Radio- und Photosensitivierung von Plasmid-DNA durch den DNA-bindenden Liganden Propidiumiodid: Untersuchungen zur Auger-Elektronen-Induktion und zum Nachweis von Cherenkov-Strahlung

Zusammenfassung

Ziel Untersucht wurde, ob Propidiumiodid (PI) die DNA-schädigende Wirkung von ionisierender und nicht ionisierender Strahlung (Röntgenstrahlung, Alpha-, Beta-, Auger-Elektronen-Strahlung bzw. Licht diverser Wellenlängen) verstärken kann. Diese biophysikalische Versuchsanordnung ermöglicht es zu überprüfen, ob Cherenkov-Strahlung in relevantem Umfang via photodynamischer Effekte und erhöhter DNA-Schädigung nachweisbar ist.

Material und Methoden Konformationsänderungen der Plasmid-DNA durch DNA-Schäden wurden mittels Gelelektrophorese und Fluoreszenzfärbung detektiert und quantifiziert. Wasserstoffperoxid, Zinndichlorid und Dimethylsulfoxid wurden als chemische Modulatoren, Tc-99m, Re-188, Ra-223 und Röntgenstrahlung (32 kV und 200 kV) zur Bestimmung der Radiotoxizität und Licht (λ = 254 nm, 366 nm und 530–575 nm) zur Bestimmung der Phototoxizität eingesetzt.

Ergebnisse Die Radiotracer und die Röntgenstrahlung verursachten dosisabhängige DNA-Schäden. PI fungierte nicht als Radio-Sensitizer bei den Radionukliden und nur in geringem Maß bei Röntgenstrahlung. Die Phototoxizität war abhängig von der Wellenlänge. Licht im Wellenlängenbereich von 530–575 nm (VIS) resultierte in Kombination mit PI in direkten DNA-Schäden. Die Ausbeuten der Cherenkov-Strahlung lagen weit unter der Photonen-Emission der Lichtbestrahlung und konnten daher von der Radiotoxizität nicht unterschieden werden.

Schlussfolgerung PI bindet an Plasmid-DNA, ist nicht chemotoxisch und steigert kaum die Radiotoxizität. Die Phototoxizität und die Wirkung von PI sind abhängig von der Wellenlänge. Keine Art der Energiezufuhr konnte via PI eine Auger-Elektronen-Kaskade induzieren. Auch eine erhöhte DNA-Schädigung durch photodynamische Effekte via Cherenkov-Strahlung war nicht nachweisbar.

Abstract

Purpose We investigated whether propidium iodide (PI) enhances DNA damaging effects of ionizing and non-ionizing radiation species (X-rays, alpha-, beta-, auger electron emission and light of various wavelengths, respectively). This biophysical experimental setting allowed us, furthermore, to investigate whether Cherenkov emission can be detected by photodynamic effects and increased DNA damage.

Material and methods Conformation changes of plasmid DNA were detected and quantified by gelelectrophoresis and fluorescence imaging. Hydrogen peroxide, stannous dichloride, and dimethylsulfoxide were used as chemical modulators, Tc-99m, Re-188, Ra-223, and x-ray (32 kV and 200 kV) reflected radiotoxicity and light (λ = 254 nm, 366 nm and 530–575 nm) induced phototoxicity.

Results Radiotracers and x-rays induced dose dependent DNA damage. PI did not serve as radiosensitizer in radioisotopes, while a low effect was detected in X-rays. The phototoxicity was dependent on the wavelengths of light. Light with a wavelength range of 530–575 nm in combination with PI resulted in direct DNA damage. The yield of Cherenkov emission was far below the photon emission of light irradiation and not distinguishable from general radiotoxicity.

Conclusions PI binds to plasmid DNA, is not chemotoxic, and increases radiotoxicity only to minor extent. Phototoxicity and its stimulation by PI is dependent on the wavelength of the light. No kind of energy deposition was capable of inducing an Auger electron cascade. Furthermore, no increase in DNA damage induced by photodynamic effects from Cherenkov emission was detectable.

