Skip to main content
SpringerLink
Log in

Targeting of photooxidative damage on single-stranded DNA representing the bcr-abl chimeric gene using oligonucleotide-conjugates containing [Ru(phen)3]2+-like photosensitiser groups

  • Paper
  • Published:
Photochemical & Photobiological Sciences Aims and scope Submit manuscript

Abstract

Photooxidative damage was induced predominantly at a single guanine base in a target DNA by irradiation (λ > 330 nm) in the presence of complementary oligodeoxynucleotide conjugates (ODN-5′-linker-[Ru(phen)3]2+) (phen = 1,10-phenanthroline). The target DNA represents the b2a2 variant of the chimeric bcr-abl gene implicated in the pathogenesis of chronic myeloid leukaemia, and the sequence of the 17mer ODN component of the conjugate (3′ G G T A G T T A T T C C T T C T T 5′) was complementary to the junction region of the sense strand sequence of this oncogene. Two different conjugates were prepared, both of them by reaction of the appropriate succinimide ester with 5′-hexylamino-derivatised 17mer ODN. In Ru—ODN-1 (7) the linker was—(CH2)6-NHCO-bpyMe (-bpyMe = 4′-[4-methyl-2,2′-bipyridyl]), whereas in Ru—ODN-2 (13) it was—(CH2)6-NHCO—(CH2)3-CONH-phen. Photoexcitation of either of the conjugates when hybridised with the 32P-5′-end-labelled target 34mer 5′T G A C̲ C̲ A̲ T̲ C̲ A̲ A̲ T̲ A̲ A̲ G̲ G̲ A̲ A̲ G̲ A̲ A̲ G21 C C C T T C A G C G G C C 3′ (ODN binding site underlined) led to an alkali-labile site predominantly (> 90%) at the G21 base, which is at the junction of double-stranded and single-stranded regions of the hybrid. Greater yields were found with Ru—ODN-1 (7) than with Ru—ODN-2 (13). In contrast to this specific cleavage with Ru—ODN-1 (7) or Ru—ODN-2 (13), alkali-labile sites were generated at all guanines when the 34mer was photolysed in the presence of the free sensitiser [Ru(phen)3]2+. Since [Ru(phen)3]2+ was shown to react with 2′-deoxyguanosine to form the diastereomers of a spiroiminodihydantoin derivative (the product from 1O2 reaction), 1O2 might also be an oxidizing species in the case of Ru—ODN-1 (7) and Ru—ODN-2 (13). Therefore to determine the range of reaction, a series of ‘variant’ targets was prepared, in which G21 was replaced with a cytosine and a guanine substituted for a base further towards the 3′-end (e.g. Variant 3; 5′T G A C C A T C A A T A A G G A A G A A C C G23 C T T C A G C G G32 C C 3′). While it was noted that efficient reaction took place at distances apparently remote from the photosensitiser (e.g. at G32, but not G23 for Variant 3), this effect could be attributed to hairpinning of the single-stranded region of the target. These results are therefore consistent with the photooxidative damage being induced by a reaction close to the photosensitiser rather than by a diffusible species such as 1O2.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Similar content being viewed by others

References

  1. K. E. Erikkila, D. T. Odom, J. K. Barton, Recognition and reaction of metallointercalators with DNA, Chem. Rev., 1999, 99, 2777–2795.

    Article  CAS  Google Scholar 

  2. B. Nordén, P. Lincoln, B. Akerman and E. Tuite, Probing of nucleic acids by metal ion complexes of small molecules, in Metal Ions in Biological Systems, ed. A. Sigel and H. Sigel, Marcel Dekker, New York, 1996, vol. 33, pp. 177–252.

  3. T. Da Ros, G. Spalluto, A. S. Boutorine, R. V. Bensasson, M. Prato, DNA-photocleavage agents, Curr. Pharm. Des., 2001, 7, 1781–1821.

    PubMed  Google Scholar 

  4. C. Moucheron, A. Kirsch-De Mesmaeker, J. M. Kelly, Photoreactions of ruthenium(II) and osmium(II) complexes with deoxyribonucleic acid (DNA), J. Photochem. Photobiol. B., 1997, 40, 91–106.

