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Volume 25, Issue 12
  • ISSN: 1389-4501
  • E-ISSN: 1873-5592

Abstract

Background

Nitazoxanide not only exhibits a broad spectrum of activities against various pathogens infecting animals and humans but also induces cellular autophagy. Currently, the pattern of action and subcellular targets of nitazoxanide-induced cellular autophagy are still unclear.

Methods

To identify potential targets of nitazoxanide in mammalian cells, we developed an affinity chromatography system using tizoxanide, a deacetyl derivative of nitazoxanide, as a ligand. Affinity chromatography was performed using VERO cell extracts on tizoxanide-biotin, and the isolated binding proteins were identified by mass spectrometry. Candidate target proteins obtained using affinity chromatography were co-analysed with the drug affinity response target stability method. Fluorescent probes obtained by coupling rhodamine B to nitazoxanide were used for intracellular localisation of the binding targets. Solvent-induced protein precipitation profiling and thermal proteome profiling were used to further validate the binding proteins.

Results

The joint analysis of the drug affinity response target stability method and affinity chromatography resulted in the screening of six possible candidate target proteins. Fluorescent probes localised the nitazoxanide-binding protein around the nuclear membrane. Molecular docking revealed that the binding proteins mainly formed hydrogen bonds with the nitro group of nitazoxanide. Solvent-induced protein precipitation profiling and thermal proteome profiling further validated SEC61A, PSMD12, and PRKAG1 as potential target proteins of nitazoxanide.

Conclusion

The data supports the idea that nitazoxanide is a multifunctional compound with multiple targets.

© 2024 Bentham Science Publishers
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2024-11-22

