Skip to main content
SpringerLink
Log in
Book cover

HIV Reservoirs pp 429–445Cite as

Pathways Toward a Functional HIV-1 Cure: Balancing Promise and Perils of CRISPR Therapy

  • Protocol
  • First Online:

Part of the book series: Methods in Molecular Biology ((MIMB,volume 2407))

Abstract

First identified as a viral defense mechanism, clustered regularly interspaced short palindromic repeats (CRISPR) and CRISPR-associated proteins (Cas) has been transformed into a gene-editing tool. It now affords promise in the treatment and potential eradication of a range of divergent genetic, cancer, infectious, and degenerative diseases. Adapting CRISPR-Cas into a programmable endonuclease directed guide RNA (gRNA) has attracted international attention. It was recently awarded the 2020 Nobel Prize in Chemistry. The limitations of this technology have also been identified and work has been made in providing potential remedies. For treatment of the human immunodeficiency virus type one (HIV-1), in particular, a CRISPR-Cas9 approach was adapted to target then eliminate latent proviral DNA. To this end, we reviewed the promise and perils of CRISPR-Cas gene-editing strategies for HIV-1 elimination. Obstacles include precise delivery to reservoir tissue and cell sites of latent HIV-1 as well as assay sensitivity and specificity. The detection and consequent excision of common viral strain sequences and the avoidance of off-target activity will serve to facilitate a final goal of HIV-1 DNA elimination and accelerate testing in infected animals ultimately for use in man.

This is a preview of subscription content, log in via an institution.

Protocol
USD   49.95
Price excludes VAT (USA)
  • Available as PDF
  • Read on any device
  • Instant download
  • Own it forever
eBook
USD   99.00
Price excludes VAT (USA)
  • Available as EPUB and PDF
  • Read on any device
  • Instant download
  • Own it forever
Softcover Book
USD   129.99
Price excludes VAT (USA)
  • Compact, lightweight edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info
Hardcover Book
USD   199.99
Price excludes VAT (USA)
  • Durable hardcover edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info

Tax calculation will be finalised at checkout

Purchases are for personal use only

Learn about institutional subscriptions

Springer Nature is developing a new tool to find and evaluate Protocols. Learn more

References

  1. Panfil AR, London JA, Green PL, Yoder KE (2018) CRISPR/Cas9 genome editing to disable the latent HIV-1 provirus. Front Microbiol 9:3107–3107. https://doi.org/10.3389/fmicb.2018.03107

    Article  PubMed  PubMed Central  Google Scholar 

  2. Jinek M, Chylinski K, Fonfara I, Hauer M, Doudna JA, Charpentier E (2012) A programmable dual-RNA-guided DNA endonuclease in adaptive bacterial immunity. Science 337:816–821. https://doi.org/10.1126/science.1225829

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  3. Cohen J (2020) CRISPR, the revolutionary genetic ‘scissors,’ honored by Chemistry Nobel. 2020. https://www.sciencemag.org/news/2020/10/crispr-revolutionary-genetic-scissors-honored-chemistry-nobel

  4. Yoder KE, Bundschuh R (2016) Host double strand break repair generates HIV-1 strains resistant to CRISPR/Cas9. Sci Rep 6:29530. https://doi.org/10.1038/srep29530

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  5. Liao HK, Gu Y, Diaz A, Marlett J, Takahashi Y, Li M, Suzuki K, Xu R, Hishida T, Chang CJ, Esteban CR, Young J, Izpisua Belmonte JC (2015) Use of the CRISPR/Cas9 system as an intracellular defense against HIV-1 infection in human cells. Nat Commun 6:6413. https://doi.org/10.1038/ncomms7413

    Article  CAS  PubMed  Google Scholar 

  6. Wang G, Zhao N, Berkhout B, Das AT (2016) CRISPR-Cas9 can inhibit HIV-1 replication but NHEJ repair facilitates virus escape. Mol Ther 24:522–526. https://doi.org/10.1038/mt.2016.24

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  7. Wang Q, Liu S, Liu Z, Ke Z, Li C, Yu X, Chen S, Guo D (2018) Genome scale screening identification of SaCas9/gRNAs for targeting HIV-1 provirus and suppression of HIV-1 infection. Virus Res 250:21–30. https://doi.org/10.1016/j.virusres.2018.04.002

