Amino Acids

, Volume 44, Issue 5, pp 1279–1292 | Cite as

Quinacrine reactivity with prion proteins and prion-derived peptides

  • Zbigniew Zawada
  • Martin Šafařík
  • Eva Dvořáková
  • Olga Janoušková
  • Anna Březinová
  • Ivan Stibor
  • Karel Holada
  • Petr Bouř
  • Jan Hlaváček
  • Jaroslav Šebestík
Original Article


Quinacrine is a drug that is known to heal neuronal cell culture infected with prions, which are the causative agents of neurodegenerative diseases called transmissible spongiform encephalopathies. However, the drug fails when it is applied in vivo. In this work, we analyzed the reason for this failure. The drug was suggested to “covalently” modify the prion protein via an acridinyl exchange reaction. To investigate this hypothesis more closely, the acridine moiety of quinacrine was covalently attached to the thiol groups of cysteines belonging to prion-derived peptides and to the full-length prion protein. The labeled compounds were conveniently monitored by fluorescence and absorption spectroscopy in the ultraviolet and visible spectral regions. The acridine moiety demonstrated characteristic UV–vis spectrum, depending on the substituent at the C-9 position of the acridine ring. These results confirm that quinacrine almost exclusively reacts with the thiol groups present in proteins and peptides. The chemical reaction alters the prion properties and increases the concentration of the acridine moiety in the prion protein.


Quinacrine Prion protein and peptide model reactions Solid phase and recombinant synthesis 



This work was supported by the Czech Science Foundation (GA CR) Grant No. 203/07/1517. KH, OJ and ED were supported by the projects of Charles University in Prague: PRVOUK-P24/LF1/3, UNCE 204022 and SVV-2012- 264506. English language revision was made by American Journal Experts,

Conflict of interest

Authors declare that they have no conflict of interest.

Supplementary material

726_2013_1460_MOESM1_ESM.doc (1.1 mb)
Supplementary material 1 (DOC 1149 kb)


