The Role of Topoisomerases and Apoptosis in the Mechanism of Action of Preactivated Compounds

  • Kirpal S. Gulliya
Part of the Medical Intelligence Unit book series (MIU.LANDES)


In this chapter a description of the experimental data obtained to enhance our understanding of the underlying mechanism of action of pMC540, merocil and merodantoin is provided. The first set of experiments was designed to assess the potential involvement of oxygen derived species in the pMC540 mediated cytotoxicity. The rationale for this approach, at the time, was based on the observations that the in vitro cytotoxicity profile of pMC540 paralleled the one observed during conventional photodynamic therapy with native MC540, a process known to involve reactive oxygen species. Therefore, involvement of reactive oxygen species was chosen as a first step towards investigating the potential mechanism of action of pMC540. A brief description of these studies is followed by a discussion of the data related to the involvement of topoisomerases, apoptosis, mitochondrial morphology and function in the observed cytotoxicity mediated by preactivated compounds.


Nuclear Extract Daudi Cell Free Radical Biology Negative Breast Cancer Cell Line Estrogen Receptor Negative Breast Cancer 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.


Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.


  1. 1.
    Halliwell B, Gutteridge JMC. Free Radicals in Biology and Medicine. Oxford: Clarendon, 1985.Google Scholar
  2. 2.
    McCord JM, Fridovich I. Superoxide dismutase. An enzymeic function for erythrocuprein (hemocuprein). J Biol Chem 1969; 244: 6049–55.PubMedGoogle Scholar
  3. 3.
    Babior BM. Oxidants from phagocytes: agents of defense and destruction. Blood 1984; 64 959–66.PubMedGoogle Scholar
  4. 4.
    Comporti M. Lipid peroxidation and cellular damage in toxic liver injury. Lab Invest 1985; 53: 599–623.PubMedGoogle Scholar
  5. 5.
    Gilbert DL, ed. Oxygen and Living Processes. An interdisciplinary approach. New York: Springer-Verlag, 1981.Google Scholar
  6. 6.
    Taeishi T, Yoshimine N, Kucuya F. Serum lipid peroxide assay by a new colorimetric method. Exp Gerontol 1987; 22: 103–11.CrossRefGoogle Scholar
  7. 7.
    Pervaiz S, Harriman A, Gulliya KS. Protein damage by photoproducts of merocyanine 540. Free Rad Biol and Med 1992; 12: 389–96.Google Scholar
  8. 8.
    Davies KJA, Delsignore ME, Lin SW. Protein damage and degradation by oxygen radicals. J Biol Chem 1987; 262: 9902–7.PubMedGoogle Scholar
  9. 9.
    Davies KJA and Delsignore ME. Protein damage and degradation by oxygen radicals. J Biol Chem 1987; 262: 9908–13.PubMedGoogle Scholar
  10. 10.
    Davies KJA, Lin S, and Pacifici RE. Protein damage and degradation by oxygen radicals. J Biol Chem 1987; 262: 9914–20.PubMedGoogle Scholar
  11. 11.
    Gross AJ, Sizer IW. The oxidation of tyramine, tyrosine, and related compounds by peroxidase. J Biol Chem 1959; 234: 1611–14.PubMedGoogle Scholar
  12. 12.
    Prutz WA, Butler J, Land EJ. Phenol coupling initiated by one-electron oxidation of tyrosine units in peptides and histones. Int J Radiat Biol Relat Stud Phy Chem Med 1983; 44: 183–96.CrossRefGoogle Scholar
  13. 13.
    Bohlen P, Stein S, Dairman W, et al. Fluorometric assay of proteins in the nanogram range. Arch Biochem Biophys 1973; 155: 213–20.PubMedCrossRefGoogle Scholar
  14. 14.
    Thor H, Smith MT, Hartzel P et al. The metabolism of menadi-one (2-methyl-1,4-naphtholquinone by isolated hepatocytes. J Biol Chem 1982; 257: 12419–25.PubMedGoogle Scholar
  15. 15.
    Doroshow JH. Role of hydrogen peroxide and hydroxide radical formation in the killing of Ehrlich tumor cells by anticancer quinones. Proc Natl Acad Sci USA, 1985; 83: 4515–18.Google Scholar
  16. 16.