    Article  CAS  PubMed  Google Scholar 

  8. Gao Z, Fan M, Das AT, Herrera-Carrillo E, Berkhout B (2020) Extinction of all infectious HIV in cell culture by the CRISPR-Cas12a system with only a single crRNA. Nucleic Acids Res 48:5527–5539. https://doi.org/10.1093/nar/gkaa226

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  9. Kaminski R, Chen Y, Fischer T, Tedaldi E, Napoli A, Zhang Y, Karn J, Hu W, Khalili K (2016) Elimination of HIV-1 genomes from human T-lymphoid cells by CRISPR/Cas9 gene editing. Sci Rep 6:22555. https://doi.org/10.1038/srep22555

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  10. Hu W, Kaminski R, Yang F, Zhang Y, Cosentino L, Li F, Luo B, Alvarez-Carbonell D, Garcia-Mesa Y, Karn J, Mo X, Khalili K (2014) RNA-directed gene editing specifically eradicates latent and prevents new HIV-1 infection. Proc Natl Acad Sci U S A 111:11461–11466. https://doi.org/10.1073/pnas.1405186111

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  11. Yin L, Hu S, Mei S, Sun H, Xu F, Li J, Zhu W, Liu X, Zhao F, Zhang D, Cen S, Liang C, Guo F (2018) CRISPR/Cas9 inhibits multiple steps of HIV-1 infection. Hum Gene Ther 29:1264–1276. https://doi.org/10.1089/hum.2018.018

    Article  CAS  PubMed  Google Scholar 

  12. Yin L, Zhao F, Sun H, Wang Z, Huang Y, Zhu W, Xu F, Mei S, Liu X, Zhang D, Wei L, Cen S, Hu S, Liang C, Guo F (2020) CRISPR-Cas13a inhibits HIV-1 infection. Mol Ther Nucleic Acids 21:147–155. https://doi.org/10.1016/j.omtn.2020.05.030

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  13. Yin C, Zhang T, Li F, Yang F, Putatunda R, Young WB, Khalili K, Hu W, Zhang Y (2016) Functional screening of guide RNAs targeting the regulatory and structural HIV-1 viral genome for a cure of AIDS. AIDS 30:1163–1174. https://doi.org/10.1097/QAD.0000000000001079

    Article  CAS  PubMed  Google Scholar 

  14. Binda CS, Klaver B, Berkhout B, Das AT (2020) CRISPR-Cas9 dual-gRNA attack causes mutation, excision and inversion of the HIV-1 proviral DNA. Viruses 12:330. https://doi.org/10.3390/v12030330

    Article  CAS  PubMed Central  Google Scholar 

  15. Ophinni Y, Inoue M, Kotaki T, Kameoka M (2018) CRISPR/Cas9 system targeting regulatory genes of HIV-1 inhibits viral replication in infected T-cell cultures. Sci Rep 8:7784. https://doi.org/10.1038/s41598-018-26190-1

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  16. Zhu W, Lei R, Le Duff Y, Li J, Guo F, Wainberg MA, Liang C (2015) The CRISPR/Cas9 system inactivates latent HIV-1 proviral DNA. Retrovirology 12:22. https://doi.org/10.1186/s12977-015-0150-z

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  17. Vergara-Mendoza M, Gomez-Quiroz LE, Miranda-Labra RU, Fuentes-Romero LL, Romero-Rodriguez DP, Gonzalez-Ruiz J, Hernandez-Rizo S, Viveros-Rogel M (2020) Regulation of Cas9 by viral proteins Tat and Rev for HIV-1 inactivation. Antivir Res 180:104856. https://doi.org/10.1016/j.antiviral.2020.104856

    Article  CAS  PubMed  Google Scholar 

  18. Darcis G, Binda CS, Klaver B, Herrera-Carrillo E, Berkhout B, Das AT (2019) The impact of HIV-1 genetic diversity on CRISPR-Cas9 antiviral activity and viral escape. Viruses 11:225. https://doi.org/10.3390/v11030255

    Article  CAS  Google Scholar 

  19. Dampier W, Sullivan NT, Chung CH, Mell JC, Nonnemacher MR, Wigdahl B (2017) Designing broad-spectrum anti-HIV-1 gRNAs to target patient-derived variants. Sci Rep 7:14413. https://doi.org/10.1038/s41598-017-12612-z