  1. Bolognesi ML, Cavalli A, Valgimigli L, Bartolini M, Rosini M, Andrisano V, Recanatini M, Melchiorre C (2007) Multi-target-directed drug design strategy: from a dual binding site Acetylcholinesterase inhibitor to a trifunctional compound against Alzheimer’s disease. J Med Chem 50:6446–6449PubMedCrossRefGoogle Scholar
  2. Burnett JC, Schmidt JJ, Stafford RG, Panchal RG, Nguyen TL, Hermone AR, Vennerstrom JL, McGrath CF, Lane DJ, Sausville EA, Zaharevitz DW, Gussio R, Bavari S (2003) Novel small molecule inhibitors of Botulinum neurotoxin. A metalloprotease activity. Biochem Biophys Res Commun 310:84–93PubMedCrossRefGoogle Scholar
  3. Chang CD, Waki M, Ahmad M, Meienhofer J, Lundell EO, Haug JD (1980) Preparation and properties of N-9-fluorenylmethyloxycarbonylamino acids bearing tert-butyl side chain protection. Int J Pept Prot Res 15:59–66CrossRefGoogle Scholar
  4. Claude S, Lehn JM, Vigneron JP (1989) Bicyclo-bis-intercalands: synthesis of triply bridged bis-intercalands based on acridine subunits. Tetrahedron Lett 30:941–944CrossRefGoogle Scholar
  5. Cobb NJ, Surewicz WK (2009) Prion diseases and their biochemical mechanisms. Biochemistry 48:2574–2585PubMedCrossRefGoogle Scholar
  6. Collinge J, Gorham M, Hudson F, Kennedy A, Keogh G, Pal S, Rossor M, Rudge P, Siddique D, Spyer M, Thomas D, Walker S, Webb T, Wroe S, Darbyshire J (2009) Safety and efficacy of quinacrine in human prion disease (PRION-1 study): a patient-preference trial. Lancet Neurol 8:334–344PubMedCrossRefGoogle Scholar
  7. Collins SJ, Lewis V, Brazier M, Hill AF, Fletcher A, Masters CL (2002) Quinacrine does not prolong survival in a murine Creutzfeldt-Jakob Disease model. Ann Neurol 52:503–506PubMedCrossRefGoogle Scholar
  8. Demeunynck M, Charmantray F, Martelli A (2001) Interest of acridine derivatives in the anticancer chemotherapy. Curr Pharm Des 7:1703–1724PubMedCrossRefGoogle Scholar
  9. Denny WA (2003) Acridine-4-carboxamides and the concept of minimal DNA intercalators. In: Demeunynck M, Bailly C, Wilson WD (eds) In small molecule DNA and RNA binders. Wiley-VCH Verlag GmbH & Co, Weinheim, pp 482–502Google Scholar
  10. Dollinger S, Lober S, Klingenstein R, Korth C, Gmeiner PA (2006) Chimeric ligand approach leading to potent antiprion active acridine derivatives: design, synthesis, and biological investigations. J Med Chem 49:6591–6595PubMedCrossRefGoogle Scholar
  11. Dondi F (1982) Approximation properties of the Edgeworth-Cramer series and determination of peak parameters of chromatographic peaks. Anal Chem 54:473–477CrossRefGoogle Scholar
  12. Dringen R (2000) Metabolism and functions of glutathione in brain. Progress Neurobiol 62:649–671CrossRefGoogle Scholar
  13. Eiter LC, Hall NW, Day CS, Saluta G, Kucera GL, Bierbach U, Gold I (2009) Analogues of a platinum-acridine antitumor agent are only moderately cytotoxic but show potent activity against Mycobacterium tuberculosis. J Med Chem 52:6519–6522PubMedCrossRefGoogle Scholar
  14. Fields GB, Noble RL (1990) Solid phase peptide synthesis utilizing 9-fluorenylmethoxycarbonyl amino acids. Int J Pept Prot Res 35:161–214CrossRefGoogle Scholar
  15. Flavell RR, Huse M, Goger M, Trester-Zedlitz M, Kuriyan J, Muir TW (2002) Efficient semisynthesis of a tetraphosphorylated analogue of the type I TGFβ receptor. Org Lett 4:165–168PubMedCrossRefGoogle Scholar
  16. Franks NP, Abraham MH, Lieb WR (1993) Molecular-organization of liquid n-octanol: an X-ray diffraction analysis. J Pharm Sci 82:466–470PubMedCrossRefGoogle Scholar
  17. Gaydukevich AN, Kazakov GP, Kravchenko AA, Porokhnyak LA, Pinchuk VV, Velikii DL (1987) Synthesis and biological activity of acridinyl-9-thioacetic acids and their derivatives. Pharm Chem J 21:633–636CrossRefGoogle Scholar
  18. Ghaemmaghami S, Ahn M, Lessard P, Giles K, Legname G, DeArmond SJ, Prusiner SB (2009) Continuous Quinacrine treatment results in the formation of drug-resistant prions. PLoS Pathogens 5: art no e1000673Google Scholar
  19. Goodell JR, Ougolkov AV, Hiasa H, Kaur H, Remmel R, Billadeau DD, Ferguson DM (2008) Acridine-based agents with Topoisomerase II activity inhibit pancreatic cancer cell proliferation and induce apoptosis. J Med Chem 51:179–182PubMedCrossRefGoogle Scholar
  20. Guddneppanavar R, Saluta G, Kucera GL, Bierbach U (2006) Synthesis, biological activity and DNA-damage profile of platinum-threading intercalator conjugates designed to target adenine. J Med Chem 49:3204–3214PubMedCrossRefGoogle Scholar