    Arslan P, Di Virgilio F, Beltrame M. Cytosolic Cat+ homeostasis in Ehrlich and Yoshida carcinomas. J Biol Chem 1985; 260: 2719–27.PubMedGoogle Scholar
  17. 17.
    Mello Filho AC, Hoffmann ME, Meneghini R. Cell killing and DNA damage by hydrogen peroxide are mediated by intracellular iron. Biochem J 1984; 218: 273–75.PubMedGoogle Scholar
  18. 18.
    Rosen H, Klebanoff SJ. Role of iron and ethylenediaminetetraacetic acid in the bactericidal activity of a superoxide anion-generating system. Arch Biochem Biophys 1981; 208: 512–19.PubMedCrossRefGoogle Scholar
  19. 19.
    Bates DA, Winterbourn CC. Deoxyribose breakdown by the adriamycin semiquinone and H2O2: evidence for hydroxyl radical participation. FEBS Lett 1982; 145: 137–42.PubMedCrossRefGoogle Scholar
  20. 20.
    Ward PA, Till GO, Kunkel R et al. Evidence for role of hydroxyl radical in complement and neutrophil-dependent tissue injury. J Clin Invest 1983; 72: 789–801.PubMedCrossRefGoogle Scholar
  21. 21.
    Rowe TC, Chen GC, Hsiang V et al. DNA damage by antitumor acridines mediated by mammalian DNA topoisomerase II. Cancer Res 1986; 46: 2021–26.PubMedGoogle Scholar
  22. 22.
    Yang L, Rowe TC, Liu LF. Identification of topoisomerase II as the intracellular target of antitumor epipodophyllotoxins in simian virus 40 infected monkey cells. Cancer Res 1985; 45: 5872–76.PubMedGoogle Scholar
  23. 23.
    Hsiang YM, Lin ZF. Identification of mammalian DNA topoisomeras I as an intracellular target of the anticancer drug Camptothecin. Cancer Res 1988; 48: 1722–26.PubMedGoogle Scholar
  24. 24.
    Wall ME, Wani MD, Cook CE et al. Plant antitumor agens. I. the isolation and structure of camptothecin, a novel leukemia and tumor inhibitor from Camptotheca acuminata. J Am Chem Soc 1966; 88: 3888–90.CrossRefGoogle Scholar
  25. 25.
    Perdue RE Jr, Smith RL, Wall ME et al. Camptotheca acuminata Decaisne (Nyssaceae). Source of campothecin. U S Dept \Agr, Agr Research Service 1970; Technical Bulletin No. 1415: 1–26.Google Scholar
  26. 26.
    Heckendorf AH, Mattes KC, Hutchinson CR et al. Sterochemistry and conformation of biogenetic precursors of indole alkaloids. J Org Chem 1976; 41: 2045–47.PubMedCrossRefGoogle Scholar
  27. 27.
    Moertel CG, Schutt HJ, Reitmerer RJ et al. Phase II study of campthecin (NSC-100880) in the treatment of advanced gastrointestinal cancer. Cancer Chemother Rep Part I 1972; 56: 95.Google Scholar
  28. 28.
    Gottleib JA, Luce JK. Treatment of malignant melanoma with camptothecin (NSC-100880). Cancer Chemother Rep 1972; 56: 103.Google Scholar
  29. 29.
    Wani MC, Ronman PE, Lindley JT et al. Plant antitumor agents. 18. Synthesis and biological activity of camptothecin analogs. J Med Chem 1980; 23: 554–60.PubMedCrossRefGoogle Scholar
  30. 30.
    Andoh T, Ishii K, Suzuki Y et al. Characterization of mammalian mutant with camptothecin-resistant DNA topoisomerase I. Proc Natl Acad Sci USA 1987; 84: 5565–69.PubMedCrossRefGoogle Scholar
  31. 31.
    Gupta RS, Gupta R, Eng B et al. Camptothecin-resistant mutant of Chinese hamster ovary cells containing a resistant form of topoisomerase I. Cancer Res 1988; 48: 6404–10.PubMedGoogle Scholar
  32. 32.