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References

  1. Castillo-SalazarM. Sánchez-MuñozF. Springall del VillarR. Nitazoxanide exerts immunomodulatory effects on peripheral blood mononuclear cells from type 2 diabetes patients.Biomolecules20211112181710.3390/biom1112181734944461
     [Google Scholar]
  2. RossignolJ.F. MaisonneuveH. Nitazoxanide in the treatment of Taenia saginata and Hymenolepis nana infections.Am. J. Trop. Med. Hyg.198433351151210.4269/ajtmh.1984.33.5116731683
     [Google Scholar]
  3. RossignolJ.F. Nitazoxanide: A first-in-class broad-spectrum antiviral agent.Antiviral Res.20141109410310.1016/j.antiviral.2014.07.01425108173
     [Google Scholar]
  4. EspositoM. MüllerN. HemphillA. Structure-activity relationships from in vitro efficacies of the thiazolide series against the intracellular apicomplexan protozoan Neospora caninum.Int. J. Parasitol.200737218319010.1016/j.ijpara.2006.10.00917141783
     [Google Scholar]
  5. HemphillA. MuellerJ. EspositoM. Nitazoxanide, a broad-spectrum thiazolide anti-infective agent for the treatment of gastrointestinal infections.Expert Opin. Pharmacother.20067795396410.1517/14656566.7.7.95316634717
     [Google Scholar]
  6. BroekhuysenJ. StockisA. LinsR.L. GraeveJ.D. RossignolJ.F. Nitazoxanide: Pharmacokinetics and metabolism in man.Int. J. Clin. Pharmacol. Ther.200038838739410.5414/CPP3838710984012
     [Google Scholar]
  7. La FraziaS. CiucciA. ArnoldiF. Thiazolides, a new class of antiviral agents effective against rotavirus infection, target viral morphogenesis, inhibiting viroplasm formation.J. Virol.20138720110961110610.1128/JVI.01213‑1323926336
     [Google Scholar]
  8. HuangZ. ZhengH. WangY. The modulation of metabolomics and antioxidant stress is involved in the effect of nitazoxanide against influenza A virus in vitro.Acta Virol.2023671161210.3389/av.2023.11612
     [Google Scholar]
  9. ShiZ. WeiJ. DengX. Nitazoxanide inhibits the replication of Japanese encephalitis virus in cultured cells and in a mouse model.Virol. J.20141111010.1186/1743‑422X‑11‑1024456815
     [Google Scholar]
  10. StelitanoD. La FraziaS. AmbrosinoA. Antiviral activity of nitazoxanide against Morbillivirus infections.J. Virus Erad.20239410035310.1016/j.jve.2023.10035338028567
     [Google Scholar]
  11. PiacentiniS. RiccioA. SantopoloS. The FDA-approved drug nitazoxanide is a potent inhibitor of human seasonal coronaviruses acting at postentry level: Effect on the viral spike glycoprotein.Front. Microbiol.202314120695110.3389/fmicb.2023.1206951
     [Google Scholar]
  12. Al-kuraishyH.M. Al-GareebA.I. ElekhnawyE. BatihaG.E.S. Nitazoxanide and COVID-19: A review.Mol. Biol. Rep.20224911111691117610.1007/s11033‑022‑07822‑236094778
     [Google Scholar]
  13. LüZ. LiX. LiK. Nitazoxanide and related thiazolides induce cell death in cancer cells by targeting the 20S proteasome with novel binding modes.Biochem. Pharmacol.202219711491310.1016/j.bcp.2022.11491335032461
     [Google Scholar]
  14. Abd El-FadealNM NafieMS K El-Kherbetawy M, et al Antitumor activity of nitazoxanide against colon cancers: Molecular docking and experimental studies based on Wnt/β-Catenin signaling inhibition.Int. J. Mol. Sci.20212210521310.3390/ijms2210521334069111
     [Google Scholar]
  15. SunH.Y. OuT. HuJ.Y. Nitazoxanide impairs mitophagy flux through ROS-mediated mitophagy initiation and lysosomal dysfunction in bladder cancer.Biochem. Pharmacol.202119011458810.1016/j.bcp.2021.114588
     [Google Scholar]
  16. YeC. WeiM. HuangH. Nitazoxanide inhibits osteosarcoma cells growth and metastasis by suppressing AKT/mTOR and Wnt/β-catenin signaling pathways.Biol. Chem.20224031092994310.1515/hsz‑2022‑014835946850
     [Google Scholar]
  17. ShouJ. KongX. WangX. Tizoxanide inhibits inflammation in LPS-Activated RAW264.7 macrophages via the suppression of NF-κB and MAPK activation.Inflammation20194241336134910.1007/s10753‑019‑00994‑330937840