  21. Kaiser E, Colescot RL, Bossinger CD, Cook PI (1970) Color test for detection of free terminal amino groups in the solid-phase synthesis of peptides. Anal Biochem 34:595–598PubMedCrossRefGoogle Scholar
  22. Korth C, May BCH, Cohen FE, Prusiner SB (2001) Acridine and phenothiazine derivatives as pharmacotherapeutics for prion disease. Proc Natl Acad Sci USA 98:9836–9841PubMedCrossRefGoogle Scholar
  23. Krauth-Siegel RL, Bauer H, Schirmer H (2005) Dithiol proteins as guardians of the intracellular redox milieu in parasites: old and new drug targets in trypanosomes and malaria-causing plasmodia. Angew Chem Int Ed 44:690–715CrossRefGoogle Scholar
  24. Krchňák V, Vágner J, Lebl M (1988a) Non-invasive continuous monitoring of solid-phase peptide synthesis by acid-base indicator a. Int J Pept Prot Res 32:415–416CrossRefGoogle Scholar
  25. Krchňák V, Vágner J, Šafař P, Lebl M (1988b) Non-invasive continuous monitoring of solid phase peptide synthesis by acid–base indicator b. Collect Czech Chem Commun 33:2542–2548CrossRefGoogle Scholar
  26. Kunikowski A, Ledochowski A (1981) Reactions at C9 of acridine-derivatives 28. Kinetics of hydrolysis of N-(1-nitroacridyl-9)-d,l-amino acids. Polish J Chem 55:1979–1984Google Scholar
  27. Lehn JM (1993) Supramolecular chemistry. Science 260:1762–1763PubMedCrossRefGoogle Scholar
  28. Lewis JS (1949) The N-acylation of N-(4-methoxyphenyl)-4-chloro-anthranilic acid. J Org Chem 14:285–288CrossRefGoogle Scholar
  29. Madden HH (1978) Comments on the Savitzky-Golay convolution method for least-squares fit smoothing and differentiation of digital data. Anal Chem 50:1383–1386CrossRefGoogle Scholar
  30. May BCH, Fafarman AT, Hong SB, Rogers M, Deady LWn Prusiner SB, Cohen FE (2003) Potent inhibition of scrapie prion replication in cultured cells by bis-acridines. Proc Natl Acad Sci USA 100:3416–3421Google Scholar
  31. Meienhofer J, Waki M, Heimer EP, Lambros TJ, Makofske RC, Chang CD (1979) Solid phase synthesis without repetitive acidolysis. Preparation of leucyl-alanyl-glycyl-valine using 9-fluorenylmethyloxycarbonylamino acids. Int J Pept Prot Res 13:35–42CrossRefGoogle Scholar
  32. Nadal RC, Abdelraheim SR, Brazier MW, Rigby SEJ, Brown DR, Vile JH (2007) Prion protein does not redox-silence Cu2+, but is a sacrificial quencher of hydroxyl radicals. Free Radical Biol Med 42:79–89CrossRefGoogle Scholar
  33. Orzáez M, Mondragon L, Garcia-Jareno A, Mosulen S, Pineda-Lucena A, Perez-Paya E (2009) Deciphering the antitumoral activity of Quinacrine: binding to and inhibition of Bcl-xL. Bioorg Med Chem Lett 19:1592–1595PubMedCrossRefGoogle Scholar
  34. Paul A, Ladame S (2009) 9-Amino acridines undergo reversible amine exchange reactions in water: implications on their mechanism of action in vivo. Org Lett 11:4894–4897PubMedCrossRefGoogle Scholar
  35. Pavlíček A, Bednárová L, Holada K (2007) Production, purification and oxidative folding of the mouse recombinant prion protein. Folia Microbiol 52:391–397Google Scholar
  36. Phuan PW, Zorn JA, Safar J, Giles K, Prusiner SB, Cohen FE, May BCH (2007) Discriminating between cellular and misfolded prion protein by using affinity to 9-aminoacridine compounds. J General Virol 88:1392–1401CrossRefGoogle Scholar
  37. Rodriguez-Franco MI, Fernandez-Bachiller MI, Perez C, Hernandez-Ledesma B, Bartolome B (2006) Novel tacrine-melatonin hybrids as dual-acting drugs for Alzheimer disease, with improved acetylcholinesterase inhibitory and antioxidant properties. J Med Chem 49:459–462PubMedCrossRefGoogle Scholar
  38. Ronga L, Palladino P, Costantini S, Facchiano A, Ruvo M, Benedetti E, Ragone R, Rossi F (2007) Conformational diseases and structure-toxicity relationships: lessons from prion-derived peptides. Curr Prot Pept Sci 8:83–90CrossRefGoogle Scholar
  39. Rosini M, Simoni E, Bartolini M, Cavalli A, Ceccarini L, Pascu N, McClymont DW, Tarozzi A, Bolognesi ML, Minarini A, Tumiatti V, Andrisano V, Mellor IR, Melchiorre C (2008) Inhibition of Acetylcholinesterase, beta-Amyloid aggregation, and NMDA receptors in Alzheimer’s disease: a promising direction for the multi-target-directed ligands gold rush. J Med Chem 51:4381–4384PubMedCrossRefGoogle Scholar
  40. Saravanamuthu A, Vickers TJ, Bond CS, Peterson MR, Hunter WN, Fairlamb AH (2004) Two interacting binding sites for quinacrine derivatives in the active site of trypanothione reductase. A template for drug design. J Biol Chem 279:29493–29500PubMedCrossRefGoogle Scholar
