Advertisement

Gene Editing in Clinical Practice: Where are We?

  • Rama Devi MittalEmail author
Review Article
  • 44 Downloads

Abstract

Multitude of gene-altering capabilities in combination with ease of design and low cost have all led to the adoption of the sophisticated and yet simple gene editing system that are clustered regularly interspaced short palindromic repeats (CRISPR)-associated protein 9 (CRISPR). The CRISPR/Cas9 system holds promise for the correction of deleterious mutations by taking advantage of the homology directed repair pathway and by supplying a correction template to the affected patient’s cells. CRISPR is a tool that allows researchers to edit genes very precisely, easily and quickly. It does this by harnessing a mechanism that already existed in bacteria. Basically, there’s a protein that acts like a scissors and cuts the DNA, and there’s an RNA molecule that directs the scissors to any point on the genome one wants which results basically a word processor for genes. An entire gene can be taken out, put one in, or even edit just a single letter within a gene. Several platforms for molecular scissors that enable targeted genome engineering have been developed, including zinc-finger nucleases, transcription activator-like effector nucleases and, most recently, CRISPR/CRISPR-associated-9 (Cas9). The CRISPR/Cas9 system’s simplicity, facile engineering and amenability to multiplexing make it the system of choice for many applications. CRISPR/Cas9 has been used to generate disease models to study genetic diseases. Improvements are urgently needed for various aspects of the CRISPR/Cas9 system, including the system’s precision, delivery and control over the outcome of the repair process. However, there are still some glitches to be mended like how to regulate gene drives and its safeguards. The creation of gene knockouts is one of the first and most widely used applications of the CRISPR–Cas9 system. Nuclease-active Cas9 creates a double-strand break at the single guide RNA-targeted locus. These breaks can be repaired by homologous recombination, which can be used to introduce new mutations. When the double-strand break is repaired by the error-prone nonhomologous end joining process, indels are introduced which can produce frame shifts and stop codons, leading to functional knockout of the gene. Precedence modification have to be done on mechanism of CRISPR/Cas9, including its biochemical and structural implications incorporating the latest improvements in the CRISPR/Cas9 system, especially Cas9 protein modifications for customization. Current applications, where the versatile CRISPR/Cas9 system is to be used to edit the genome, epigenome, or RNA of various organisms is debated. Although CRISPR/Cas9 allows convenient genome editing accompanied by many benefits, one should not ignore the significant ethical and biosafety concerns that it raises. Conclusively lot of prospective applications and challenges of several promising techniques adapted from CRISPR/Cas9. Is discussed. Although many mechanistic questions remain to be answered and several challenges to be addressed yet, the use of CRISPR–Cas9-based genome technologies will increase our knowledge of disease process and their treatment in near future. Undoubtedly this field is revolutionizing in current era and may open new vistas in the treatment of fatal genetic disease.

Keywords

Genome editing CASPR Cas9 systems Molecular scissors Palindromic repeats Nucleases Gene targeting Gene therapy embryo Ethics Potential pitfalls 