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  20. Roychoudhury P, De Silva Feelixge H, Reeves D, Mayer BT, Stone D, Schiffer JT, Jerome KR (2018) Viral diversity is an obligate consideration in CRISPR/Cas9 designs for targeting the HIV reservoir. BMC Biol 16:75. https://doi.org/10.1186/s12915-018-0544-1

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  21. Dampier W, Sullivan NT, Mell JC, Pirrone V, Ehrlich GD, Chung CH, Allen AG, DeSimone M, Zhong W, Kercher K, Passic S, Williams JW, Szep Z, Khalili K, Jacobson JM, Nonnemacher MR, Wigdahl B (2018) Broad-spectrum and personalized guide RNAs for CRISPR/Cas9 HIV-1 therapeutics. AIDS Res Hum Retrovir 34:950–960. https://doi.org/10.1089/AID.2017.0274

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  22. Sullivan NT, Dampier W, Chung CH, Allen AG, Atkins A, Pirrone V, Homan G, Passic S, Williams J, Zhong W, Kercher K, Desimone M, Li L, Antell GC, Mell JC, Ehrlich GD, Szep Z, Jacobson JM, Nonnemacher MR, Wigdahl B (2019) Novel gRNA design pipeline to develop broad-spectrum CRISPR/Cas9 gRNAs for safe targeting of the HIV-1 quasispecies in patients. Sci Rep 9:17088. https://doi.org/10.1038/s41598-019-52353-9

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  23. Nerys-Junior A, Braga-Dias LP, Pezzuto P, Cotta-de-Almeida V, Tanuri A (2018) Comparison of the editing patterns and editing efficiencies of TALEN and CRISPR-Cas9 when targeting the human CCR5 gene. Genet Mol Biol 41:167–179. https://doi.org/10.1590/1678-4685-GMB-2017-0065

    Article  PubMed  PubMed Central  Google Scholar 

  24. Qi C, Li D, Jiang X, Jia X, Lu L, Wang Y, Sun J, Shao Y, Wei M (2018) Inducing CCR5Delta32/Delta32 homozygotes in the human jurkat CD4+ cell line and primary CD4+ cells by CRISPR-Cas9 Genome-Editing Technology. Mol Ther Nucleic Acids 12:267–274. https://doi.org/10.1016/j.omtn.2018.05.012

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  25. Xu L, Wang J, Liu Y, Xie L, Su B, Mou D, Wang L, Liu T, Wang X, Zhang B, Zhao L, Hu L, Ning H, Zhang Y, Deng K, Liu L, Lu X, Zhang T, Xu J, Li C, Wu H, Deng H, Chen H (2019) CRISPR-edited stem cells in a patient with HIV and acute lymphocytic leukemia. N Engl J Med 381:1240–1247. https://doi.org/10.1056/NEJMoa1817426

    Article  CAS  PubMed  Google Scholar 

  26. Liu S, Wang Q, Yu X, Li Y, Guo Y, Liu Z, Sun F, Hou W, Li C, Wu L, Guo D, Chen S (2018) HIV-1 inhibition in cells with CXCR4 mutant genome created by CRISPR-Cas9 and piggyBac recombinant technologies. Sci Rep 8:8573. https://doi.org/10.1038/s41598-018-26894-4

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  27. Hou P, Chen S, Wang S, Yu X, Chen Y, Jiang M, Zhuang K, Ho W, Hou W, Huang J, Guo D (2015) Genome editing of CXCR4 by CRISPR/cas9 confers cells resistant to HIV-1 infection. Sci Rep 5:15577. https://doi.org/10.1038/srep15577

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  28. Schumann K, Lin S, Boyer E, Simeonov DR, Subramaniam M, Gate RE, Haliburton GE, Ye CJ, Bluestone JA, Doudna JA, Marson A (2015) Generation of knock-in primary human T cells using Cas9 ribonucleoproteins. Proc Natl Acad Sci U S A 112:10437–10442. https://doi.org/10.1073/pnas.1512503112

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  29. Hultquist JF, Schumann K, Woo JM, Manganaro L, McGregor MJ, Doudna J, Simon V, Krogan NJ, Marson A (2016) A Cas9 ribonucleoprotein platform for functional genetic studies of HIV-host interactions in primary human T cells. Cell Rep 17:1438–1452. https://doi.org/10.1016/j.celrep.2016.09.080