     [Google Scholar]
  18. AraújoJ.A.A. GomesT.C. LimaV.C.N. SilvaY.B. LinoJunior R.S. VinaudM.C. Oxfendazole nitazoxanide combination in experimental neurocysticercosis - anti-inflammatory and cysticidal effects.Exp. Parasitol.202426210876410.1016/j.exppara.2024.10876438677580
     [Google Scholar]
  19. AmireddyN. DulamV. KaulS. PakkiriR. KalivendiS.V. The mitochondrial uncoupling effects of nitazoxanide enhances cellular autophagy and promotes the clearance of α-synuclein: Potential role of AMPK-JNK pathway.Cell. Signal.202310911076910.1016/j.cellsig.2023.11076937315747
     [Google Scholar]
  20. ShouJ. WangM. ChengX. Tizoxanide induces autophagy by inhibiting PI3K/Akt/mTOR pathway in RAW264.7 macrophage cells.Arch. Pharm. Res.202043225727010.1007/s12272‑019‑01202‑431894502
     [Google Scholar]
  21. ElaidyS.M. HussainM.A. El-KherbetawyM.K. Time-dependent therapeutic roles of nitazoxanide on high-fat diet/streptozotocin-induced diabetes in rats: Effects on hepatic peroxisome proliferator-activated receptor-gamma receptors.Can. J. Physiol. Pharmacol.201896548549710.1139/cjpp‑2017‑053329244961
     [Google Scholar]
  22. LiX. LuJ. XuY. Discovery of nitazoxanide-based derivatives as autophagy activators for the treatment of Alzheimer’s disease.Acta Pharm. Sin. B202010464666610.1016/j.apsb.2019.07.00632322468
     [Google Scholar]
  23. MaM.H. LiF.F. LiW.F. Repurposing nitazoxanide as a novel anti‐atherosclerotic drug based on mitochondrial uncoupling mechanisms.Br. J. Pharmacol.20231801627910.1111/bph.1594936082580
     [Google Scholar]
  24. DubreuilL. HouckeI. MoutonY. RossignolJ.F. In vitro evaluation of activities of nitazoxanide and tizoxanide against anaerobes and aerobic organisms.Antimicrob. Agents Chemother.199640102266227010.1128/AAC.40.10.22668891127
     [Google Scholar]
  25. de CarvalhoL.P.S. DarbyC.M. RheeK.Y. NathanC. Nitazoxanide Disrupts Membrane Potential and Intrabacterial pH Homeostasis of Mycobacterium tuberculosis.ACS Med. Chem. Lett.201121184985410.1021/ml200157f22096616
     [Google Scholar]
  26. ChahalesP. HoffmanP.S. ThanassiD.G. Nitazoxanide inhibits pilus biogenesis by interfering with folding of the usher protein in the outer membrane.Antimicrob. Agents Chemother.20166042028203810.1128/AAC.02221‑1526824945
     [Google Scholar]
  27. PsonisJ.J. ChahalesP. HendersonN.S. RigelN.W. HoffmanP.S. ThanassiD.G. The small molecule nitazoxanide selectively disrupts BAM-mediated folding of the outer membrane usher protein.J. Biol. Chem.201929439143571436910.1074/jbc.RA119.00961631391254
     [Google Scholar]
  28. HoffmanP.S. SissonG. CroxenM.A. Antiparasitic drug nitazoxanide inhibits the pyruvate oxidoreductases of Helicobacter pylori, selected anaerobic bacteria and parasites, and Campylobacter jejuni.Antimicrob. Agents Chemother.200751386887610.1128/AAC.01159‑0617158936
     [Google Scholar]
  29. MüllerJ. WastlingJ. SandersonS. MüllerN. HemphillA. A novel Giardia lamblia nitroreductase, GlNR1, interacts with nitazoxanide and other thiazolides.Antimicrob. Agents Chemother.20075161979198610.1128/AAC.01548‑0617438059
     [Google Scholar]
  30. MüllerJ. NaguleswaranA. MüllerN. HemphillA. Neospora caninum: Functional inhibition of protein disulfide isomerase by the broad-spectrum anti-parasitic drug nitazoxanide and other thiazolides.Exp. Parasitol.20081181808810.1016/j.exppara.2007.06.00817720161
     [Google Scholar]
  31. AhmedT. RahmanS.M.A. AsaduzzamanM. IslamA.B.M.M.K. ChowdhuryA.K.A. Synthesis, in vitro bioassays, and computational study of heteroaryl nitazoxanide analogs.Pharmacol. Res. Perspect.202193e0080010.1002/prp2.80034086411
     [Google Scholar]
  32. MüllerJ. SidlerD. NachburU. WastlingJ. BrunnerT. HemphillA. Thiazolides inhibit growth and induce glutathione‐S‐transferase Pi (GSTP1)‐dependent cell death in human colon cancer cells.Int. J. Cancer200812381797180610.1002/ijc.2375518688861