    Per SR, Mattern MR, Mirabelli CK et al. Characterization of a subline of P388 leukemia resistant to amsacrine: Evidence of altered topoisomerase II function. Mol Pharmacol 1987; 32: 17–25.PubMedGoogle Scholar
  33. 33.
    Robson CN, Hoban PR, Harris AL et al. Cross-sensitivity to topoisomerase II inhibitors in cytotoxic drug-hypersensitive Chinese hamster ovary cell lines. Cancer Res 1987; 47: 1560–65.PubMedGoogle Scholar
  34. 34.
    Wang JC. DNA topoisomerases. Annu Rev Biochem 1985; 54: 665–97.PubMedCrossRefGoogle Scholar
  35. 35.
    Vosberg HP. DNA topoisomerases: Enzymes that control DNA conformation. Curr Top Microbiol Immunol 1985; 114: 19–102.PubMedCrossRefGoogle Scholar
  36. 36.
    Osheroff N. Biochemical basis for the interaction of type I and type II topoisomerases with DNA. Pharmacol Ther 1989; 41: 223–41.PubMedCrossRefGoogle Scholar
  37. 37.
    D’Arpa P, Liu LF. Topoisomerase-targeting antitumor drugs. Biochim Biophys Acta 1989; 989: 163–77.PubMedGoogle Scholar
  38. 38.
    Liu LF. DNA topoisomerase poisons as antitumor drugs. Annu Rev Biochem 1989; 58: 351–57.PubMedCrossRefGoogle Scholar
  39. 39.
    Rowe TC, Chen GL, Hsiang YH et al. DNA damage by antitumor acridines mediated by mammalian topoisomerase II. Cancer Res 1986; 46: 2021–26.PubMedGoogle Scholar
  40. 40.
    Osheroff N, Zechiedrich EL. Calcium-promoted DNA cleavage by eukaryotic topoisomerase II: Trapping the covalent enzyme-DNA complex in an active form. Biochemistry 1987; 26: 4303–9.PubMedCrossRefGoogle Scholar
  41. 41.
    Gibson BS, Ross WE. DNA topoisomerase II: a primer on the enzyme and its unique role as a multidrug target in cancer chemotherapy. Pharmacol Ther 1987; 32: 89–106.CrossRefGoogle Scholar
  42. 42.
    Kohn KW, Pommier Y, Kerriga D et al. Topoisomerase II as a target of anticancer drug action in mammalian cells. Natl Cancer Inst Monogr 1987; 4: 61–71.Google Scholar
  43. 43.
    Zwelling LA. Topoisomerase II as a target of antileukemia drugs: a review of controversial areas. Hum Pathol 1989; 3: 101–12.Google Scholar
  44. 44.
    Gulliya KS, Franck B, Schneider U et al. Topoisomerase II-dependent novel antitumor compounds Merocil and Merodantoin induce apoptosis in Daudi cells. Anticancer Drugs 1994; 5: 557–66.PubMedCrossRefGoogle Scholar
  45. 45.
    Sharma R, Arnold L, Gulliya KS. Correlation between DNA topoisomerase II and cytotoxicity in pMC540 and merodantoin sensitive and resistant human breast cancer cells. Anticancer Res 1995; 15: 295–304.PubMedGoogle Scholar
  46. 46.
    Marsh W, Center MS. Adriamycin resistance in HL-60 cells and accompanying modification of a surface membrane protein contained in drug-sensitive cells. Cancer Res 1987; 47: 5080–86.Google Scholar
  47. 47.
    Bhalla K, Hindenburg A, Taub RN et al. Isolation and characterization of an anthracycline-resistant human leukemia cell line. Cancer Res 1985; 45: 3657–62.PubMedGoogle Scholar
  48. 48.
    Beck WT, Cirtain MC, Danks MK et al. Pharmacological molecular, and cytogenetic analysis of “atypical” multidrug resistant human leukemic cells. Cancer Res 1987; 47: 5455–60.PubMedGoogle Scholar
  49. 49.
    Sinha BK, Haim N, Dusre L et al. DNA strand breaks produced by etaposide (VP-16, 213) in sensitive and resistant human breast tumor cells; Implication for the mechanism of action. Cancer Res 1988; 48: 5096–5100.Google Scholar
  50. 50.
