Effect of the reverse transcriptase inhibitor efavirenz on 5-azacytidine-induced LINE1 long interspersed repeat expression and genetic instability in acute myeloid leukemia cells

Cover Page

Cite item

Full Text

Abstract

Introduction. The use of hypomethylating agents in the treatment of acute myeloid leukemia has increased overall patient survival by 12 %. However, alongside their potent antitumor effect, hypomethylating agents negatively impact genome stability due to the transposition of activated LINE1 elements, which contributes to the short duration of remission. Since LINE1 retrotransposition requires the ORF2-encoded reverse transcriptase, homologous to the retroviral enzyme, we proposed combining hypomethylating agents with a non-nucleoside inhibitor of this enzyme to reduce the genotoxic effects of 5-azacytidine.

Aim. To evaluate the effect of efavirenz on 5-azacytidine in cultured acute myeloid leukemia cells regarding: сytotoxic activity, expression of long interspersed nuclear elements (LINE1), the level of genetic instability.

Materials and methods. The study was conducted on THP-1 and KG-1 acute myeloid leukemia cell lines. 5-Azacytidine (hypomethylating agents) and efavirenz (reverse transcriptase inhibitor) were used. Cytotoxicity was assessed via the resazurin assay, apoptosis and necrosis rates were measured by flow cytometry, LINE1 expression was quantified using real-time polymerase chain reaction, and DNA damage was evaluated via the comet assay.

Results. Efavirenz did not affect the cytotoxicity of 5-azacytidine. Immunofluorescence staining of LINE1-encoded ORF1 protein and flow cytometry confirmed that efavirenz did not alter LINE1 expression levels. However, comet assay data indicated that combining 5-azacytidine with efavirenz reduced its genotoxic effects.

Conclusion. Our findings demonstrate, for the first time, the potential of a novel acute myeloid leukemia treatment strategy combining hypomethylating agents with reverse transcriptase inhibitors.

About the authors

Kh. M. Magomedova

N.N. Blokhin National Medical Research Center of Oncology, Ministry of Health of Russia

Author for correspondence.
Email: fake@neicon.ru
ORCID iD: 0009-0004-8514-3859

24 Kashirskoe Shosse, Moscow 115522

Russian Federation

I. A. Antonova

N.N. Blokhin National Medical Research Center of Oncology, Ministry of Health of Russia

Email: fake@neicon.ru
ORCID iD: 0009-0004-3482-8954

24 Kashirskoe Shosse, Moscow 115522

Russian Federation

M. S. Strelkov

N.N. Blokhin National Medical Research Center of Oncology, Ministry of Health of Russia; RUDN Universtiy

Email: fake@neicon.ru
ORCID iD: 0009-0007-4778-0584

24 Kashirskoe Shosse, Moscow 115522

6 Miklukho-Maklaya St., Moscow 117198

Russian Federation

V. A. Nurtdinova

N.N. Blokhin National Medical Research Center of Oncology, Ministry of Health of Russia

Email: fake@neicon.ru

24 Kashirskoe Shosse, Moscow 115522

Russian Federation

V. T. Cheshchevik

Polessky State University

Email: fake@neicon.ru
ORCID iD: 0000-0002-1457-8570

23 Dneprovskoy Flotilla St., Pinsk 225710

Belarus

K. I. Kirsanov

N.N. Blokhin National Medical Research Center of Oncology, Ministry of Health of Russia; RUDN Universtiy

Email: fake@neicon.ru
ORCID iD: 0000-0002-8599-6833

24 Kashirskoe Shosse, Moscow 115522

6 Miklukho-Maklaya St., Moscow 117198

Russian Federation

G. A. Belitsky

N.N. Blokhin National Medical Research Center of Oncology, Ministry of Health of Russia

Email: fake@neicon.ru
ORCID iD: 0000-0002-3167-7204

24 Kashirskoe Shosse, Moscow 115522

Russian Federation

M. G. Yakubovskaya

N.N. Blokhin National Medical Research Center of Oncology, Ministry of Health of Russia; RUDN Universtiy

Email: mgyakubovskaya@mail.ru
ORCID iD: 0000-0002-9710-8178

24 Kashirskoe Shosse, Moscow 115522

6 Miklukho-Maklaya St., Moscow 117198

Russian Federation

O. A. Vlasova

N.N. Blokhin National Medical Research Center of Oncology, Ministry of Health of Russia