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  30. Liu Z, Chen S, Jin X, Wang Q, Yang K, Li C, Xiao Q, Hou P, Liu S, Wu S, Hou W, Xiong Y, Kong C, Zhao X, Wu L, Li C, Sun G, Guo D (2017) Genome editing of the HIV co-receptors CCR5 and CXCR4 by CRISPR-Cas9 protects CD4(+) T cells from HIV-1 infection. Cell Biosci 7:47. https://doi.org/10.1186/s13578-017-0174-2

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  31. Yu S, Yao Y, Xiao H, Li J, Liu Q, Yang Y, Adah D, Lu J, Zhao S, Qin L, Chen X (2018) Simultaneous knockout of CXCR4 and CCR5 genes in CD4+ T cells via CRISPR/Cas9 confers resistance to both X4- and R5-tropic human immunodeficiency virus type 1 infection. Hum Gene Ther 29:51–67. https://doi.org/10.1089/hum.2017.032

    Article  CAS  PubMed  Google Scholar 

  32. Dickerson JE, Pinney JW, Robertson DL (2010) The biological context of HIV-1 host interactions reveals subtle insights into a system hijack. BMC Syst Biol 4:80. https://doi.org/10.1186/1752-0509-4-80

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  33. Park RJ, Wang T, Koundakjian D, Hultquist JF, Lamothe-Molina P, Monel B, Schumann K, Yu H, Krupzcak KM, Garcia-Beltran W, Piechocka-Trocha A, Krogan NJ, Marson A, Sabatini DM, Lander ES, Hacohen N, Walker BD (2017) A genome-wide CRISPR screen identifies a restricted set of HIV host dependency factors. Nat Genet 49:193–203. https://doi.org/10.1038/ng.3741

    Article  CAS  PubMed  Google Scholar 

  34. Taylor JP, Cash MN, Santostefano KE, Nakanishi M, Terada N, Wallet MA (2018) CRISPR/Cas9 knockout of USP18 enhances type I IFN responsiveness and restricts HIV-1 infection in macrophages. J Leukoc Biol 103:1225–1240. https://doi.org/10.1002/JLB.3MIA0917-352R

    Article  CAS  Google Scholar 

  35. Osei Kuffour E, Schott K, Jaguva Vasudevan AA, Holler J, Schulz WA, Lang PA, Lang KS, Kim B, Haussinger D, Konig R, Munk C (2018) USP18 (UBP43) abrogates p21-mediated inhibition of HIV-1. J Virol 92:e00592-18. https://doi.org/10.1128/JVI.00592-18

    Article  PubMed  PubMed Central  Google Scholar 

  36. Teng Y, Luo M, Yu T, Chen L, Huang Q, Chen S, Xie L, Zeng Y, Luo F, Xiong H, Liu Y, Hou W, Feng Y (2019) CRISPR/Cas9-mediated deletion of miR-146a enhances antiviral response in HIV-1 infected cells. Genes Immun 20:327–337. https://doi.org/10.1038/s41435-018-0036-x

    Article  CAS  PubMed  Google Scholar 

  37. Dufour C, Claudel A, Joubarne N, Merindol N, Maisonnet T, Masroori N, Plourde MB, Berthoux L (2018) Editing of the human TRIM5 gene to introduce mutations with the potential to inhibit HIV-1. PLoS One 13:e0191709. https://doi.org/10.1371/journal.pone.0191709

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  38. Ohainle M, Kim K, Komurlu Keceli S, Felton A, Campbell E, Luban J, Emerman M (2020) TRIM34 restricts HIV-1 and SIV capsids in a TRIM5alpha-dependent manner. PLoS Pathog 16:e1008507. https://doi.org/10.1371/journal.ppat.1008507

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  39. Meng B, Ip NCY, Abbink TEM, Kenyon JC, Lever AML (2020) ESCRT-II functions by linking to ESCRT-I in human immunodeficiency virus-1 budding. Cell Microbiol 22:e13161. https://doi.org/10.1111/cmi.13161

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  40. Meng B, Ip NC, Prestwood LJ, Abbink TE, Lever AM (2015) Evidence that the endosomal sorting complex required for transport-II (ESCRT-II) is required for efficient human immunodeficiency virus-1 (HIV-1) production. Retrovirology 12:72. https://doi.org/10.1186/s12977-015-0197-x

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  41. Lampi Y, Van Looveren D, Vranckx LS, Thiry I, Bornschein S, Debyser Z, Gijsbers R (2019) Targeted editing of the PSIP1 gene encoding LEDGF/p75 protects cells against HIV infection. Sci Rep 9:2389. https://doi.org/10.1038/s41598-019-38718-0

