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Mechanisms of induction of apoptosis by anthraquinone anticancer drugs aclarubicin and mitoxantrone in comparison with doxorubicin: Relation to drug cytotoxicity and caspase-3 activation

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Abstract

We examined molecular events and morphological features associated with apoptosis induced by anthraquinone anticancer drugs aclarubicin, mitoxantrone and doxorubicin in two spontaneously immortalized cell lines (NIH 3T3 and B14) in relation to cytotoxicity of these drugs. The investigated cells showed similar sensitivity to aclarubicin but different sensitivity to doxorubicin and mitoxantrone: mitoxantrone was the most cytotoxic drug in both cell lines. All three drugs triggered both apoptosis and necrosis but none of these processes was positively correlated with their cytotoxicity. Apoptosis was the prevalent form of cell kill by aclarubicin, while doxorubicin and mitoxantrone induced mainly the necrotic mode of cell death. The extent and the timing of apoptosis were strongly dependent on the cell line, the type of the drug and its dose, and were mediated by caspase-3 activation. A significant increase in caspase-3 activity and the percentage of apoptotic cells, oligonucleosomal DNA fragmentation, chromatin condensation and formation of apoptotic bodies was observed predominantly in B14 cells. NIH 3T3 cells showed lesser changes and a lack of DNA fragmentation. Aclarubicin was the fastest acting drug, inducing DNA fragmentation 12 h earlier than doxorubicin, and 24 h earlier than mitoxantrone. Caspase-3 inhibitor Ac-DEVD-CHO did not show any significant effect on drug cytotoxicity and DNA nucleosomal fragmentation.

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References

  1. Murphy GP, Lawrence W. Jr, Lenhard RE, eds. American society textbook of clinical oncology, 2nd ed. Atlanta, GA: American Cancer Society, 1995.

    Google Scholar 

  2. Doroshow JH. Anthracyclines and anthracenediones. In: Chabner BA, Longo DL, eds. Cancer chemotherapy and biotherapy, 2nd ed., New York: Lippincott-Raven Publishers, Philadelphia, 1996: 409–434.

    Google Scholar 

  3. Faulds D, Balfour JA, Chrisp P, Langtry HD. Mitoxantrone, a review of its pharmacodynamic and pharmacokinetic properties, and therapeutic potential in the chemotherapy of cancer. Drugs 1991; 3: 400–449.

    Google Scholar 

  4. Nicholson DW, Thornberry NA. Caspases:Killer proteases. Trends Biochem Sci 1997; 22: 299–306.

    Article  CAS  PubMed  Google Scholar 

  5. Leist M, Jäättelä M. Four deaths and a funeral: from caspases to alternative mechanisms. Nat Rev Mol Cell Biol 2001; 2: 589–598.

    Article  CAS  PubMed  Google Scholar 

  6. Woo M, Hakem R, Soengas MS, et al. Essential contribution of caspase 3/CPP32 to apoptosis and its associated nuclear changes. Genes Dev 1998; 12: 806–819.

    CAS  PubMed  Google Scholar 

  7. Jänicke RU, Sprengart ML, Wati MR, Porter AG. Caspase-3 is required for DNA fragmentation and morphological changes associated with apoptosis. J Biol Chem 1998; 273: 9357–9360.

    PubMed  Google Scholar 

  8. Susin SA, Douglas E, Ravagnan L, et al. Two distinct pathways leading to nuclear apoptosis. J Exp Med 2000; 192: 571–579.

    Article  CAS  PubMed  Google Scholar 

  9. Joza N, Susin SA, Douglas E, et al. Essential role of the mitochondrial apoptosis-inducing factor in programmed cell death. Nature 2001; 410: 549–554.

    Article  CAS  PubMed  Google Scholar 

  10. Li LY, Luo X, Wang X. Endonuclease G is an apoptotic Dnase when released from mitochondria. Nature 2001; 412: 95–99.

    CAS  PubMed  Google Scholar 

  11. Nakano H, Shinohara K. Time sequence analysis of caspase-3 independent programmed cell death and apoptosis in X-irradiated human leukaemic MOLT-4 cells. Cell Tissue Res 2001; 310: 305–311.

    Google Scholar 

  12. Coelho D, Holl V, Weltin D, et al. Caspase-3-like activity determines the type of cell death following ionising radiationin MOLT-4 human leukaemia cells. Br J Cancer 2000; 83: 642–649.