Email: olya_vlasov@mail.ru
ORCID iD: 0000-0002-1498-849X

24 Kashirskoe Shosse, Moscow 115522

Russian Federation

References

  1. Granatkin M.A., Nikitin E.A., Mikhailov E.S. et al. Combination of azacitidine and venetoclax in first-line therapy in elderly patients with acute myeloid leukemia: first experience. Clin Oncohematol 2022;15(3):282–8. doi: 10.21320/2500-2139-2022-15-3-282-288
  2. Dokshina I.A., Minaeva N.V., Pozdeev N.M. et al. Azacitidine is the preparation of choice in the treatment of myelodysplastic syndromes patients. Gematologiya i transfusiologiya = Russian Journal of Hematology and Transfusiology 2017;62(2):60–4. (In Russ.). doi: 10.18821/0234-5730-2017-62-2-60-64
  3. Jani C.T., Ahmed A., Singh H. et al. 1990–2019: estimates from the global burden of disease study. JCO Glob Oncol 2023;9:e2300229. doi: 10.1200/GO.23.00229
  4. Seymour J.F., Döhner H., Butrym A. et al. Azacitidine improves clinical outcomes in older patients with acute myeloid leukaemia with myelodysplasia-related changes compared with conventional care regimens. BMC Cancer 2017;17(1):852. doi: 10.1186/s12885-017-3803-6
  5. Christman J. 5-azacytidine and 5-aza-2’-deoxycytidine as inhibitors of DNA methylation: mechanistic studies and their implications for cancer therapy. Oncogene 2002;21:5483–95. doi: 10.1038/sj.onc.1205699
  6. Gu X., Tohme R., Tomlinson B. et al. Decitabine- and 5-azacytidine resistance emerges from adaptive responses of the pyrimidine metabolism network. Leukemia 2021;35(4):1023–36. doi: 10.1038/s41375-020-1003-x
  7. Brocks D., Schmidt C.R., Daskalakis M. et al. DNMT and HDAC inhibitors induce cryptic transcription start sites encoded in long terminal repeats. Nat Genet 2017;49(7):1052–60. doi: 10.1038/ng.3889
  8. Daskalakis M., Brocks D., Sheng Y.H. et al. Reactivation of endogenous retroviral elements via treatment with DNMT- and HDAC-inhibitors. Cell Cycle 2018;17(7):811–22. doi: 10.1080/15384101.2018.1442623
  9. Chiappinelli K.B., Strissel P.L., Desrichard A. et al. Inhibiting DNA methylation causes an interferon response in cancer via dsRNA including endogenous retroviruses. Cell 2015;162(5):974–86. doi: 10.1016/j.cell.2015.07.011
  10. Vlasova O.A., Antonova I.A., Magomedova Kh.M. et al. Potential to use of viral reverse transcriptase inhibitors in oncology. Uspekhi molekulyarnoy onkologii = Advances in Molecular Oncology 2024;11(2):8–28. (In Russ.). DOI: 17650/2313-805X-2024-11-2-8-28
  11. Singh A.K., Kumar A., Arora S. et al. Current insights and molecular docking studies of HIV-1 reverse transcriptase inhibitors. Chem Biol Drug Des 2024;103(1):e14372. doi: 10.1111/cbdd.14372
  12. Lander E.S., Linton L.M., Birren B. et al. Initial sequencing and analysis of the human genome. Nature 2001;409(6822):860–921. doi: 10.1038/35057062
  13. Cordaux R., Batzer M.A. The impact of retrotransposons on human genome evolution. Nat Rev Genet 2009;10(10):691–703. doi: 10.1038/nrg2640
  14. Orgel L.E., Crick F.H. Selfish DNA: the ultimate parasite. Nature 1980;284(5757):604–7. doi: 10.1038/284604a0
  15. Ardeljan D., Taylor M.S., Ting D.T., Burns K.H. The human long interspersed element-1 retrotransposon: an emerging biomarker of neoplasia. Clin Chem 2017;63(4):816–22. doi: 10.1373/clinchem.2016.257444
  16. Howard G., Eiges R., Gaudet F. et al. Activation and transposition of endogenous retroviral elements in hypomethylation induced tumors in mice. Oncogene 2008;27(3):404–8. doi: 10.1038/sj.onc.1210631
  17. Payer L.M., Burns K.H. Transposable elements in human genetic disease. Nat Rev Genet 2019;20(12):760–72. doi: 10.1038/s41576-019-0165-8
  18. Scott E.C., Devine S.E. The role of somatic L1 retrotransposition in human cancers. Viruses 2017;9(6):131. doi: 10.3390/v9060131
  19. Burns K.H. Transposable elements in cancer. Nat Rev Cancer 2017;17(7):415–24. doi: 10.1038/nrc.2017.35 20. Rodriguez-Martin B., Alvarez E.G., Baez-Ortega A. et al. Pan cancer analysis of whole genomes identifies driver rearrangements promoted by LINE-1 retrotransposition. Nat Genet 2020;52(3):306–19. doi: 10.1038/s41588-019-0562-0
  20. Toropov S.E., Rudakova A.V., Zakharova N.G. et al. Pharmacoeconomic analysis of firs-line antiretroviral therapy. VICH-infektsiya i immunosupressii = HIV Infection and Immunosuppressive Disorders 2015;7(1):29–39. (In Russ.). doi: 10.22328/2077-9828-2015-7-1-29-39
  21. Costa B., Vale N. Efavirenz: history, development and future. Biomolecules 2022;13(1):88. doi: 10.3390/biom13010088