References

  1. 1.
    Veltman JA, Brunner HG. De novo mutations in human genetic disease. Nat Rev Genet. 2012;13:565–75.CrossRefGoogle Scholar
  2. 2.
    Yoshimi K, Kaneko T, Voigt B, Mashimo T. Allele-specific genome editing and correction of disease-associated phenotypes in rats using the CRISPR–Cas platform. Nat Commun. 2014;5:4240–58.CrossRefGoogle Scholar
  3. 3.
    Ishino Y, Krupovic M, Forterre P. History of CRISPR–Cas from encounter with a mysterious repeated sequence to genome editing technology. J Bacteriol. 2018.  https://doi.org/10.1128/JB.00580-17.CrossRefPubMedPubMedCentralGoogle Scholar
  4. 4.
    Scharf I, Bierbaumer L, Huber H, Wittmann P, Haider C, Pirker C, Berger W, Mikulits W. Dynamics of CRISPR/Cas9-mediated genomic editing of the AXL locus in hepatocellular carcinoma cells. Oncol Lett. 2018;15(2):2441–50.PubMedGoogle Scholar
  5. 5.
    Motta BM, Pramstaller PP, Hicks AA, Rossini A. The impact of CRISPR/Cas9 technology on cardiac research: from disease modelling to therapeutic approaches. Stem Cells Int. 2017.  https://doi.org/10.1155/2017/8960236.CrossRefPubMedPubMedCentralGoogle Scholar
  6. 6.
    Lemay ML, Horvath P, Moineau S. The CRISPR–Cas app goes viral. Curr Opin Microbiol. 2017;37:103–9.CrossRefGoogle Scholar
  7. 7.
    Fellmann C, Gowen BG, Lin PC, Doudna JA, Corn JE. Cornerstones of CRISPR–Cas in drug discovery and therapy. Nat Rev Drug Discov. 2017;16(2):89–100.CrossRefGoogle Scholar
  8. 8.
    Doudna JA, Gersbach CA. Genome editing: the end of the beginning. Genome Biol. 2015;16:292–4.CrossRefGoogle Scholar
  9. 9.
    Hille F, Charpentier E. CRISPR–Cas: biology, mechanisms and relevance. Philos Trans R Soc Lond B Biol Sci. 2016;371(1707):1–11.CrossRefGoogle Scholar
  10. 10.
    Cong L, Ran FA, Cox D, Lin S, Barretto R, Zhang F, et al. Multiplex genome engineering using CRISPR/Cas systems. Science. 2013;339(6121):819–23.CrossRefGoogle Scholar
  11. 11.
    Platt RJ, Chen S, Zhou Y, Yim MJ, Swiech L, Zhang F. CRISPR–Cas9 knockin mice for genome editing and cancer modeling. Cell. 2014;159(2):440–55.CrossRefGoogle Scholar
  12. 12.
    Stupp R, Hegi ME, Mason WP, van den Bent MJ, Taphoorn MJ, Janzer RC, et al. Effects of radiotherapy with concomitant and adjuvant temozolomide versus radiotherapy alone on survival in glioblastoma in a randomised phase III study: 5-year analysis of the EORTC-NCIC trial. Lancet Oncol. 2009;10:459–66.CrossRefGoogle Scholar
  13. 13.
    Ran FA, Hsu PD, Wright J, Agarwala V, Scott DA, Zhang F. Genome engineering using the CRISPR–Cas9 system. Nat Protoc. 2013;8(11):2281–307.CrossRefGoogle Scholar
  14. 14.
    Chin A. CRISPR–Cas9 therapeutics: a technology overview. Oxford: Oxford Biotechnology; 2015.Google Scholar
  15. 15.
    Chowdhury R, Chinmaya AM. Translating cancer genomes and transcriptomes for precision oncology. CA. 2016;66:75–88.Google Scholar
  16. 16.
    Khan FA, Pandupuspitasari PS, Chun-Jie H, Zhou A, Jamal M, Zohaib S, et al. CRISPR/Cas9 therapeutics: a cure for cancer and other genetic diseases. Oncotarget. 2016;7(32):52541–52.CrossRefGoogle Scholar
  17. 17.
    Cai M, Yang Y. Targeted genome editing tools for disease modelling and gene therapy. Curr Gene Ther. 2014;14:2–9.CrossRefGoogle Scholar
  18. 18.
    Rongxue P, Guigao L, Jinming L. Potential pitfalls of CRISPR/Cas9-mediated genome editing. FEBS J. 2016;283:1218–23.CrossRefGoogle Scholar
  19. 19.
    Plaza RA, Lanner F. Towards a CRISPR view of early human development: applications, limitations and ethical concerns of genome editing in human embryos. Development. 2017;144:3–7.CrossRefGoogle Scholar
  20. 20.
    Eid A, Magdy MM. Genome editing: the road of CRISPR/Cas9 from bench to clinic. Exp Mol Med. 2016;48:1–11.CrossRefGoogle Scholar
  21. 21.
    Tatjana IC, Claudio M, Toni C. Refining strategies to translate genome editing to the clinic. Nat Med. 2017;23(4):415–23.CrossRefGoogle Scholar