    Article  CAS  PubMed  Google Scholar 

  5. C. Moucheron, A. Kirsch-De Mesmaeker, J. M. Kelly, Photophysics and photochemistry of metal polypyridyl and related complexes with nucleic acids, Struct. Bonding, 1998, 92, 163–216.

    Article  CAS  Google Scholar 

  6. J. M. Kelly, A. B. Tossi, D. J. McConnell, C. OhUigin, A study of the interactions of some polypyridylruthenium(II) complexes with DNA using fluorescence spectroscopy, topoisomerisation and thermal denaturation, Nucleic Acid Research, 1985, 13, 6017–6034

    Article  CAS  Google Scholar 

  7. A. B. Tossi, J. M. Kelly, A study of some polypyridylruthenium(II) complexes as DNA binders and photocleavage reagents, Photochem. Photobiol., 1989, 49, 545–556.

    Article  CAS  PubMed  Google Scholar 

  8. A. Aboul-Enein, D. Schulte-Frohlinde, Biological deactivation and single-strand breakage of plasmid DNA by photosensitization using tris(2,2′-bipyridyl)ruthenium(II) and peroxydisulfate, Photochem. Photobiol., 1988, 48, 27–34.

    Article  CAS  PubMed  Google Scholar 

  9. C. Sentage, J. C. Chambron, J. P. Sauvage, N. Paillous, Tuning the mechanism of DNA cleavage photosensitized by ruthenium dipyridophenazine complexes by varying the structure of the two non intercalating ligands, J. Photochem. Photobiol. B., 1994, 26, 165–174.

    Article  Google Scholar 

  10. J. M. Kelly, A. B. Tossi, D. J. McConnell, C. OhUigin, C. Hélène and T. le Doan, Interaction of ruthenium polypyridyl complexes with DNA and their use as sensitisers for its cleavage, in Free Radicals, Metal Ions and Biopolymers, ed. P. C. Beaumont, D. J. Deeble, B. J. Parsons and C. Rice Evans, Richelieu Press, London, 1989, p. 143.

  11. S. Delaney, M. Pascaly, P. K. Bhattacharya, K. Han, J. K. Barton, Oxidative damage by ruthenium complexes containing the dipyridophenazine ligand or its derivatives: a focus on intercalation, Inorg. Chem., 2002, 41, 1966–1974.

    Article  CAS  PubMed  Google Scholar 

  12. L. Jaquet, R. J. H. Davies, A. Kirsch-De Mesmaeker, J. M. Kelly, Photoaddition of [Ru(tap)2(bpy)]2+ to DNA: A new mode of covalent attachment of metal complexes to duplex DNA, J. Am. Chem. Soc., 1997, 119, 11763–11768.

    Article  Google Scholar 

  13. J-P. Lecomte, A. Kirsch-De Mesmaeker, M. M. Feeney, J. M. Kelly, Ruthenium(II) complexes with 1,4,5,8,9,12-hexaazatriphenylene and 1,4,5,8-tetraazaphenanthrene ligands: key role played by the photoelectron transfer in DNA cleavage and adduct formation, Inorg. Chem., 1995, 34, 6481–6491.

    Article  CAS  Google Scholar 

  14. Y. Jenkins, J. K. Barton, A sequence-specific molecular light switch: tethering of an oligonucleotide to a dipyridophenazine complex of ruthenium(II), J. Am. Chem. Soc., 1992, 114, 8736–8738.

    Article  CAS  Google Scholar 

  15. X. Hu, G. D. Smith, M. Sykora, S. J. Lee, M. W. Grinstaff, Automated solid-phase synthesis and photophysical properties of oligodeoxynucleotides labeled at 5′-aminothymidine with Ru(bpy)2(4-m-4′-cam-bpy)2+, Inorg. Chem., 2000, 39, 2500–2504.