    Article  CAS  PubMed  Google Scholar 

  13. Blagosklonny MV. Cell death beyond apoptosis. Leukemia 2000; 14: 1502–1508.

    CAS  PubMed  Google Scholar 

  14. Schulze-Osthoff K, Walczak H, Droge W, Krammer PH. Cell nucleus and DNA fragmentation are not required for apoptosis. J Cell Biol 1994; 127: 15–20.

    Article  CAS  PubMed  Google Scholar 

  15. Johnson DE. Programmed cell death regulation: Basic mechanisms and therapeutic opportunities. Leukemia 2000; 14: 1340–1344.

    CAS  PubMed  Google Scholar 

  16. Skladanowski A, Konopa J. Adriamycin and daunomycin induce programmed cell death (apoptosis) in tumor cells. Biochem Pharmacol 1993; 46: 375–382.

    Article  CAS  PubMed  Google Scholar 

  17. Ling Y, Priebe W, Perez-Soler R. Apotosis induced by anthracycline antibiotics in P388 parent and multidrug-resistant cells. Cancer Res 1993; 53: 1845–1852.

    CAS  PubMed  Google Scholar 

  18. Han J, Dionne CA, Kedersha NL, Goldmacher VS. p53 status affects the rate of the onset but not the overall extent of doxorubicin-induced death in Rat-1 fibroblasts constitutively expressing c-myc. Cancer Res 199757: 176–182.

    CAS  PubMed  Google Scholar 

  19. Thakkar NS, Potten CS. Abrogation of adriamycin toxicity in vivo by cycloheximide. Biochem Pharmacol 1992; 43: 1683–1691.

    Article  CAS  PubMed  Google Scholar 

  20. Fornari Jr, FA, Jarvis WD, Grant S, Orr MS, Randolph JK, White FKH, Gewitz DA. Growth arrest and non-apoptotic cell death associated with the suppression of c-myc expression in MCF-7 breast tumor cells following acute exposure to doxorubicin. Biochem Pharmacol 1996; 51: 931–940.

    Article  CAS  PubMed  Google Scholar 

  21. Gruber BM, Anuszewska EL, Skierski JS. Activation of programmed cell death (apoptosis) by adriamycin in human neoplastic cells. Mutation Res 2001; 484: 87–93.

    CAS  PubMed  Google Scholar 

  22. Dartsch DC, Schaefer A, Boldt S, Kolch W, Marquardt H. Comparison of anthracycline-induced death of human leukaemia cells: Programmed cells death versus necrosis. Apoptosis 2000; 7: 537–548.

    Google Scholar 

  23. Mossman T. Rapid colorimetric assay for cellular growth and survival: Application to proliferation and cytotoxicity assays. J Immunol Methods 1983; 65: 55–63.

    Google Scholar 

  24. Ormerod M, Collins M, Rodriguez-Jarduchy G, Robertson D. Apoptosis in interleukin-3-dependent haemopoietic cells. Quantification by two flow cytometric methods. J Immunol Methods 1992; 153: 57–65.

    Article  CAS  PubMed  Google Scholar 

  25. Darzynkiewicz Z, Bruno S, Del Bino G, et al. Features of apoptotic cells measured by flow cytometry. Cytometry 1992; 13: 795–808.

    Article  CAS  PubMed  Google Scholar 

  26. Hotz MA, Gong J, Traganos F, Darzynkiewicz Z. Flow cytometric detection of apoptosis: Comparison of the assay oof in situ DNA degradation and chromatin changes. Cytometry 1994; 15: 237–244.

    Article  CAS  PubMed  Google Scholar 

  27. Elstein KH, Zucker RM. Comparison of cellular and nuclear flow cytometric techniques for discriminating apoptotic subpopulations. Exp Cell Res 1994; 211: 322–331.

    Article  CAS  PubMed  Google Scholar 

  28. Gasiorowski K, Brokos B, Kulma A, Ogorzalek A, Skorkowska K. A comparison of the methods applied to detect apoptosis in genotoxically-damaged lymphocytes cultured in the presence of four antimutagens. Cell Biol Mol Lett 2001; 6: 141–159.

    CAS  Google Scholar 

  29. Nicholson DW, Ali A, Thornberry NA, et al. Identification and inhibition of the ICE/CED-3 protease necessary for mammalian apoptosis. Nature 1995; 376: 37–43.

    Article  CAS  PubMed  Google Scholar 

  30. Gonzales VM, Fuertes MA, Alonso C, Perez JM. Is cisplatin-induced cell death always produced by apoptosis? Mol Pharmacol 2001; 59: 657–663.