     [Google Scholar]
  33. StachulskiA.V. TaujanskasJ. PateS.L. Therapeutic potential of nitazoxanide: An appropriate choice for repurposing versus SARS-CoV-2?ACS Infect. Dis.2021761317133110.1021/acsinfecdis.0c0047833352056
     [Google Scholar]
  34. XuJ. XueY. BolingerA.A. Therapeutic potential of salicylamide derivatives for combating viral infections.Med. Res. Rev.202343489793110.1002/med.2194036905090
     [Google Scholar]
  35. HossainM.J. RahmanS.M.A. Repurposing therapeutic agents against SARS-CoV-2 infection: Most promising and neoteric progress.Expert Rev. Anti Infect. Ther.20211981009102710.1080/14787210.2021.186432733355520
     [Google Scholar]
  36. LokhandeA.S. DevarajanP.V. A review on possible mechanistic insights of Nitazoxanide for repurposing in COVID-19.Eur. J. Pharmacol.202189117374810.1016/j.ejphar.2020.17374833227285
     [Google Scholar]
  37. AgrawalM. SarafS. SarafS. In-line treatments and clinical initiatives to fight against COVID-19 outbreak.Respir. Med.202219110619210.1016/j.rmed.2020.10619233199136
     [Google Scholar]
  38. Bello-PerezM. SolaI. NovoaB. KlionskyD.J. FalcoA. Canonical and noncanonical autophagy as potential targets for COVID-19.Cells202097161910.3390/cells9071619
     [Google Scholar]
  39. LomenickB. HaoR. JonaiN. Target identification using drug affinity responsive target stability (DARTS).Proc. Natl. Acad. Sci. USA200910651219842198910.1073/pnas.091004010619995983
     [Google Scholar]
  40. BurleyS.K. BhikadiyaC. BiC. RCSB Protein Data Bank: Powerful new tools for exploring 3D structures of biological macromolecules for basic and applied research and education in fundamental biology, biomedicine, biotechnology, bioengineering and energy sciences.Nucleic Acids Res.202149D1D437D45110.1093/nar/gkaa103833211854
     [Google Scholar]
  41. ZhangX. WangQ. LiY. Solvent-induced protein precipitation for drug target discovery on the proteomic scale.Anal. Chem.20209211363137110.1021/acs.analchem.9b0453131794197
     [Google Scholar]
  42. ZhangX. WangK. WuS. Highly effective identification of drug targets at the proteome level by pH-dependent protein precipitation.Chem. Sci. (Camb.)20221342124031241810.1039/D2SC03326G36382280
     [Google Scholar]
  43. PanS. ZhangH. WangC. YaoS.C.L. YaoS.Q. Target identification of natural products and bioactive compounds using affinity-based probes.Nat. Prod. Rep.201633561262010.1039/C5NP00101C26580476
     [Google Scholar]
  44. ShiyamaT. FuruyaM. YamazakiA. TeradaT. TanakaA. Design and synthesis of novel hydrophilic spacers for the reduction of nonspecific binding proteins on affinity resins.Bioorg. Med. Chem.200412112831284110.1016/j.bmc.2004.03.05215142543
     [Google Scholar]
  45. TanidaI. UenoT. KominamiE. LC3 conjugation system in mammalian autophagy.Int. J. Biochem. Cell Biol.200436122503251810.1016/j.biocel.2004.05.00915325588
     [Google Scholar]
  46. NayarisseriA. KhandelwalR. TanwarP. Artificial intelligence, big data and machine learning approaches in precision medicine & drug discovery.Curr. Drug Targets202122663165510.2174/18735592MTEzsMDMnz33397265
     [Google Scholar]
  47. LiuW. ZhangZ. ZhangZ.M. HaoP. DingK. LiZ. Integrated phenotypic screening and activity-based protein profiling to reveal potential therapy targets of pancreatic cancer.Chem. Commun. (Camb.)201955111596159910.1039/C8CC08753A30656306
     [Google Scholar]
  48. SharmaV. SinghA. ChauhanS. Role of artificial intelligence in drug discovery and target identification in cancer.Curr. Drug Deliv.20232169062110.2174/156720182166623090509062137670704
     [Google Scholar]
  49. HwangH.Y. KimT.Y. SzászM.A. Profiling the protein targets of unmodified bio‐active molecules with drug affinity responsive target stability and liquid chromatography/tandem mass spectrometry.Proteomics2020209190032510.1002/pmic.20190032531926115