    Article  CAS  PubMed  Google Scholar 

  16. D. J. Hurley, Y. Tor, Metal-containing oligonucleotides: solid-phase synthesis and luminescence properties, J. Am. Chem. Soc., 1998, 120, 2194–2195.

    Article  CAS  Google Scholar 

  17. E. Meggers, D. Kusch, B. Giese, An effective synthesis of enantiomerically pure Δ- and Λ-ruthenium(II) labelled oligonucleotides, Helv. Chim. Acta., 1997, 80, 640–652.

    Article  CAS  Google Scholar 

  18. D. Ossipov, P. I. Pradeepkumar, M. Holmer, J. Chattopadhyaya, Synthesis of [Ru(phen)2dppz]2+-tethered oligo-DNA and studies on the metallointercalation mode into the DNA duplex, J. Am. Chem. Soc., 2001, 123, 3551–3562.

    Article  CAS  PubMed  Google Scholar 

  19. W. Bannwarth, D. Schmidt, R. L. Stallard, C. Hornung, R. Knorr, F. Müller, Bathophenanthroline-ruthenium(II) complexes as non-radioactive labels for oligonucleotides which can be measured by time-resolved fluorescence techniques, Helv. Chim. Acta., 1988, 71, 2085–2099.

    Article  CAS  Google Scholar 

  20. J. Telser, A. Cruickshank, K. S. Schanze, T. L. Netzel, DNA oligomers and duplexes containing a covalently attached derivative of tris (2, 2′ -bipyridine) ruthenium(II): synthesis and characterization by thermodynamic and optical spectroscopic measurements, J. Am. Chem. Soc., 1989, 111, 7221–7226.

    Article  CAS  Google Scholar 

  21. G. N. Grimm, A. S. Boutorine, P. Lincoln, B. Nordén, C. Hélène, Formation of DNA triple helices by an oligonucleotide conjugated to a fluorescent ruthenium complex, ChemBioChem., 2002, 3, 324–331.

    Article  CAS  PubMed  Google Scholar 

  22. D. García-Fresnadillo, N. Boutonnet, S. Schumm, C. Moucheron, A. Kirsch-De Mesmaeker, E. Defrancq, J. F. Constant, J. Lhomme, Luminescence quenching of Ru-labeled oligonucleotides by targeted complementary strands, Biophys. J., 2002, 82, 978–987.

    Article  PubMed  PubMed Central  Google Scholar 

  23. S. Schumm, M. Prévost, D. García-Fresnadillo, O. Lentzen, C. Moucheron, A. Kirsch-De Mesmaeker, Influence of the sequence dependent ionisation potentials of guanines on the luminescence quenching of Ru-labeled oligonucleotides: a theoretical and experimental study, J. Phys. Chem. B, 2002, 106, 2763–2768.

    Article  CAS  Google Scholar 

  24. C. G. Coates, J. J. McGarvey, P. L. Callaghan, M. Colleti, J. G. Hamilton, Probing the interaction of [Ru(phen)2dppz]2+ with single-stranded DNA–what degree of protection is required for operation of the “light-switch effect”?, J. Phys. Chem. B, 2001, 105, 730–735.

    Article  CAS  Google Scholar 

  25. P. K. Bhattacharya, J. K. Barton, Influence of intervening mismatches on long-range guanine oxidation in DNA duplexes, J. Am. Chem. Soc., 2001, 123, 8649–8656.

    Article  CAS  PubMed  Google Scholar 

  26. I. Ortmans, S. Content, N. Boutonnet, A. Kirsch-De Mesmaeker, W. Bannwarth, J-F. Constant, E. Defrancq, J. Lhomme, Ru-labeled oligonucleotides for photoinduced reactions on targeted DNA guanines, Chem. Eur. J., 1999, 5, 2712–2721.

    Article  CAS  Google Scholar 

  27. R. E. Clarke, Antisense therapeutics in chronic myeloid leukaemia: the promise, the progress and the problems, Leukaemia, 2000, 14, 347–355.