    Google Scholar 

  31. Kiaris H, Schally AV. Apoptosis versus necrosis: Which should be the aim of cancer therapy? Proc Soc Exp Biol Med 1999; 221: 87–88.

    Article  CAS  PubMed  Google Scholar 

  32. Fukushima T, Kawai Y, Nakayama T, et al. Superior cytotoxic potency of mitoxantrone in interaction with DNA: Comparison with that of daunorubicin. Oncology Res 1996; 8: 95–100.

    CAS  Google Scholar 

  33. Kaspers GJL, Veerman AJP, Pieters R, et al. In vitro cytotoxicity of mitoxantrone, daunorubicin, and doxorubicin in untreated childhood acute leukaemia. Leukemia 1994; 8: 24–29.

    CAS  PubMed  Google Scholar 

  34. Van der Graaf WTA, de Vries ECE. Mitoxantrone bluebeard for malignancies. Anti-Cancer Drugs 1990; 1: 109–125.

    PubMed  Google Scholar 

  35. Borner C, Money L. Apoptosis without caspases: An inefficient molecular guillotine? Cell Death & Diff 1999; 6: 497–507.

    CAS  Google Scholar 

  36. McCarthy NJ, Whyte MKB, Gilbert CS, Evan GI. Inhibition of Ced-3/ICE-related proteases does not prevent cell death induced by oncogenes, DNA damage, or the bcl-2 homologue bak. J Cell Biol 1997; 136: 215–227.

    Article  CAS  PubMed  Google Scholar 

  37. Nicotera P. Apoptosis and age-related disorders: Role of caspase-dependent and caspase-independent pathways. Toxicol Lett 2002; 127: 189–195.

    Article  CAS  PubMed  Google Scholar 

  38. Hirsch T, Marchetti P, Susin SA, et al. The apoptosis-necrosis paradox. Apoptogenic proteases activated after mitochondrial permeability transition determine the mode of cell death. Oncogene 1997; 15: 1573–1581.

    Article  CAS  PubMed  Google Scholar 

  39. Sarin A, Williams MS, Alexander-Miller MA, Berezofsky JA, Zacharczuk CM, Henkart PA. Target cell lysis by CTL granule exocytosis is independent of ICE/Ced-3 family proteases. Immunity 1997; 6: 209–215.

    Article  CAS  PubMed  Google Scholar 

  40. Vercammen D, Beyaert R, Denecker G, et al. Inhibition of caspases increases the sensitivity of L929 cells to necrosis mediated by tumor necrosis factor. J Exp Med 1998; 187: 1477–1485.

    Article  CAS  PubMed  Google Scholar 

  41. Susin SA, Zamzani N, Castedo M, et al. The central executioner of apoptosis: Multiple connection between protease activation and mitochondria in Fas/APO-1?CD95- and ceramide-induced apoptosis. J Exp Med 1997; 186: 25–37.

    Article  CAS  PubMed  Google Scholar 

  42. Turnbull KJ, Brown BL, Dobson PRM. Caspase-3-like activity is necessary but not sufficient for daunorubicin-induced apoptosis in Jurkat human lymphoblastic leukaemia cells. Leukaemia 1999; 13: 1056–1061.

    CAS  Google Scholar 

  43. Matsura T, Kai M, Fujii Y, Ito H, Yamada K. Hydrogen peroxide-induced apoptosis in HL-60 cells requires caspase-3 activation. Free Rad Res 1999; 30: 73–83.

    CAS  Google Scholar 

  44. Drexler HC. Activation of the cell death program by inhibition of proteasome function. Proc Natl Acad Sci USA 1997; 94: 855–860.

    Article  CAS  PubMed  Google Scholar 

  45. Sakahira H, Enari M, Ohsawa Y, Uchiyama Y, Nagata S. Apoptotic nuclear morphological change without DNA fragmentation. Curr Biol 1999; 9: 543–546.

    Article  CAS  PubMed  Google Scholar 

  46. Zenebergh A, Baurain R, Trouet A. Cellular pharmacokinetics of aclacinomycin A in cultured L1210 cells. Comparison with daunorubicin and doxorubicin, Cancer Chemother. Pharmacol 1982; 8: 243–249.