    Article  PubMed  PubMed Central  Google Scholar 

  42. Qu D, Sun WW, Li L, Ma L, Sun L, Jin X, Li T, Hou W, Wang JH (2019) Long noncoding RNA MALAT1 releases epigenetic silencing of HIV-1 replication by displacing the polycomb repressive complex 2 from binding to the LTR promoter. Nucleic Acids Res 47:3013–3027. https://doi.org/10.1093/nar/gkz117

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  43. Hirakawa MP, Krishnakumar R, Timlin JA, Carney JP, Butler KS (2020) Gene editing and CRISPR in the clinic: current and future perspectives. Biosci Rep 40:BSR20200127. https://doi.org/10.1042/BSR20200127

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  44. Günthard HF, Saag MS, Benson CA, del Rio C, Eron JJ, Gallant JE, Hoy JF, Mugavero MJ, Sax PE, Thompson MA, Gandhi RT, Landovitz RJ, Smith DM, Jacobsen DM, Volberding PA (2016) Antiretroviral drugs for treatment and prevention of HIV infection in adults: 2016 recommendations of the International Antiviral Society-USA Panel. JAMA 316:191–210. https://doi.org/10.1001/jama.2016.8900

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  45. Link RW, Nonnemacher MR, Wigdahl B, Dampier W (2018) Prediction of human immunodeficiency virus type 1 subtype-specific off-target effects arising from CRISPR-Cas9 gene editing therapy. CRISPR J 1:294–302. https://doi.org/10.1089/crispr.2018.0020

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  46. Ye L, Wang J, Beyer AI, Teque F, Cradick TJ, Qi Z, Chang JC, Bao G, Muench MO, Yu J, Levy JA, Kan YW (2014) Seamless modification of wild-type induced pluripotent stem cells to the natural CCR5Delta32 mutation confers resistance to HIV infection. Proc Natl Acad Sci U S A 111:9591–9596. https://doi.org/10.1073/pnas.1407473111

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  47. Sessions KJ, Chen YY, Hodge CA, Hudson TR, Eszterhas SK, Hayden MS, Howell AL (2020) Analysis of CRISPR/Cas9 guide RNA efficiency and specificity against genetically diverse HIV-1 isolates. AIDS Res Hum Retrovir 36:862–874. https://doi.org/10.1089/AID.2020.0055

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  48. Chung CH, Allen AG, Atkins AJ, Sullivan NT, Homan G, Costello R, Madrid R, Nonnemacher MR, Dampier W, Wigdahl B (2020) Safe CRISPR-Cas9 inhibition of HIV-1 with high specificity and broad-spectrum activity by targeting LTR NF-kappaB binding sites. Mol Ther Nucleic Acids 21:965–982. https://doi.org/10.1016/j.omtn.2020.07.016

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  49. Zhang Y, Arango G, Li F, Xiao X, Putatunda R, Yu J, Yang XF, Wang H, Watson LT, Zhang L, Hu W (2018) Comprehensive off-target analysis of dCas9-SAM-mediated HIV reactivation via long noncoding RNA and mRNA profiling. BMC Med Genet 11:78. https://doi.org/10.1186/s12920-018-0394-2

    Article  CAS  Google Scholar 

  50. Xiao Q, Chen S, Wang Q, Liu Z, Liu S, Deng H, Hou W, Wu D, Xiong Y, Li J, Guo D (2019) CCR5 editing by Staphylococcus aureus Cas9 in human primary CD4(+) T cells and hematopoietic stem/progenitor cells promotes HIV-1 resistance and CD4(+) T cell enrichment in humanized mice. Retrovirology 16:15. https://doi.org/10.1186/s12977-019-0477-y

    Article  PubMed  PubMed Central  Google Scholar 

  51. Liu Z, Liang J, Chen S, Wang K, Liu X, Liu B, Xia Y, Guo M, Zhang X, Sun G, Tian G (2020) Genome editing of CCR5 by AsCpf1 renders CD4(+)T cells resistance to HIV-1 infection. Cell Biosci 10:85. https://doi.org/10.1186/s13578-020-00444-w

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  52. Wang Z, Pan Q, Gendron P, Zhu W, Guo F, Cen S, Wainberg MA, Liang C (2016) CRISPR/Cas9-derived mutations both inhibit HIV-1 replication and accelerate viral escape. Cell Rep 15:481–489. https://doi.org/10.1016/j.celrep.2016.03.042

