UK Medical Research Council (FC001120), and the Wellcome Trust (FC001120).
The authors declare no competing interests.
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A. Lea, R., K. Niakan, K. Human germline genome editing. Nat Cell Biol 21, 1479–1489 (2019). https://doi.org/10.1038/s41556-019-0424-0
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