  41. Sarin VK, Kent SBH, Tam JP, Merrifield RB (1981) Quantitative monitoring of solid phase peptide synthesis by the ninhydrin reaction. Anal Biochem 117:147–157PubMedCrossRefGoogle Scholar
  42. Savitzky A, Golay MJE (1964) Smoothing and differentiation of data by simplified least squares procedures. Anal Chem 36:1627–1639CrossRefGoogle Scholar
  43. Schantl JG, Tűrk W (1990) Synthesen von 1-(9-acridinyl)-3-hydroxy-harnstoff und 9-acridanon-oxim. Arch Pharm 323:720–726Google Scholar
  44. Šebestík J, Šafařík M, Stibor I, Hlaváček J (2006) Acridin-9-yl exchange: a proposal for the action of some 9-aminoacridine drugs. Biopolymers 84:605–614PubMedCrossRefGoogle Scholar
  45. Šebestík J, Hlaváček J, Stibor I (2007) A role of the 9-aminoacridines and their conjugates in a life science. Curr Prot Pept Sci 8:471–483CrossRefGoogle Scholar
  46. Spilman P, Lessard P, Sattavat M, Bush C, Tousseyn T, Huang EJ, Giles K, Golde T, Das P, Fauq A, Prusiner SB, DeArmond SJ (2008) A γ-secretase inhibitor and quinacrine reduce prions and prevent dendritic degeneration in murine brains. Proc Natl Acad Sci USA 105:10595–10600PubMedCrossRefGoogle Scholar
  47. Steinier J, Termonia Y, Deltour J (1972) Comments on smoothing and differentiation of data by simplified least square procedure. Anal Chem 44:1906–1909PubMedCrossRefGoogle Scholar
  48. Tompa P, Tusnady GE, Friedrich P, Simon I (2002) The role of dimerization in prion replication. Biophys J 82:1711–1718PubMedCrossRefGoogle Scholar
  49. Vogtherr M, Grimme S, Elshorst B, Jacobs DM, Fiebig K, Griesinger C, Zahn R (2003) Antimalarial drug quinacrine binds to C-terminal helix of cellular prion protein. J Med Chem 46:3563–3564PubMedCrossRefGoogle Scholar
  50. Vojkovský T (1995) Detection of secondary amines on solid phase. Pept Res 8:236–237PubMedGoogle Scholar
  51. Wallace DJ (1989) The use of quinacrine (Atabrine) in rheumatic diseases—a reexamination. Semin Arthritis Rheum 18:282–297PubMedCrossRefGoogle Scholar
  52. Weltrowski M, Ledochowski A, Sowinski P (1982) Research on tumor-inhibiting compounds 70. Reactions of 1-nitroacridines with ethanethiol. Polish J Chem 56:77–82Google Scholar
  53. Wild F, Young JM (1965) The reaction of Mepacrine with thiols. J Chem Soc, 7261–7274. doi: 10.1039/JR9650007261
  54. Wille H, Bian W, McDonald M, Kendall A, Colby DW, Bloch L, Ollesch J, Borovinskiy AL, Cohen FE, Prusiner SB, Stubbs G (2009) Natural and synthetic prion structure from X-ray fiber diffraction. Proc Natl Acad Sci USA 106:16990–16995Google Scholar
  55. Wysocka-Skrzela B (1986) Research on tumor inhibiting compounds. Part LXXVI. Reactions of 1-nitro-9-aminoacridine derivatives, new antitumor agents with nucleophiles. Polish J Chem 60:317–318Google Scholar
  56. Yamamoto T, Miller WH (2005) Path integral evaluation of the quantum instanton rate constant for proton transfer in a polar solvent. J Chem Phys 122:art no 044106Google Scholar
  57. Yung L, Huang PY, Lessard P, Legname G, Lin ET, Baldwin M, Prusiner SB, Ryou C, Guglielmo BJ (2004) Pharmacokinetics of Quinacrine in the treatment of prion disease. BMC infectious diseases 4:art no 53Google Scholar
  58. Zawada Z, Šebestík J, Šafařík M, Krejčiříková A, Březinová A, Hlaváček J, Stibor I, Holada K, Bouř P (2010) What could be the role of Quinacrine in Creutzfeldt–Jakob Disease treatment? In: Lebl M, Meldal M, Jensen K, Hoeg-Jensen T (eds) Peptides 2010. Proc 31st Eur Pep Symp. Eur Pep Soc, Copenhagen, pp 84–85Google Scholar
  59. Zawada Z, Šebestík J, Šafařík M, Bouř P (2011) Dependence of the reactivity of acridine on its substituents: a computational and kinetic study. Eur J Org Chem 2011:6989–6997CrossRefGoogle Scholar

Copyright information

© Springer-Verlag Wien 2013

Authors and Affiliations

  • Zbigniew Zawada
    • 1
    • 2
  • Martin Šafařík
    • 1
  • Eva Dvořáková
    • 3
  • Olga Janoušková
    • 3
  • Anna Březinová
    • 1
  • Ivan Stibor
    • 1
    • 4
  • Karel Holada
    • 3
  • Petr Bouř
    • 1
  • Jan Hlaváček
    • 1
  • Jaroslav Šebestík
    • 1
  1. 1.Institute of Organic Chemistry and BiochemistryAcademy of Sciences of the Czech RepublicPrague 6Czech Republic
  2. 2.Institute of Chemical TechnologyPrague 6Czech Republic
  3. 3.First Faculty of MedicineCharles University in PraguePrague 2Czech Republic
  4. 4.Technical University of LiberecLiberec 1Czech Republic

Personalised recommendations