  22. Hecht M., Harrer T., Körber V. et al. Cytotoxic effect of efavirenz in BxPC-3 pancreatic cancer cells is based on oxidative stress and is synergistic with ionizing radiation. Oncol Lett 2018;15(2):1728–36. doi: 10.3892/ol.2017.7523
  23. Houédé N., Pulido M., Mourey L. et al. A phase II trial evaluating the efficacy and safety of efavirenz in metastatic castration-resistant prostate cancer. Oncologist 2014;19(12):1227–8. doi: 10.1634/theoncologist.2014-0345
  24. Heaton B.J., Jensen R.L., Line J. et al. Exposure of human immune cells, to the antiretrovirals efavirenz and lopinavir, leads to lower glucose uptake and altered bioenergetic cell profiles through interactions with SLC2A1. Biomed Pharmacother 2022;150:112999. doi: 10.1016/j.biopha.2022.112999
  25. Tang H., Yang J., Xu J. et al. The transcription factor PAX5 activates human LINE1 retrotransposons to induce cellular senescence. EMBO Rep 2024;25(8):3263–75. doi: 10.1038/s44319-024-00176-9
  26. Tan Z., Jia X., Ma F. et al. Increased MMAB level in mitochondria as a novel biomarker of hepatotoxicity induced by efavirenz. PLoS One 2017;12(11):e0188366. doi: 10.1371/journal.pone.0188366
  27. Martinez-Arroyo O., Gruevska A., Victor V.M. et al. Mitophagy in human astrocytes treated with the antiretroviral drug efavirenz: lack of evidence or evidence of the lack. Antiviral Res 2019;168:36–50. doi: 10.1016/j.antiviral.2019.04.015
  28. Ganta K.K., Mandal A., Chaubey B. Depolarization of mitochondrial membrane potential is the initial event in non-nucleoside reverse transcriptase inhibitor efavirenz induced cytotoxicity. Cell Biol Toxicol 2017;33(1):69–82. doi: 10.1007/s10565-016-9362-9 Вклад авторов
  29. Apostolova N., Gomez-Sucerquia L.J., Gortat A. et al. Autophagy as a rescue mechanism in efavirenz-induced mitochondrial dysfunction: a lesson from hepatic cells. Autophagy 2011;7(11):1402–4. doi: 10.4161/auto.7.11.17653
  30. Blas-García A., Apostolova N., Ballesteros D. et al. Inhibition of mitochondrial function by efavirenz increases lipid content in hepatic cells. Hepatology 2010;52(1):115–25. doi: 10.1002/hep.23647
  31. Apostolova N., Funes H.A., Blas-Garcia A. et al. Involvement of nitric oxide in the mitochondrial action of efavirenz: a differential effect on neurons and glial cells. J Infect Dis 2015;211(12):1953–8. doi: 10.1093/infdis/jiu825
  32. Imaizumi N., Kwang Lee K., Zhang C., Boelsterli U.A. Mechanisms of cell death pathway activation following drug induced inhibition of mitochondrial complex I. Redox Biol 2015;4:279–88. doi: 10.1016/j.redox.2015.01.005
  33. Li M., Sopeyin A., Paintsil E. Combination of tenofovir and emtricitabine with efavirenz does not moderate inhibitory effect of efavirenz on mitochondrial function and cholesterol biosynthesis in human T lymphoblastoid cell line. Antimicrob Agents Chemother 2018;62(9):e00691–18. doi: 10.1128/AAC.00691-18
  34. Khan R., Schmidt-Mende J., Karimi M. et al. Hypomethylation and apoptosis in 5-azacytidine-treated myeloid cells. Exp Hematol 2008;36(2):149–57. doi: 10.1016/j.exphem.2007.10.002
  35. Kim D.Y., Cheong H.T., Ra C.S. et al. Effect of 5-azacytidine (5-aza) on UCP2 expression in human liver and colon cancer cells. Int J Med Sci 2021;18(10):2176–86. doi: 10.7150/ijms.56564
  36. Derdak Z., Mark N.M., Beldi G. et al. The mitochondrial uncoupling protein-2 promotes chemoresistance in cancer cells. Cancer Res 2008;68(8):2813–9. doi: 10.1158/0008-5472.CAN-08-0053
  37. Dalla Pozza E., Fiorini C., Dando I. et al. Role of mitochondrial uncoupling protein 2 in cancer cell resistance to gemcitabine. Biochim Biophys Acta 2012;1823(10):1856–63. doi: 10.1016/j.bbamcr.2012.06.007
  38. Boada M., Grille S. Myelodysplastic syndrome in patients living with HIV infection. Hematol Transfus Cell Ther 2023;45(1):119–23. doi: 10.1016/j.htct.2021.06.006
  39. Forghieri F., Nasillo V., Bettelli F. et al. Acute myeloid leukemia in patients living with HIV infection: several questions, fewer answers. Int J Mol Sci 2020;21(3):1081. doi: 10.3390/ijms21031081

Supplementary files

Supplementary Files
Action
1. JATS XML
Download

Copyright (c)


СМИ зарегистрировано Федеральной службой по надзору в сфере связи, информационных технологий и массовых коммуникаций (Роскомнадзор).
Регистрационный номер и дата принятия решения о регистрации СМИ: серия ПИ № ФС 77 - 57560 от  08.04.2014.