     [Google Scholar]
  50. DrewesG. KnappS. Chemoproteomics and chemical probes for target discovery.Trends Biotechnol.201836121275128610.1016/j.tibtech.2018.06.00830017093
     [Google Scholar]
  51. BantscheffM. ScholtenA. HeckA.J.R. Revealing promiscuous drug-target interactions by chemical proteomics.Drug Discov. Today20091421-221021102910.1016/j.drudis.2009.07.00119596079
     [Google Scholar]
  52. PichlerC.M. KrysiakJ. BreinbauerR. Target identification of covalently binding drugs by activity-based protein profiling (ABPP).Bioorg. Med. Chem.201624153291330310.1016/j.bmc.2016.03.05027085673
     [Google Scholar]
  53. TopçuA.A. KılıçS. ÖzgürE. TürkmenD. DenizliA. Inspirations of biomimetic affinity ligands: A review.ACS Omega2022737328973290710.1021/acsomega.2c0353036157742
     [Google Scholar]
  54. SeoS.Y. CorsonT.W. Small molecule target identification using photo-affinity chromatography. Methods Enzymol201962234737410.1016/bs.mie.2019.02.028]31155061
     [Google Scholar]
  55. RylovaG. OzdianT. VaranasiL. Affinity-based methods in drug-target discovery.Curr. Drug Targets2015161607610.2174/138945011566614112011032325410410
     [Google Scholar]
  56. ChangJ. KimY. KwonH.J. Advances in identification and validation of protein targets of natural products without chemical modification.Nat. Prod. Rep.201633571973010.1039/C5NP00107B26964663
     [Google Scholar]
  57. RixU. GridlingM. Superti-FurgaG. Compound immobilization and drug-affinity chromatography.Methods Mol. Biol.2012803253810.1007/978‑1‑61779‑364‑6_322065216
     [Google Scholar]
  58. SaxenaC. HiggsR.E. ZhenE. HaleJ.E. Small-molecule affinity chromatography coupled mass spectrometry for drug target deconvolution.Expert Opin. Drug Discov.20094770171410.1517/1746044090300556523489165
     [Google Scholar]
  59. NagatsukaT. UzawaH. SatoK. OhsawaI. SetoY. NishidaY. Glycotechnology for decontamination of biological agents: A model study using ricin and biotin-tagged synthetic glycopolymers.ACS Appl. Mater. Interfaces20124283283710.1021/am201493q22214533
     [Google Scholar]
  60. RybakJ.N. ScheurerS.B. NeriD. EliaG. Purification of biotinylated proteins on streptavidin resin: A protocol for quantitative elution.Proteomics2004482296229910.1002/pmic.20030078015274123
     [Google Scholar]
  61. BishopE. BradshawT.D. Autophagy modulation: A prudent approach in cancer treatment?Cancer Chemother. Pharmacol.201882691392210.1007/s00280‑018‑3669‑630182146
     [Google Scholar]
  62. PaiP.P. MondalS. Applying knowledge of enzyme biochemistry to the prediction of functional sites for aiding drug discovery.Curr. Top. Med. Chem.201717212401242110.2174/156802661766617032915385828359251
     [Google Scholar]
  63. PiazzaI. KochanowskiK. CappellettiV. A map of protein-metabolite interactions reveals principles of chemical communication.Cell20181721-2358372.e2310.1016/j.cell.2017.12.00629307493
     [Google Scholar]
  64. TanieY. TanabeN. KuboyamaT. TohdaC. Extracellular neuroleukin enhances neuroleukin secretion from astrocytes and promotes axonal growth in vitro and in vivo.Front. Pharmacol.20189122810.3389/fphar.2018.0122830459611
     [Google Scholar]
  65. ChristensenK.E. MirzaI.A. BerghuisA.M. MacKenzieR.E. Magnesium and phosphate ions enable NAD binding to methylenetetrahydrofolate dehydrogenase-methenyltetrahydrofolate cyclohydrolase.J. Biol. Chem.200528040343163432310.1074/jbc.M50521020016100107
     [Google Scholar]
  66. ZhangH. ZhuS. ZhouH. LiR. XiaX. XiongH. Identification of MTHFD2 as a prognostic biomarker and ferroptosis regulator in triple-negative breast cancer.Front. Oncol.202313109835710.3389/fonc.2023.109835736726381
     [Google Scholar]
  67. LeGrosH.L.Jr HalimA.B. GellerA.M. KotbM. Cloning, expression, and functional characterization of the beta regulatory subunit of human methionine adenosyltransferase (MAT II).J. Biol. Chem.200027542359236610.1074/jbc.275.4.235910644686