    Ferguson PJ, Fisher MH, Stephenson J et al. Combined modalities of resistance in etoposide-resistant human KB cell lines. Cancer Res 1988; 48: 5956–64.PubMedGoogle Scholar
  51. 51.
    Marini JC, Miller KG, Englund PT. Decatenation of kinetoplast DNA by topoisomerase II. J Biol Chem 1980; 255: 4976–79.PubMedGoogle Scholar
  52. 52.
    Miller KG, Liu LF, Englund PT. A homogenous type II DNA topoisomerase from HeLa cell nuclei. J Biol Chem 1981; 256: 9334–39.PubMedGoogle Scholar
  53. 53.
    Liu LF, Miller KG. Eukaryotic DNA topoisomerases: two forms of type I DNA topoisomerases from HeLa cell nuclei. Proc Natl Acad Sci USA 1981; 78: 3487–91.PubMedCrossRefGoogle Scholar
  54. 54.
    Muller MT, Spitzner JR, Di Donato JA et al. Single-stranded DNA cleavage by eukaryotic topoisomerase II. Biochemistry 1988; 27: 8369–79.PubMedCrossRefGoogle Scholar
  55. 55.
    Spitzner JR, Chung IK, Muller MT. Eukaryotic topoisomerase II preferentially cleaves alternating purine-pyrimidine repeats. Nucleic Acids Res 1990; 18: 1–11.PubMedCrossRefGoogle Scholar
  56. 56.
    Foglesong PD, Reckord C. Improved electrophoretic separation of supercoiled and relaxed DNA in the presence of ethidium bromide. Biotechniques 1992; 13: 402–4.PubMedGoogle Scholar
  57. 57.
    Sullivan DM, Latham MD, Ross WE. Proliferation dependent topoisomerase II content as a determinant of antineoplastic drug action in human, mouse, and Chinese hamster ovary cells. Cancer Res 1987; 47: 3973–79.PubMedGoogle Scholar
  58. 58.
    Tsai-Pflugfelder M, Liu LF, Liu A et al. Cloning and sequencing of cDNA encoding human DNA topoisomerase II and localization of the gene to chromosome region 17q 21–22. Proc Natl Acad Sci USA 1988; 85: 7177–81.PubMedCrossRefGoogle Scholar
  59. 59.
    Jenkins JR, Ayton P, Jones T et al. Isolation of cDNA clones encoding the p isozyme of human DNA topoisomerase II and localization of the gene to chromosome 3p24. Nucl Acid Res 1992; 20: 5587–92.CrossRefGoogle Scholar
  60. 60.
    Woessner RD, Mattem MR. Proliferation and cell cycle dependent differences in expression of the 170 kilodalton and 180 kilodalton forms of topoisomerase II in NIH 3T3. Cell Growth and Differ 1991; 2: 209–14.Google Scholar
  61. 61.
    Drake FH et al.. Biochemical and Pharmacological properties of p170 and p180 forms of Topoisomerase II. Biochemistry 1989; 18:8154–60.Google Scholar
  62. 62.
    Taylor SS, Buechler JA, Yonemoto W. cAMP-dependent protein kinase: framework for a diverse family of regulatory enzymes. Annu Rev Biochem 1990; 59: 971–1005.PubMedCrossRefGoogle Scholar
  63. 63.
    Cho-Chung YS. Role of cyclic AMP receptor proteins in growth differentiation and suppression of malignancy: new approaches to therapy. Cancer Res 1990; 50: 7093–7100.PubMedGoogle Scholar
  64. 64.
    Tortora G, Yokozaki H, Pepe S et al. Differentiation of HL-60 leukemia by type I regulatory subunit antisense oligodeoxynucleotide of cAMP-dependent protein kinase. Proc Natl Acad Sci USA 1991; 88: 2011–15.PubMedCrossRefGoogle Scholar
  65. 65.
    Rohlff C, Clair T, Cho-Chung YS. 8-cl-cAMP induces down-regulation of the RIa subunit and up regulation of RIp subunit of cAMP-dependent protein kinase leading to type II holoenzyme-dependent growth inhibition and differentiation of HL-60 leukemia cells. J Biol Chem 1993; 268: 5774–82.PubMedGoogle Scholar
  66. 66.