    Article  CAS  PubMed  Google Scholar 

  53. Wang Z, Wang W, Cui YC, Pan Q, Zhu W, Gendron P, Guo F, Cen S, Witcher M, Liang C (2018) HIV-1 employs multiple mechanisms to resist Cas9/single guide RNA targeting the viral primer binding site. J Virol 92:e01135-18. https://doi.org/10.1128/JVI.01135-18

    Article  PubMed  PubMed Central  Google Scholar 

  54. Ueda S, Ebina H, Kanemura Y, Misawa N, Koyanagi Y (2016) Anti-HIV-1 potency of the CRISPR/Cas9 system insufficient to fully inhibit viral replication. Microbiol Immunol 60:483–496. https://doi.org/10.1111/1348-0421.12395

    Article  CAS  PubMed  Google Scholar 

  55. Lebbink RJ, de Jong DC, Wolters F, Kruse EM, van Ham PM, Wiertz EJ, Nijhuis M (2017) A combinational CRISPR/Cas9 gene-editing approach can halt HIV replication and prevent viral escape. Sci Rep 7:41968. https://doi.org/10.1038/srep41968

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  56. Wang G, Zhao N, Berkhout B, Das AT (2016) A combinatorial CRISPR-Cas9 attack on HIV-1 DNA extinguishes all infectious provirus in infected T cell cultures. Cell Rep 17:2819–2826. https://doi.org/10.1016/j.celrep.2016.11.057

    Article  CAS  PubMed  Google Scholar 

  57. Scott T, Urak R, Soemardy C, Morris KV (2019) Improved Cas9 activity by specific modifications of the tracrRNA. Sci Rep 9:16104. https://doi.org/10.1038/s41598-019-52616-5

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  58. Yan M, Wen J, Liang M, Lu Y, Kamata M, Chen IS (2015) Modulation of gene expression by polymer nanocapsule delivery of DNA cassettes encoding small RNAs. PLoS One 10:e0127986. https://doi.org/10.1371/journal.pone.0127986

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  59. Campbell LA, Coke LM, Richie CT, Fortuno LV, Park AY, Harvey BK (2019) Gesicle-mediated delivery of CRISPR/Cas9 Ribonucleoprotein complex for inactivating the HIV provirus. Mol Ther 27:151–163. https://doi.org/10.1016/j.ymthe.2018.10.002

    Article  CAS  PubMed  Google Scholar 

  60. Kaushik A, Yndart A, Atluri V, Tiwari S, Tomitaka A, Gupta P, Jayant RD, Alvarez-Carbonell D, Khalili K, Nair M (2019) Magnetically guided non-invasive CRISPR-Cas9/gRNA delivery across blood-brain barrier to eradicate latent HIV-1 infection. Sci Rep 9:3928. https://doi.org/10.1038/s41598-019-40222-4

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  61. Choi JG, Dang Y, Abraham S, Ma H, Zhang J, Guo H, Cai Y, Mikkelsen JG, Wu H, Shankar P, Manjunath N (2016) Lentivirus pre-packed with Cas9 protein for safer gene editing. Gene Ther 23:627–633. https://doi.org/10.1038/gt.2016.27

    Article  CAS  PubMed  Google Scholar 

  62. Wang Q, Chen S, Xiao Q, Liu Z, Liu S, Hou P, Zhou L, Hou W, Ho W, Li C, Wu L, Guo D (2017) Genome modification of CXCR4 by Staphylococcus aureus Cas9 renders cells resistance to HIV-1 infection. Retrovirology 14:51. https://doi.org/10.1186/s12977-017-0375-0

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  63. Wang B, Zuo J, Kang W, Wei Q, Li J, Wang C, Liu Z, Lu Y, Zhuang Y, Dang B, Liu Q, Kang W, Sun Y (2018) Generation of Hutat2:Fc knockin primary human monocytes using CRISPR/Cas9. Mol Ther Nucleic Acids 11:130–141. https://doi.org/10.1016/j.omtn.2018.01.012

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  64. Li C, Guan X, Du T, Jin W, Wu B, Liu Y, Wang P, Hu B, Griffin GE, Shattock RJ, Hu Q (2015) Inhibition of HIV-1 infection of primary CD4+ T-cells by gene editing of CCR5 using adenovirus-delivered CRISPR/Cas9. J Gen Virol 96:2381–2393. https://doi.org/10.1099/vir.0.000139