     [Google Scholar]
  68. MonsonM.S. SettlageR.E. McMahonK.W. Response of the hepatic transcriptome to aflatoxin B1 in domestic turkey (Meleagris gallopavo).PLoS One201496e10093010.1371/journal.pone.010093024979717
     [Google Scholar]
  69. WangZ. LiZ. XuH. PSMD12 promotes glioma progression by upregulating the expression of Nrf2.Ann. Transl. Med.20219870010.21037/atm‑21‑148133987398
     [Google Scholar]
  70. HuiX. CaoL. XuT. PSMD12-mediated M1 ubiquitination of influenza A Virus at K102 regulates viral replication.J. Virol.20229615e00786e2210.1128/jvi.00786‑2235861516
     [Google Scholar]
  71. JuricaM.S. LickliderL.J. GygiS.P. GrigorieffN. MooreM.J. Purification and characterization of native spliceosomes suitable for three-dimensional structural analysis.RNA20028442643910.1017/S135583820202108811991638
     [Google Scholar]
  72. GeuensT. BouhyD. TimmermanV. The hnRNP family: Insights into their role in health and disease.Hum. Genet.2016135885186710.1007/s00439‑016‑1683‑527215579
     [Google Scholar]
  73. HaßdenteufelS. JohnsonN. PatonA.W. PatonJ.C. HighS. ZimmermannR. Chaperone-mediated sec61 channel Gating during ER import of small precursor proteins overcomes Sec61 inhibitor-reinforced energy barrier.Cell Rep.20182351373138610.1016/j.celrep.2018.03.12229719251
     [Google Scholar]
  74. PandaD. RoseP.P. HannaS.L. Genome-wide RNAi screen identifies SEC61A and VCP as conserved regulators of Sindbis virus entry.Cell Rep.2013561737174810.1016/j.celrep.2013.11.02824332855
     [Google Scholar]
  75. OakhillJ.S. SteelR. ChenZ.P. AMPK is a direct adenylate charge-regulated protein kinase.Science201133260361433143510.1126/science.120009421680840
     [Google Scholar]
  76. LöfflerA.S. AlersS. DieterleA.M. Ulk1-mediated phosphorylation of AMPK constitutes a negative regulatory feedback loop.Autophagy20117769670610.4161/auto.7.7.1545121460634
     [Google Scholar]
  77. MüllerJ. HemphillA. Identification of a host cell target for the thiazolide class of broad-spectrum anti-parasitic drugs.Exp. Parasitol.2011128214515010.1016/j.exppara.2011.02.00721335006
     [Google Scholar]
  78. SunC.P. ZhouJ.J. YuZ.L. Kurarinone alleviated Parkinson’s disease viastabilization of epoxyeicosatrienoic acids in animal model.Proc. Natl. Acad. Sci. USA20221199e211881811910.1073/pnas.211881811935217618
     [Google Scholar]
  79. FrankenH. MathiesonT. ChildsD. Thermal proteome profiling for unbiased identification of direct and indirect drug targets using multiplexed quantitative mass spectrometry.Nat. Protoc.201510101567159310.1038/nprot.2015.10126379230
     [Google Scholar]
  80. TuY. TanL. TaoH. LiY. LiuH. CETSA and thermal proteome profiling strategies for target identification and drug discovery of natural products.Phytomedicine202311615486210.1016/j.phymed.2023.15486237216761
     [Google Scholar]
  81. Martinez MolinaD. NordlundP. The cellular thermal shift assay: A novel biophysical assay for in situ drug target engagement and mechanistic biomarker studies.Annu. Rev. Pharmacol. Toxicol.201656114116110.1146/annurev‑pharmtox‑010715‑10371526566155
     [Google Scholar]
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Joint Screening and Identification of Potential Targets of Nitazoxanide by Affinity Chromatography and Label-Free Techniques
Curr Drug Targets 25, 819 (2024); https://doi.org/10.2174/0113894501297697240805103744
/content/journals/cdt/10.2174/0113894501297697240805103744
/content/journals/cdt/10.2174/0113894501297697240805103744
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  • Article Type:     Research Article
Keyword(s): affinity chromatography; Alzheimer's disease; atherosclerosis; autophagy; drug target; nitazoxanide

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