    Article  CAS  Google Scholar 

  28. A. Hergueta-Bravo, M. E. Jiménez-Hernández, F. Montero, E. Oliveros, G. Orellana, Singlet oxygen-mediated DNA photocleavage with Ru(II) polypyridyl complexes, J. Phys. Chem. B, 2002, 106, 4010–4017.

    Article  CAS  Google Scholar 

  29. F. Schubert, A. Knaf, U. Möller, D. Cech, Covalent attachment of methylene blue to oligonucleotides, Nucleosides Nucleotides, 1995, 14, 1437–1443.

    Article  CAS  Google Scholar 

  30. S. I. Khan, A. E. Beilstein, G. D. Smith, M. Sykora, M. W. Grinstaff, Synthesis and excited-state properties of a novel ruthenium nucleoside: 5-[Ru(bpy)2(4-m-4′-pa-bpy)]2+-deoxyuridine, Inorg. Chem., 1999, 38, 2411–2415.

    Article  CAS  Google Scholar 

  31. J. Feely, P. V. Kavanagh, S. M. McNamara, J. E. O’Brien, Simple preparation of the major urinary metabolites of flunitrazepam and nitrazepam, Iran. J. Med. Sci., 1999, 168, 8–9.

    Article  CAS  Google Scholar 

  32. B. H. Han, D. H. Shin, S. Y. Cho, Graphite catalysed reduction of aromatic and aliphatic nitro compounds with hydrazine hydrate, Tetrahedron Lett., 1985, 26, 6233–6234.

    Article  CAS  Google Scholar 

  33. N. Y. Sardesai, S. C. Lin, K. Zimmermann, J. K. Barton, Construction of coordinatively saturated rhodium complexes containing appended peptides, Bioconjugate Chem., 1995, 6, 302–312.

    Article  CAS  Google Scholar 

  34. I. Saito, T. Nakamura, K. Natatani, Mapping of highest occupied molecular orbitals of duplex DNA by cobalt-mediated guanine oxidation, J. Am. Chem. Soc., 2000, 122, 3001–3006.

    Article  CAS  Google Scholar 

  35. F. Wilkinson, W. P. Helman, A. B. Ross, Rate constants for the decay and reactions of the lowest electronically excited singlet-state of molecular-oxygen in solution-an expanded and revised compliation, J. Phys. Chem. Ref. Data, 1995, 24, 663–1021.

    Article  CAS  Google Scholar 

  36. J. Van Houten, R. J. Watts, Temperature dependence of the photophysical and photochemical properties of the tris(2,2′-bipyridyl)ruthenium(II) ion in aqueous solution, J. Am. Chem. Soc., 1976, 98, 4853–4858.

    Article  Google Scholar 

  37. J-L. Ravanat, T. Douki, M-F. Incardona, J. Cadet, HPLC separations of normal and modified nucleobases and nucleosides on an amino-silica gel column, J. Liq. Chromatogr., 1993, 36, 3185–3202.

    Article  Google Scholar 

  38. J. Cadet and P. Vigny, in Bioorganic Chemistry, ed. H. Morrison, Wiley, New York, 1990, vol. 1, pp. 1–272.

  39. J. C. Niles, J. S. Wishnok, S. R. Tannenbaum, Spiroimino-dihydantoin is the major product of the 8-oxo-7,8-dihydroguanosine reaction with peroxynitrite in the presence of thiols and guanosine photoxidation by methylene blue, Org. Lett., 2001, 3, 963–966.

    CAS  PubMed  Google Scholar 

  40. W. Adam, M. A. Arnold, M. Grüne, W. M. Nau, U. Pischel, C. R. Saha-Müller, Spiroiminodihydantoin is a major product in the photooxidation of 2′-deoxyguanosine by the triplet states and oxyl radicals generated from hydroxyacetophenone photolysis and dioxetane thermolysis, Org. Lett., 2002, 4, 537–540.

    Article  CAS  PubMed  Google Scholar 

  41. T. Douki, J. Cadet, Modification of DNA bases by photosensitized one-electron oxidation, Int. J. Radiat. Biol., 1999, 75, 571–581.