Schlüsselwörter

Propidiumiodid - Plasmid-DNA - Auger-Elektronen - Cherenkov-Strahlung

Key words

Propidium iodide - plasmid DNA - Auger electrons - Cherenkov emission

• Literatur

• 1 Martin RF, Murray V, D’Cunha G. et al. Radiation sensitization by an iodine-labelled DNA ligand. Int J Radiat Biol 1990; 57: 939-946
• 2 Nath R, Bongiorni P, Rockwell S. Enhancement of IUdR radiosensitization by low energy photons. Int J Radiat Oncol Biol Phys 1987; 13: 1071-1079
• 3 Shinohara K, Nakano H, Ohara H. Detection of Auger enhancement induced in HeLa cells labeled with iododeoxyuridine and irradiated with 150 kV x-rays--Effects of cysteamine and dimethylsulfoxide. Acta Oncol 1996; 35: 869-875
• 4 Kobayashi K, Usami N, Sasaki I. et al. Study of Auger effect in DNA when bound to molecules containing platinum. A possible application to hadrontherapy. Nucl Instrum Meth B 2003; 199: 348-355 doi:10.1016/S0168–583x(02)01532-X
• 5 Maeda M, Kobayashi K, Hieda K. Efficiencies of induction of DNA double strand breaks in solution by photoabsorption at phosphorus and platinum. Int J Radiat Biol 2004; 80: 841-847 doi:10.1080/09553000400017598
• 6 Lobachevsky PN, Martin RF. DNA breakage by decay of Auger electron emitters: experiments with 123I-iodoHoechst 33258 and plasmid DNA. Radiation research 2005; 164: 766-773
• 7 Kral T, Widerak K, Langner M. et al. Propidium iodide and PicoGreen as dyes for the DNA fluorescence correlation spectroscopy measurements. J Fluoresc 2005; 15: 179-183 doi:10.1007/s10895–005–2526–2
• 8 Kotzerke J, Punzet R, Runge R. et al. 99mTc-labeled HYNIC-DAPI causes plasmid DNA damage with high efficiency. PLoS One 2014; 9: e104653 doi:10.1371/journal.pone.0104653
• 9 Reissig F, Mamat C, Steinbach J. et al. Direct and Auger Electron-Induced, Single- and Double-Strand Breaks on Plasmid DNA Caused by 99mTc-Labeled Pyrene Derivatives and the Effect of Bonding Distance. PLoS One 2016; 11: e0161973 doi:10.1371/journal.pone.0161973
• 10 Dolmans DE, Fukumura D, Jain RK. Photodynamic therapy for cancer. Nat Rev Cancer 2003; 3: 380-387 doi:10.1038/nrc1071
• 11 Dothager RS, Goiffon RJ, Jackson E. et al. Cerenkov radiation energy transfer (CRET) imaging: a novel method for optical imaging of PET isotopes in biological systems. PLoS One 2010; 5: e13300 doi:10.1371/journal.pone.0013300
• 12 Thorek DL, Ogirala A, Beattie BJ. et al. Quantitative imaging of disease signatures through radioactive decay signal conversion. Nat Med 2013; 19: 1345-1350 doi:10.1038/nm.3323
• 13 Grimm J. Cancer nanomedicine: Therapy from within. Nat Nanotechnol 2015; 10: 299-300 doi:10.1038/nnano.2015.63
• 14 Kotagiri N, Sudlow GP, Akers WJ. et al. Breaking the depth dependency of phototherapy with Cerenkov radiation and low-radiance-responsive nanophotosensitizers. Nat Nanotechnol 2015; 10: 370-379 doi:10.1038/nnano.2015.17
• 15 Runge R, Oehme L, Kotzerke J. et al. The effect of dimethyl sulfoxide on the induction of DNA strand breaks in plasmid DNA and colony formation of PC Cl3 mammalian cells by alpha-, beta-, and Auger electron emitters (223)Ra, (188)Re, and (99 m)Tc. EJNMMI Res 2016; 6: 48 doi:10.1186/s13550–016–0203-x