    Tortora G, Pepe S, Cirafici AM et al. THS-regulated growth and cell cycle distribution of rat thyroid cells involve type I isozyme of cAMP-dependent protein kinase. Cell Growth and Differ 1993; 4: 359–65.Google Scholar
  67. 67.
    Totora G, Ciardiello F, Damiano V et al. Cyclic AMP-dependent protein kinase type I in hypersensitivity of human breast cells to topoisomerase II inhibitors. Clin Cancer Res 1995; 1: 49–56.Google Scholar
  68. 68.
    Gulliya KS, Pervaiz S, Dowben RM et al. Tumor cell specific dark cytotoxicity of preactivated merocyanine 540: implications for systemic therapy without light. Photochem Photobiol 1990; 52: 831–38.PubMedCrossRefGoogle Scholar
  69. 69.
    Glucksmann, A. Cell death in normal vertebrate ontogeny Biological Reviews 1951; 26: 59–86.Google Scholar
  70. 70.
    Kerr JFR, Wyllie AH, Currie AR. Apoptosis: a biological phenomenon with wide-ranging implications in tissue kinetics. Br J Cancer 1972; 26: 239–57.PubMedCrossRefGoogle Scholar
  71. 71.
    Wyllie AH, Kerr JFR, Currie AR. Cell death: the significance of apoptosis. Int Rev Cytol 1980; 68: 251–306.PubMedCrossRefGoogle Scholar
  72. 72.
    Wyllie AH. Glucocorticoid-induced thymocyte apoptosis is associated with endogenous endonuclease activation. Nature (London) 1980; 284: 555–56.CrossRefGoogle Scholar
  73. 73.
    Kerr John FR, Winterford Clay M, Harmon Brian V. Its significance in cancer and cancer therapy. Cancer 1994; 73: 2013–26.CrossRefGoogle Scholar
  74. 74.
    Fallon JF, Saunders JW. In vitro analysis of the control of cell death in a zone of prospective necrosis from the chich wing bud. Developmental Biology 1968; 18: 553–70.PubMedCrossRefGoogle Scholar
  75. 75.
    Schwartz LM, Truman JW. Peptide and steroid regulation of muscle degeneration in an insect. Science 1982; 215: 1420–21.PubMedCrossRefGoogle Scholar
  76. 76.
    Truman JW. Programmed cell death in the nervous system of an adult insect. Journal of Comparative Neurology 1983; 216: 445–52.PubMedCrossRefGoogle Scholar
  77. 77.
    Truman JW. Schwartz LM. Insect systems for the study of programmed neuronal death. Neuroscience Commentaries 1982; 1: 66–72.Google Scholar
  78. 78.
    Wyllie AH, Kerr JFR, Currie AR. Cell death in the normal neonatal rat adrenal cortex. J of Path 1973; 111: 255–61.CrossRefGoogle Scholar
  79. 79.
    Wyllie AH, Kerr JFR, MacAskill IAM, Currie AR. Adrenocortical cell deletion: The role of ACTH. J of Path 1973; 111: 85–94.CrossRefGoogle Scholar
  80. 80.
    Kerr JFR, Searle J. Deletion of cells by apoptosis during castration-induced involution of the rat prostate. Virchows Archiv (Cell Pathology) 1973; 13: 87–102.Google Scholar
  81. 81.
    Sandow BA, West NB, Norman RL, Brenner RM. Hormonal control of apoptosis in hamster uterine lumina epithelium. American Journal of Anatomy 1979; 156: 15–36.PubMedCrossRefGoogle Scholar
  82. 82.
    Ferguson DJP, Anderson TJ. Morphological evaluation of cell turnover in relation to the menstrual cycle of the “resting” breast. British Journal of Cancer 1981; 44: 177–81.PubMedCrossRefGoogle Scholar
  83. 83.
    Ferguson DJP, Anderson TJ. Ultrastructural observations on cell death by apoptosis in the `resting’ human breast. Virchows Archiv (Pathology, Anatomy) 1981; 393: 193–203.Google Scholar
  84. 84.
    Lennon SV, Martin SJ, Cotter TG. Induction of apoptosis (programmed cell death) in tumor cell lines by widely diverging stimuli. Biochem Soc Trans 1990; 18: 343–51.PubMedGoogle Scholar
  85. 85.