  22. 22.
    Otieno MO. CRISPR–Cas9 human genome editing: challenges, ethical concerns and implications. J Clin Res Bioeth. 2015;6(6):1–3.Google Scholar
  23. 23.
    Chu VT, et al. Increasing the efficiency of homology-directed repair for CRISPR–Cas9-induced precise gene editing in mammalian cells. Nat Biotechnol. 2015;33:543–8.CrossRefGoogle Scholar
  24. 24.
    Maruyama T, et al. Increasing the efficiency of precise genome editing with CRISPR–Cas9 by inibition of nonhomologous end joining. Nat Biotechnol. 2015;33:538–42.CrossRefGoogle Scholar
  25. 25.
    Lanphier E, Urnov F, Haecker SE, Werner M, Smolenski J. Don’t edit the human germ line. Nature. 2015;519(7544):410–1.CrossRefGoogle Scholar
  26. 26.
    Baltimore D, Berg P, Botchan M, Carroll D, Charo RA, Church G, et al. Biotechnology. A prudent path forward for genomic engineering and germline gene modification. Science. 2015;348(6230):36–8.CrossRefGoogle Scholar
  27. 27.
    Men K, Xingmei D, Zhiyao H, Yang Y, Shaohua Y, Yuquan W. CRISPR/Cas9-mediated correction of human genetic disease. Sci China Life Sci. 2017;60(5):447–57.CrossRefGoogle Scholar
  28. 28.
    Wang HX, Li M, Lee CM, Chakraborty S, Kim HW, Bao G, Leong KW. CRISPR/Cas9-based genome editing for disease modeling and therapy: challenges and opportunities for non-viral delivery. Chem Rev. 2017;117(15):9874–906.CrossRefGoogle Scholar
  29. 29.
    Lau V, Davie JR. The discovery and development of the CRISPR system in applications in genome manipulation. Biochem Cell Biol. 2017;95(2):203–10.CrossRefGoogle Scholar
  30. 30.
    Xiao-Jie L, Hui-Ying X, Zun-Ping K, Jin-Lian C, Li-Juan J. CRISPR–Cas9: a new and promising player in gene therapy. J Med Genet. 2015;52(5):289–96.CrossRefGoogle Scholar
  31. 31.
    Liang P, Ding C, Sun H, Xie X, Xu Y, Zhang X. Correction of β-thalassemia mutant by base editor in human embryos. Protein Cell. 2017;8(11):811–22.CrossRefGoogle Scholar
  32. 32.
    Ma H, Marti-Gutierrez N, Mitalipov S. Correction of a pathogenic gene mutation in human embryos. Nature. 2017;548:413–9.CrossRefGoogle Scholar
  33. 33.
    Mulvihill JJ, Capps B, Joly Y, Hub T, Zwart AE, Chadwick R. Ethical issues of CRISPR technology and gene editing through the lens of solidarity: the International Human Genome Organisation (HUGO) Committee of Ethics, Law, and Society (CELS). Br Med Bull. 2017;122(1):17–29.CrossRefGoogle Scholar
  34. 34.
    Nordberg A, Minssen T, Holm S, Horst M, Mortensen K, Møller BL. Cutting edges and weaving threads in the gene editing (Я)evolution: reconciling scientific progress with legal, ethical, and social concerns. J Law Biosci. 2018;5(1):35–83.CrossRefGoogle Scholar
  35. 35.
    Cornu TI, Mussolino C, Cathomen T. Refining strategies to translate genome editing to the clinic. Nat Med. 2017;23(4):415–23.CrossRefGoogle Scholar
  36. 36.
    Thurtle-Schmidt DM, Lo TW. Molecular biology at the cutting edge: a review on CRISPR/CAS9 gene editing for undergraduates. Biochem Mol Biol Educ. 2018;46(2):195–205.CrossRefGoogle Scholar
  37. 37.
    Howard HC, van Carla G, Forzano F, Radojkovic D, Rial-Sebbag E, de Wert G, Borry P, Cornel MC. One small edit for humans, one giant edit for humankind? Points and questions to consider for a responsible way forward for gene editing in humans. Eur J Hum Genet. 2018;26:1–11.CrossRefGoogle Scholar
  38. 38.
    Roy B, Zhao J, Yang C, Luo W, Xiong T, Li Y, Fang X, Gao G, Singh CO, Madsen L, Zhou Y, Kristiansen K. CRISPR/Cascade 9-mediated genome editing-challenges and opportunities. Front Genet. 2018;9:240–50.CrossRefGoogle Scholar

Copyright information

© Association of Clinical Biochemists of India 2019

Authors and Affiliations

  1. 1.Department of Urology and Renal TransplantationSanjay Gandhi Post Graduate Institute of Medical SciencesLucknowIndia

Personalised recommendations