    Article  CAS  PubMed  Google Scholar 

  65. Fakhiri J, Nickl M, Grimm D (2019) Rapid and simple screening of CRISPR guide RNAs (gRNAs) in cultured cells using adeno-associated viral (AAV) vectors. Methods Mol Biol 1961:111–126. https://doi.org/10.1007/978-1-4939-9170-9_8

    Article  CAS  PubMed  Google Scholar 

  66. Kaminski R, Bella R, Yin C, Otte J, Ferrante P, Gendelman HE, Li H, Booze R, Gordon J, Hu W, Khalili K (2016) Excision of HIV-1 DNA by gene editing: a proof-of-concept in vivo study. Gene Ther 23:690–695. https://doi.org/10.1038/gt.2016.41

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  67. Bella R, Kaminski R, Mancuso P, Young WB, Chen C, Sariyer R, Fischer T, Amini S, Ferrante P, Jacobson JM, Kashanchi F, Khalili K (2018) Removal of HIV DNA by CRISPR from patient blood engrafts in humanized mice. Mol Ther Nucleic Acids 12:275–282. https://doi.org/10.1016/j.omtn.2018.05.021

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  68. Dash PK, Kaminski R, Bella R, Su H, Mathews S, Ahooyi TM, Chen C, Mancuso P, Sariyer R, Ferrante P, Donadoni M, Robinson JA, Sillman B, Lin Z, Hilaire JR, Banoub M, Elango M, Gautam N, Mosley RL, Poluektova LY, McMillan J, Bade AN, Gorantla S, Sariyer IK, Burdo TH, Young W-B, Amini S, Gordon J, Jacobson JM, Edagwa B, Khalili K, Gendelman HE (2019) Sequential LASER ART and CRISPR treatments eliminate HIV-1 in a subset of infected humanized mice. Nat Commun 10:2753. https://doi.org/10.1038/s41467-019-10366-y

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  69. Kong J, Wang Y, Zhang J, Qi W, Su R, He Z (2018) Rationally designed peptidyl virus-like particles enable targeted delivery of genetic cargo. Angew Chem Int Ed Engl 57:14032–14036. https://doi.org/10.1002/anie.201805868

    Article  CAS  PubMed  Google Scholar 

  70. Li L, Hu S, Chen X (2018) Non-viral delivery systems for CRISPR/Cas9-based genome editing: challenges and opportunities. Biomaterials 171:207–218. https://doi.org/10.1016/j.biomaterials.2018.04.031

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  71. Gee P, Lung MSY, Okuzaki Y, Sasakawa N, Iguchi T, Makita Y, Hozumi H, Miura Y, Yang LF, Iwasaki M, Wang XH, Waller MA, Shirai N, Abe YO, Fujita Y, Watanabe K, Kagita A, Iwabuchi KA, Yasuda M, Xu H, Noda T, Komano J, Sakurai H, Inukai N, Hotta A (2020) Extracellular nanovesicles for packaging of CRISPR-Cas9 protein and sgRNA to induce therapeutic exon skipping. Nat Commun 11:1334. https://doi.org/10.1038/s41467-020-14957-y

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  72. Geng X, Doitsh G, Yang Z, Galloway NLK, Greene WC (2014) Efficient delivery of lentiviral vectors into resting human CD4 T cells. Gene Ther 21:444–449. https://doi.org/10.1038/gt.2014.5

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  73. Fries CN, Curvino EJ, Chen J-L, Permar SR, Fouda GG, Collier JH (2021) Advances in nanomaterial vaccine strategies to address infectious diseases impacting global health. Nat Nanotechnol 16:1–14. https://doi.org/10.1038/s41565-020-0739-9

    Article  CAS  PubMed  Google Scholar 

  74. Li C, Lieber A (2019) Adenovirus vectors in hematopoietic stem cell genome editing. FEBS Lett 593:3623–3648. https://doi.org/10.1002/1873-3468.13668

    Article  CAS  PubMed  Google Scholar 

  75. Locatelli A (2020) Towards a novel therapy against AIDS. Med Hypotheses 137:109569. https://doi.org/10.1016/j.mehy.2020.109569

