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Characterizing the antiviral effect of an ATR inhibitor on human immunodeficiency virus type 1 replication

Abstract

In the search for new antiviral therapies against human immunodeficiency virus type 1 (HIV-1), several cellular targets are being investigated. Ataxia telangiectasia and Rad3-related protein (ATR) has been implicated in HIV-1 replication, namely during retroviral DNA integration. We studied the effect of the ATR inhibitor ETP-46464 on HIV-1 replication in peripheral blood mononuclear cells (PBMCs) and in the persistently HIV-1-infected cell line H61-D. After treatment with ETP-46464, a significant decrease in virus production was observed in both cell systems. Quantification of viral DNA forms in the acutely infected PBMCs suggests that inhibition could take place in the early phase of the viral life cycle before viral DNA integration. Moreover, after treatment of H61-D cells with 3’-azido-3’-deoxythymidine (AZT), which blocks new reverse transcription events, ETP-46464 decreased viral production, suggesting that inhibition of viral replication occurred in the late phase of the life cycle after viral DNA integration. A decrease in virus production after transfection of 293T cells with an HIV-1 infectious molecular clone also suggested that the effect of ETP-46464 is exerted at a post-integration step. We propose that ETP-46464 produces its inhibitory effect on HIV-1 replication by acting in both the early and late phases of the retroviral replication cycle. Thus, ATR could represent a new target for inhibition of HIV-1 replication.

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References

  1. 1.

    Adachi A, Gendelman HE, Koenig S, Folks T, Willey R, Rabson A, Martin MA (1986) Production of acquired immunodeficiency syndrome-associated retrovirus in human and nonhuman cells transfected with an infectious molecular clone. JVirol 59:284–291

  2. 2.

    Andersen JL, Le RE, Planelles V (2008) HIV-1 Vpr: mechanisms of G2 arrest and apoptosis. ExpMolPathol 85:2–10

  3. 3.

    Ariumi Y, Turelli P, Masutani M, Trono D (2005) DNA damage sensors ATM, ATR, DNA-PKcs, and PARP-1 are dispensable for human immunodeficiency virus type 1 integration. JVirol 79:2973–2978

  4. 4.

    Belzile JP, Richard J, Rougeau N, Xiao Y, Cohen EA (2010) HIV-1 Vpr induces the K48-linked polyubiquitination and proteasomal degradation of target cellular proteins to activate ATR and promote G2 arrest. JVirol 84:3320–3330

  5. 5.

    Biswas DK, Ahlers CM, Dezube BJ, Pardee AB (1994) Pentoxifylline and other protein kinase C inhibitors down-regulate HIV-LTR NF-kappa B induced gene expression. MolMed 1:31–43

  6. 6.

    Brin E, Yi J, Skalka AM, Leis J (2000) Modeling the late steps in HIV-1 retroviral integrase-catalyzed DNA integration. JBiolChem 275:39287–39295

  7. 7.

    Cooper A, Garcia M, Petrovas C, Yamamoto T, Koup RA, Nabel GJ (2013) HIV-1 causes CD4 cell death through DNA-dependent protein kinase during viral integration. Nature 498:376–379

  8. 8.

    Daniel R, Kao G, Taganov K, Greger JG, Favorova O, Merkel G, Yen TJ, Katz RA, Skalka AM (2003) Evidence that the retroviral DNA integration process triggers an ATR-dependent DNA damage response. Proc Natl Acad Sci USA 100:4778–4783

  9. 9.

    Daniel R, Marusich E, Argyris E, Zhao RY, Skalka AM, Pomerantz RJ (2005) Caffeine inhibits human immunodeficiency virus type 1 transduction of nondividing cells. JVirol 79:2058–2065

  10. 10.

    DeHart JL, Andersen JL, Zimmerman ES, Ardon O, An DS, Blackett J, Kim B, Planelles V (2005) The ataxia telangiectasia-mutated and Rad3-related protein is dispensable for retroviral integration. J Virol 79:1389–1396

  11. 11.

    Gonzalez ME (2017) The HIV-1 Vpr protein: a multifaceted target for therapeutic intervention. Int J Mol Sci 18(1):126. https://doi.org/10.3390/ijms18010126

  12. 12.

    Guendel I, Meltzer BW, Baer A, Dever SM, Valerie K, Guo J, Wu Y, Kehn-Hall K (2015) BRCA1 functions as a novel transcriptional cofactor in HIV-1 infection. VirolJ 12:40

  13. 13.

    Harper JW, Elledge SJ (2007) The DNA damage response: 10 years after. MolCell 28:739–745

  14. 14.

    Saldivara JC, Cortez D, Cimpricha KA (2017) The essential kinase ATR: ensuring faithful duplication of a challenging genome. Nat Rev Mol Cell Biol 18(10):622–636

  15. 15.

    Lai RP, Yan J, Heeney J, McClure MO, Gottlinger H, Luban J, Pizzato M (2011) Nef decreases HIV-1 sensitivity to neutralizing antibodies that target the membrane-proximal external region of TMgp41. PLoSPathog 7:e1002442

  16. 16.

    Lopez-Contreras AJ, Fernandez-Capetillo O (2010) The ATR barrier to replication-born DNA damage. DNA Repair (Amst) 9:1249–1255

  17. 17.

    Lopez-Huertas MR, Mateos E, Del Sanchez CM, Gomez-Esquer F, Diaz-Gil G, Rodriguez-Mora S, Lopez JA, Calvo E, Lopez-Campos G, Alcami J, Coiras M (2013) The presence of HIV-1 Tat protein second exon delays fas protein-mediated apoptosis in CD4+ T lymphocytes: a potential mechanism for persistent viral production. JBiolChem 288:7626–7644

  18. 18.