    Kyprianou N, English HF, Davidson NE et al. Programmed cell death during regression of the MCF-7 human breast cancer following oestrogen ablation. Cancer Res 1991; 51: 162–66.PubMedGoogle Scholar
  86. 86.
    Martikainen P, Kyprianou N, Tuker RW et al. Programmed cell death of nonproliferating androgen-independent prostatic cancer cells. Cancer Res 1991; 51: 4693–4700.PubMedGoogle Scholar
  87. 87.
    Wyllie AH. Apoptosis and the regulation of cell numbers in normal and neoplastic tissues: an overview. Cancer Metastases Reviews 1992; 11: 95–103.CrossRefGoogle Scholar
  88. 88.
    Kerr FRJ, Winterford CM. Apoptosis; Its significance in cancer and cancer therapy. Cancer 1994; 73: 2013–26.PubMedCrossRefGoogle Scholar
  89. 89.
    Duke RC, Cohen JJ, Chervenak R. Differences in target cell DNA fragmentation induced by mouse cytotoxic T lymphocytes and natural killer cells. J Immunol 1986; 137: 1442–47.PubMedGoogle Scholar
  90. 90.
    Ucker DS. Cytotoxic T-lymphocytes and glucocorticoids activate an endogenous suicide process in target cells. Nature (London) 1987; 327: 62–64.CrossRefGoogle Scholar
  91. 91.
    Tornei LD, Shapiro JP, Cope FO. Apoptosis in C3H/10T’“2 mouse embryonic cells: evidence for internucleosomal DNA modification in the absence of double-strand cleavage. Proc Natl Acad Sci U S A 1993; 90: 853–57.CrossRefGoogle Scholar
  92. 92.
    Whitfield JF, Perris AD, Youdale Y. Destruction of the nuclear morphology of thymic lymphocytes by the corticosteroid cortisol. Exp Cell Res 1968; 52: 349–62.PubMedCrossRefGoogle Scholar
  93. 93.
    Eastman A. The pathway of apoptosis activated by anticancer agents. Proceedings of the American Association for Cancer Res 1992; 33: 587–88.Google Scholar
  94. 94.
    Miyashita T, Reed JC. bd-2 transfer increases relative resistance of S49.1 and WEHI7.2 lymphoid cells to cell death and DNA fragmentation induced by glucocorticoids and multiple chemothera- peutic drugs. Cancer Res 1992; 52: 5407–541.PubMedGoogle Scholar
  95. 95.
    Hockenbery DM, Oltvai ZN, Yin X et al. Bd-2 Functions in an antioxidant pathway to prevent apoptosis. Cell 1993; 75: 241–51.PubMedCrossRefGoogle Scholar
  96. 96.
    Jacobsen MD, Burne JF, Raff MC. Programmed cell death and Bc1–2 protection in the absence of nucleus. EMBO J 1994; 13: 1899–1910.Google Scholar
  97. 97.
    Steller H. Apoptosis-mechanisms and genes of cellular suicide. Science 1995; 267: 1445–62.PubMedCrossRefGoogle Scholar
  98. 98.
    Johnson LV, Walsh ML, Brokus BJ et al. Monitoring of relative mitochondrial membrane potential in living cells by fluorescence microscopy. J Cell Biol 1981; 88: 526–35.PubMedCrossRefGoogle Scholar
  99. 99.
    Gulliya KS, Sharma R, Liu HW et al. Relationship of mitochondrial function and cellular adenosine triphosphate levels to pMC540 and merodantoin cytotoxicity in MCF-7 human breast cancer cells. Anticancer Drugs 1995; 6: 545–52.PubMedCrossRefGoogle Scholar

Copyright information

© Springer-Verlag Berlin Heidelberg 1996

Authors and Affiliations

  • Kirpal S. Gulliya
    • 1
    • 2
    • 3
  1. 1.Baylor Research InstituteBaylor University Medical CenterDallasUSA
  2. 2.Institute of Biomedical StudiesBaylor UniversityWacoUSA
  3. 3.Department of Biological ScienceUniversity of North TexasDentonUSA

Personalised recommendations