    CAS  Google Scholar 

  47. Adjei PN, Kaufman SH, Leung W-Y, Mao F, Gores GJ. Selective induction of apoptosis in Hep 3B cells by topoisomerase I inhibitors: Evidence for a protease-dependent pathway that does not activate cysteine protease P32. J Clin Invest 1996; 98: 2588–2596.

    CAS  PubMed  Google Scholar 

  48. Grimm LM, Goldberg AL, Poirier GG, Schwartz LM, Osborne BA. Proteasome play an essential role in thymocyte apoptosis. EMBO J 1996; 15: 3835–3844.

    CAS  PubMed  Google Scholar 

  49. Jaattela M, Wissing D, Kokholm K, Kallunki T, Egeblad M. Hsp70 exerts its anti-apoptotic function downstream of caspase-3-like proteases. EMBO J 1998; 17: 6124–6134.

    CAS  PubMed  Google Scholar 

  50. Leist M, Single B, Castoldi AF, Küchnle S, Nicotera P. Intracellular ATP concentration: A switch deciding between apoptosis and necrosis. J Exp Med 1997; 185: 1481–1486.

    Article  CAS  PubMed  Google Scholar 

  51. Volbracht C, Leist M, Nicotera P. ATP controls neuronal apoptosis triggered by microtubule breakdown or potassium depravation. Mol Med 1999; 5: 477–489.

    CAS  PubMed  Google Scholar 

  52. Volbracht C, Leist M, Nicotera P, Kolb SS, Nicotera P. Apoptosis in caspase-inhibited neurons. Mol Med 2001; 7: 36–48.

    CAS  PubMed  Google Scholar 

  53. Ormerod MG, O'Neill CF, Robertson D, Harrap KR. Cisplatin induces apoptosis in a human ovarian carcinoma cell line without concomitant internucleosomal degradation of DNA. Exp Cell Res 1994; 211: 231–237.

    Article  CAS  PubMed  Google Scholar 

  54. Bortner CD, Olenberg NB, Cidlowski JA. The role of DNA fragmentation in apoptosis. Trends Cell Biol 1995; 5: 21–26.

    Article  CAS  PubMed  Google Scholar 

  55. Kuo M-L, Chou Y-W, Chau Y-P, Meng T-C. Differential induction of apoptosis in oncogene-transformed NIH3T3 cells by methylmethane sulfonate. Biochem Pharmacol 1996; 52: 481–488.

    Article  CAS  PubMed  Google Scholar 

  56. Nguyen B, Gutierrez PL. Mechanism(s) for the metabolism of mitoxantrone: Electron spin resonance and electrochemical studies. Chem Biol Interact 1990; 74: 139–162.

    CAS  PubMed  Google Scholar 

  57. Zeller K-P, Mewes K, Ehninger G, Blanz J. Formation of reactive intermediates by cyt P.450 mediated oxidation of the anticancer drug mitoxantrone. Pure and Appl Chem 1994; 66: 2415–2418.

    CAS  Google Scholar 

  58. Kharasch ED, Novak RF. Inhibition of Adriamycin-stimulated microsomal lipid peroxidation by mitoxantrone and ametantrone. Biochem Biophys Res Commun 1982; 108: 1346–1352.

    Article  CAS  PubMed  Google Scholar 

  59. Morceau F, Aries A, Lachlil R, Devy L. Evidence for distinct regulation processes in the aclacinomycin- and doxorubicin-mediated differentiation of human erythroleukemic cells. Biochem Pharmacol 1996; 51: 839–845.

    Article  CAS  PubMed  Google Scholar 

  60. Gewirtz DA. A critical evaluation of the mechanisms of action proposed for the antitumor effects of the anthracycline antibiotics adriamycin and daunomycin. Biochem Pharmacol 1999; 57: 727–741.

    Article  CAS  PubMed  Google Scholar 

  61. Kapuscinski J, Darzynkiewicz Z. Relationship between the pharmacological activity of antitumor drugs ametantrone and mitoxantrone (novantrone) and their ability to condense nucleic acids. Proc Nat Acad Sci USA 1986; 83: 6302–6306.

    CAS  PubMed  Google Scholar 

  62. Booser DJ, Hortobagyi GN. Anthracycline antibiotics in cancer therapy. Drugs 1994; 47: 223–258.

    CAS  PubMed  Google Scholar 

  63. Crespi MO, Ivanier SE, Genovese J, Baldi A. Mitoxantrone affects topoisomerase activities in human breast cancer cells. Biochem Biophys Res Commun 1986; 136: 521–528.