    Mhashilkar AM, Biswas DK, LaVecchio J, Pardee AB, Marasco WA (1997) Inhibition of human immunodeficiency virus type 1 replication in vitro by a novel combination of anti-Tat single-chain intrabodies and NF-kappa B antagonists. JVirol 71:6486–6494

  19. 19.

    Nisole S, Saib A (2004) Early steps of retrovirus replicative cycle. Retrovirology 1:9

  20. 20.

    Nunnari G, Argyris E, Fang J, Mehlman KE, Pomerantz RJ, Daniel R (2005) Inhibition of HIV-1 replication by caffeine and caffeine-related methylxanthines. Virology 335:177–184

  21. 21.

    Olivares I, Shaw G, Lopez-Galindez C (1997) Phenotypic switch in a Spanish HIV type 1 isolate on serial passage on MT-4 cells. AIDS ResHumRetroviruses 13:979–984

  22. 22.

    Olivares I, Casado HC, Iglesias-Ussel MD, Dietrich U, Lopez GC (1998) Complete sequence of an infectious molecular clone derived from a Spanish HIV type I isolate. AIDS ResHumRetroviruses 14:1649–1651

  23. 23.

    Olivares I, Sanchez-Jimenez C, Vieira CR, Toledano V, Gutierrez-Rivas M, Lopez-Galindez C (2013) Evidence of ongoing replication in a human immunodeficiency virus type 1 persistently infected cell line. JGenVirol 94:944–954

  24. 24.

    Postigo ARA, Howell M, Way M (2017) Cytoplasmic ATR activation promotes vaccinia virus genome replication. Cell Rep 19:1022–1032

  25. 25.

    Roshal M, Kim B, Zhu Y, Nghiem P, Planelles V (2003) Activation of the ATR-mediated DNA damage response by the HIV-1 viral protein R. JBiolChem 278:25879–25886

  26. 26.

    Ryan EL, Hollingworth R, Grand RJ (2016) Activation of the DNA damage response by RNA viruses. Biomolecules 6:2

  27. 27.

    Sakurai Y, Komatsu K, Agematsu K, Matsuoka M (2009) DNA double strand break repair enzymes function at multiple steps in retroviral infection. Retrovirology 6:114

  28. 28.

    Sanchez-Jimenez C, Olivares I, de Avila Lucas AI, Toledano V, Gutierrez-Rivas M, Lorenzo-Redondo R, Grande-Perez A, Domingo E, Lopez-Galindez C (2012) Mutagen-mediated enhancement of HIV-1 replication in persistently infected cells. Virology 424:147–153

  29. 29.

    Sinclair A, Yarranton S, Schelcher C (2006) DNA-damage response pathways triggered by viral replication. ExpertRevMolMed 8:1–11

  30. 30.

    Smith JA, Nunnari G, Preuss M, Pomerantz RJ, Daniel R (2007) Pentoxifylline suppresses transduction by HIV-1-based vectors. Intervirology 50:377–386

  31. 31.

    Toledo LI, Murga M, Zur R, Soria R, Rodriguez A, Martinez S, Oyarzabal J, Pastor J, Bischoff JR, Fernandez-Capetillo O (2011) A cell-based screen identifies ATR inhibitors with synthetic lethal properties for cancer-associated mutations. NatStructMolBiol 18:721–727

  32. 32.

    Vandergeeten C, Fromentin R, Merlini E, Lawani MB, DaFonseca S, Bakeman W, McNulty A, Ramgopal M, Michael N, Kim JH, Ananworanich J, Chomont N (2014) Cross-clade ultrasensitive PCR-based assays to measure HIV persistence in large-cohort studies. JVirol 88:12385–12396

  33. 33.

    Vassena L, Giuliani E, Matusali G, Cohen EA, Doria M (2013) The human immunodeficiency virus type 1 Vpr protein upregulates PVR via activation of the ATR-mediated DNA damage response pathway. JGenVirol 94:2664–2669

  34. 34.

    Yang YX, Guen V, Richard J, Cohen EA, Berthoux L (2010) Cell context-dependent involvement of ATR in early stages of retroviral replication. Virology 396:272–279

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Acknowledgements

We thank Oscar Fernandez-Capetillo for providing the ATR inhibitor and for helpful suggestions, and Mayte Coiras and José Alcamí for the pCMV-Tat 101 plasmid. This work was supported by grants SAF (2010-17226) and (2016-77894-R) from MINECO (Ministerio de Economia y Competitividad, Spain) and PI 13/02269 from FIS (Fondo de Investigación en Salud del Instituto de Salud Carlos III) and in part by the RIS-RETIC grants   RD12/0017/0028 and RD16CIII/0002/0005 funded by de ISCIII FEDER). Felix Docando had a Research Assistant Grant from the Programa de Empleo Juvenil of Comunidad de Madrid.

Author information

CL-G and IO conceived the study and designed the experiments. FD and IO performed, supervised and analyzed the experiments. FD, CC, MP and AM-B performed experiments and analyzed data. IO and CL-G wrote the manuscript. All authors contributed to the critical revision of the manuscript.

Correspondence to Isabel Olivares.

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Docando, F., Casado, C., Pernas, M. et al. Characterizing the antiviral effect of an ATR inhibitor on human immunodeficiency virus type 1 replication. Arch Virol 165, 683–690 (2020). https://doi.org/10.1007/s00705-020-04531-8

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