    Article  CAS  PubMed  Google Scholar 

  42. B. Lewin, Genes V, Oxford University Press Inc., New York, 1994.

  43. F. Prat, C-C. Hou, C. S. Foote, Determination of the quenching rate constants of singlet oxygen by derivatized nucleosides in nonaqueous solution, J. Am. Chem. Soc., 1997, 119, 5051–5052.

    Article  CAS  Google Scholar 

  44. A. S. Boutorine, D. Brault, M. Takasugi, O. Delgado, C. Helene, Chlorin-oligonucleotide conjugates: synthesis, properties, and red light-induced photochemical sequence-specific DNA cleavage in duplexes and triplexes, J. Am. Chem. Soc., 1996, 118, 9469–9476.

    Article  CAS  Google Scholar 

  45. I. V. Yang, H. H. Thorp, Kinetics of metal-mediated one-electron oxidation of guanine in polymeric DNA and in oligonucleotides containing trinucleotide repeat sequences, Inorg. Chem., 2000, 39, 4969–4976.

    Article  CAS  PubMed  Google Scholar 

  46. X. Y. Zhang, M. A. J. Rodgers, Energy and electron-transfer reactions of the MLCT state of ruthenium tris(bipyridyl) with molecular-oxygen–a laser flash-photolysis study, J. Phys. Chem., 1995, 99, 12797–12803.

    Article  CAS  Google Scholar 

  47. B. P. Sullivan, D. J. Salmon, T. J. Meyer, Mixed phosphine 2,2′-bipyridine complexes of ruthenium, Inorg. Chem., 1978, 17, 3334–3341.

    Article  CAS  Google Scholar 

  48. G. Sprintschnik, H. W. Sprintschnik, P. P. Kirsch, D. G. Whitten, Preparation and photochemical reactivity of surfactant ruthenium(II) complexes in monolayer assemblies and at water-solid interfaces, J. Am. Chem. Soc., 1977, 15, 4947–4954.

    Article  Google Scholar 

  49. C. D. Ellis, L. D. Margerum, R. W. Murray, T. J. Meyer, Oxidative electropolymerization of polypyridyl complexes of ruthenium, Inorg. Chem., 1983, 22, 1283–1291.

    Article  CAS  Google Scholar 

  50. E. Koft, F. H. Case, substituted 1,10-phenanthrolines, benzo and pyrido derivatives, J. Org. Chem., 1962, 27, 865–868.

    Article  CAS  Google Scholar 

  51. T. Maniatis, E. F. Fritch, J. Sambrook, Molecular Cloning, a laboratory manual, 2nd edn., Cold Spring Harbour Lab, NY, 1989.

    Google Scholar 

Download references

Author information

Authors and Affiliations

  1. Chemistry Department, Trinity College, University of Dublin, Dublin 2, Ireland

    Conor W. Crean, Yvonne T. Kavanagh, Clare M. O’Keeffe, Peter H. Boyle & John M. Kelly

  2. Department of Haematology and Institute for Molecular Medicine, Trinity College and St. James’ Hospital, Dublin 8, Ireland

    Mark P. Lawler

  3. School of Biology and Biochemistry, Queen’s University Belfast, Belfast, UK, BT9 7BL

    Clarke Stevenson & R. Jeremy H. Davies

Authors
  1. Conor W. Crean
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Yvonne T. Kavanagh
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Clare M. O’Keeffe
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Mark P. Lawler
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Clarke Stevenson
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. R. Jeremy H. Davies
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Peter H. Boyle
    View author publications

    You can also search for this author in PubMed Google Scholar

  8. John M. Kelly
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to John M. Kelly.

Rights and permissions

Reprints and permissions

About this article

Cite this article

Crean, C.W., Kavanagh, Y.T., O’Keeffe, C.M. et al. Targeting of photooxidative damage on single-stranded DNA representing the bcr-abl chimeric gene using oligonucleotide-conjugates containing [Ru(phen)3]2+-like photosensitiser groups. Photochem Photobiol Sci 1, 1024–1033 (2002). https://doi.org/10.1039/b207387k

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1039/b207387k

Search

Navigation