    Article  CAS  PubMed  Google Scholar 

  76. Zotova A, Lopatukhina E, Filatov A, Khaitov M, Mazurov D (2017) Gene editing in human lymphoid cells: role for donor DNA, type of genomic nuclease and cell selection method. Viruses 9:325. https://doi.org/10.3390/v9110325

    Article  CAS  PubMed Central  Google Scholar 

  77. Wollebo HS, Bellizzi A, Kaminski R, Hu W, White MK, Khalili K (2015) CRISPR/Cas9 system as an agent for eliminating polyomavirus JC infection. PLoS One 10:e0136046. https://doi.org/10.1371/journal.pone.0136046

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  78. Scott T, Moyo B, Nicholson S, Maepa MB, Watashi K, Ely A, Weinberg MS, Arbuthnot P (2017) ssAAVs containing cassettes encoding SaCas9 and guides targeting hepatitis B virus inactivate replication of the virus in cultured cells. Sci Rep 7:7401. https://doi.org/10.1038/s41598-017-07642-6

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  79. Aubert M, Strongin DE, Roychoudhury P, Loprieno MA, Haick AK, Klouser LM, Stensland L, Huang ML, Makhsous N, Tait A, De Silva Feelixge HS, Galetto R, Duchateau P, Greninger AL, Stone D, Jerome KR (2020) Gene editing and elimination of latent herpes simplex virus in vivo. Nat Commun 11:4148. https://doi.org/10.1038/s41467-020-17936-5

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  80. Yang Y-C, Chen Y-H, Kao J-H, Ching C, Liu IJ, Wang C-C, Tsai C-H, Wu F-Y, Liu C-J, Chen P-J, Chen D-S, Yang H-C (2020) Permanent inactivation of HBV genomes by CRISPR/Cas9-mediated non-cleavage base editing. Mol Ther Nucleic Acids 20:480–490. https://doi.org/10.1016/j.omtn.2020.03.005

    Article  CAS  PubMed  PubMed Central  Google Scholar 

Download references

Acknowledgments

This work was supported by the National Institutes of Health R01 AG043530, P01 DA028555, P30 MH062261, R01 MH115860, R01 NS034249, R01 NS036126, and the Carol Swartz Emerging Neuroscience Fund.

Author information

Authors and Affiliations

  1. Department of Pathology and Microbiology, University of Nebraska Medical Center, Omaha, NE, USA

    Jonathan Herskovitz & Howard E. Gendelman

  2. Department of Pharmaceutical Sciences, University of Nebraska Medical Center, Omaha, NE, USA

    Mahmudul Hasan & Howard E. Gendelman

  3. Department of Pharmacology and Experimental Neuroscience, University of Nebraska Medical Center, Omaha, NE, USA

    Milankumar Patel, Bhavesh D. Kevadiya & Howard E. Gendelman

Authors
  1. Jonathan Herskovitz
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Mahmudul Hasan
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Milankumar Patel
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Bhavesh D. Kevadiya
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Howard E. Gendelman
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Howard E. Gendelman .

Editor information

Editors and Affiliations

  1. School of Medicine and Human Immuno-Virology (H.I.V.) Group, San Raffaele University and Scientific Institute, Milano, Italy

    Guido Poli

  2. Viral Pathogenesis and Biosafety Group, San Raffaele Scientific Institute, Milano, Italy

    Elisa Vicenzi

  3. Department of Molecular and Comparative Pathobiology, Johns Hopkins University School of Medicine, Baltimore, MD, USA

    Fabio Romerio

Rights and permissions

Reprints and permissions

Copyright information

© 2022 Springer Science+Business Media, LLC, part of Springer Nature

About this protocol

Check for updates. Verify currency and authenticity via CrossMark

Cite this protocol

Herskovitz, J., Hasan, M., Patel, M., Kevadiya, B.D., Gendelman, H.E. (2022). Pathways Toward a Functional HIV-1 Cure: Balancing Promise and Perils of CRISPR Therapy. In: Poli, G., Vicenzi, E., Romerio, F. (eds) HIV Reservoirs. Methods in Molecular Biology, vol 2407. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-1871-4_27

Download citation

  • DOI: https://doi.org/10.1007/978-1-0716-1871-4_27

  • Published:

  • Publisher Name: Humana, New York, NY

  • Print ISBN: 978-1-0716-1870-7

  • Online ISBN: 978-1-0716-1871-4

  • eBook Packages: Springer Protocols

Publish with us

Policies and ethics

Search

Navigation