    Article  CAS  PubMed  Google Scholar 

  64. Nitiss JL, Pourquier P, Pommier Y. Aclacinomycin A stabilizes topoisomerase I covalent complex. Cancer Res 1997; 57: 4564–4569.

    CAS  PubMed  Google Scholar 

  65. Reszka KJ, McCormick ML, Britigan BE. Peroxidase and nitrite-dependent metabolism of the anthracycline anticancer agents daunorubicin and doxorubicin. Biochemistry 2001; 40: 15349–15361.

    Article  CAS  PubMed  Google Scholar 

  66. Doroshow JH. Role of hydrogen peroxide and hydroxy radical formation in the killing of Ehrlich tumor cells by anticancer quinines. Proc Natl Acad Sci USA 1986; 83: 4514–4518.

    CAS  PubMed  Google Scholar 

  67. Kolodziejczyk P, Reszka K, Lown J. Enzymatic oxidative activation and transformation of the antitumor agent mitoxantrone. Free Radic Biol Med 1988; 5: 13–25.

    Article  CAS  PubMed  Google Scholar 

  68. Schaefer A, Dahle M, Radenz G, Steinheider G, Marquardt H. Structure-activity relationship between anthracycline-induced differentiation and inhibition of glycoprotein synthesis in friend erythroleukemia cells. Leukemia 1991; 5: 95–100.

    CAS  PubMed  Google Scholar 

  69. Ferraro C, Quemeneur L, Prigent A-F, Taverne C, Revillard J-P, Bonnefoy-Berard N. Anthracyclines trigger apoptosis of both G0/G1 and cycling peripheral blood lymphocytes and induce massive deletion of mature T and B cells. Cancer Res 2000; 60: 1901–1907.

    CAS  PubMed  Google Scholar 

  70. Serafino A, Sinibaldi-Vallebona P, Pierimarchi P, et al. Induction of apoptosis in neoplastic cells by anthracycline antitumor drugs: Nuclear and cytoplasmic triggering? Anticancer Res 1999; 19: 1909–1918.

    CAS  PubMed  Google Scholar 

  71. Orlandi L, Bertoli G, Abolafio G, Daidone MG, Zaffaroni N. Effects of liposome-entrapped annamycin in human breast cancer cells. Interference with cell cycle progression and induction of apoptosis. J Cell Biochem 2001; 81: 9–22.

    Article  CAS  PubMed  Google Scholar 

  72. Maestre N, Tritton TR, Laurent G, Jaffrezou J-P. Cell surface-directed interaction of anthracyclines leads to cytotoxixity and nuclear factor kB activation but not apoptosis signaling. Cancer Res 2001; 61: 2558–2561.

    CAS  PubMed  Google Scholar 

  73. Quillet-Mary A, Mansat V, Duchayne E, et al. Daunorubicin-induced internucleosomal DNA fragmentation in acute myeloid cell lines. Leukemia (Baltimore) 1996; 10: 417–425.

    CAS  Google Scholar 

  74. Richardson DS, Johnson SA. Anthracyclines in haematology: Preclinical studies, toxicity and delivery systems. Blood Rev 1997; 11: 201–223.

    Article  CAS  PubMed  Google Scholar 

  75. Liu FT, Kelsey SM, Newland AC, Jia L. Liposomal encapsulation diminishes daunorubicin-induced generation of reactive oxygen species, depletion of ATP and necrotic cell death in human leukaemic cells. Br J Haematol 2002; 117: 333–342.

    Article  CAS  PubMed  Google Scholar 

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Authors and Affiliations

  1. Department of Thermobiology, University of Łódź, Łódź, Poland

    A. Koceva-Chyła, M. Jedrzejczak, K. Kania & Z. Jóźwiak

  2. Flow Cytometry Laboratory, National Institute of Public Health, Warsaw, Poland

    J. Skierski

  3. Department of Thermobiology, University of Lodz, Banacha 12/16, 90-237, Lodz, Poland

    A. Koceva-Chyła

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  1. A. Koceva-Chyła
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  2. M. Jedrzejczak
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Correspondence to A. Koceva-Chyła.

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Koceva-Chyła, A., Jedrzejczak, M., Skierski, J. et al. Mechanisms of induction of apoptosis by anthraquinone anticancer drugs aclarubicin and mitoxantrone in comparison with doxorubicin: Relation to drug cytotoxicity and caspase-3 activation. Apoptosis 10, 1497–1514 (2005). https://doi.org/10.1007/s10495-005-1540-9

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