• 16 Schindelin J, Arganda-Carreras I, Frise E. et al. Fiji: an open-source platform for biological-image analysis. Nat Methods 2012; 9: 676-682 doi:10.1038/nmeth.2019
• 17 Dantas FJ, Moraes MO, de Mattos JC. et al. Stannous chloride mediates single strand breaks in plasmid DNA through reactive oxygen species formation. Toxicol Lett 1999; 110: 129-136
• 18 Lobachevsky PN, Martin RF. Plasmid DNA breakage by decay of DNA-associated auger emitters: experiments with 123I/125I-iodoHoechst 33258. Int J Radiat Biol 2004; 80: 915-920 doi:10.1080/09553000400017754
• 19 Corde S, Joubert A, Adam JF. et al. Synchrotron radiation-based experimental determination of the optimal energy for cell radiotoxicity enhancement following photoelectric effect on stable iodinated compounds. Br J Cancer 2004; 91: 544-551 doi:10.1038/sj.bjc.6601951
• 20 Fairchild RG. Contribution to radiation enhancement from Auger cascades. Int J Radiat Oncol Biol Phys 1987; 13: 1262-1263
• 21 Laster BH, Thomlinson WC, Fairchild RG. Photon activation of iododeoxyuridine: biological efficacy of Auger electrons. Radiation research 1993; 133: 219-224
• 22 Schmid E, Regulla D, Kramer HM. et al. The effect of 29 kV X rays on the dose response of chromosome aberrations in human lymphocytes. Radiation research 2002; 158: 771-777
• 23 Pettinger KH, Wimmer B, Wabner W. Atrazinentfernung aus Trinkwasser durch UV-aktiviertes Wasserstoffperoxid. Das Gas- und Wasserfach Ausgabe Wasser, Abwasser 1991; 132: 553-558
• 24 Russo D, Siciliano A, Guida M. et al. Photodegradation and ecotoxicology of acyclovir in water under UV254 and UV254/H2O2 processes. Water Res 2017; 122: 591-602 doi:10.1016/j.watres.2017.06.020
• 25 Zalazar CS, Labas MD, Brandi RJ. et al. Dichloroacetic acid degradation employing hydrogen peroxide and UV radiation. Chemosphere 2007; 66: 808-815 doi:10.1016/j.chemosphere.2006.06.044
• 26 Krishan A. Rapid flow cytofluorometric analysis of mammalian cell cycle by propidium iodide staining. J Cell Biol 1975; 66: 188-193
• 27 Grosswendt B. Nanodosimetry, the metrological tool for connecting radiation physics with radiation biology. Radiat Prot Dosimetry 2006; 122: 404-414 doi:10.1093/rpd/ncl469
• 28 Gill RK, Mitchell GS, Cherry SR. Computed Cerenkov luminescence yields for radionuclides used in biology and medicine. Phys Med Biol 2015; 60: 4263-4280 doi:10.1088/0031–9155/60/11/4263
• 29 Cho JS, Taschereau R, Olma S. et al. Cerenkov radiation imaging as a method for quantitative measurements of beta particles in a microfluidic chip. Phys Med Biol 2009; 54: 6757-6771 doi:10.1088/0031–9155/54/22/001
• 30 Kotagiri N, Niedzwiedzki DM, Ohara K. et al. Activatable probes based on distance-dependent luminescence associated with Cerenkov radiation. Angew Chem Int Ed Engl 2013; 52: 7756-7760 doi:10.1002/anie.201302564
• 31 Pratx G, Kapp DS. Is Cherenkov luminescence bright enough for photodynamic therapy?. Nat Nanotechnol 2018; 13: 354 doi:10.1038/s41565–018–0142-y
• 32 Niu G, Chen X. When radionuclides meet nanoparticles. Nat Nanotechnol 2018; 13: 359-360 doi:10.1038/s41565–018–0103–5
• 33 Tamura R, Pratt EC, Grimm J. Innovations in Nuclear Imaging Instrumentation: Cerenkov Imaging. Semin Nucl Med 2018; 48: 359-366 doi:10.1053/j.semnuclmed.2018.02.007