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LED Light Source for in vitro Study of Photosensitizing Agents for Photodynamic Therapy

LED Light Source for in vitro Study of Photosensitizing Agents for Photodynamic Therapy

Shilyagina N.Y., Plekhanov V.I., Shkunov I.V., Shilyagin P.А., Dubasova L.V., Brilkina А.А., Sokolova E.A., Turchin I.V., Balalaeva I.V.
Keywords: LED light source; photosensitizer; photodynamic activity; tumor cell culture.
СТМ, 2014, volume 6, issue 2, pages 15-24.

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The aim of the investigation was to develop a LED light source providing a homogeneous light distribution in 96-well plates and allowing an independent irradiation of individual wells, as well as its experimental testing in in vitro study of photosensitizers for photodynamic therapy.

Materials and Methods. The experiments were carried out on human cell lines of epidermoid carcinoma А-431 and human bladder carcinoma Т24. Two photosensitizers for fluorescence diagnostics and photodynamic therapy were used: Photosens®, and newly synthesized polymer brushes nanoparticles doped with porphyrazine chromophore. To study the photo-activity of the agents we created a light source with replaceable LED-arrays. Photodynamic activity of the photosensitizers was estimated in vitro by MTT assay.

Results. The created LED light source enables to expose cell cultures with narrow-band irradiation with different wavelengths and radiation intensity being up to 90 mW/cm2. Precision control of temperature conditions during the investigation is provided. Light power instability is less than 1%. Independent on and off switching of LED clusters of 4 elements is provided in order to illuminate several wells groups of a standard 96-well culture plate with different light doses simultaneously. The developed light source was tested in the study of photo-activity of agents for photodynamic therapy. The dependence of Photosens® toxicity on the light dose was evaluated and significant photodynamic activity of newly synthesized porphyrazine fluorophore was demonstrated.

Conclusion. A newly developed LED light source with replaceable LED-arrays with narrow spectral bands provides an effective in vitro study of photosensitizing preparations under development. The state-of-the-art approach enables high throughput screening of promising agents for fluorescence diagnostics and photodynamic therapy.

  1. Mroz P., Bhaumik J., Dogutan D.K., Aly Z., Kamal Z., Khalid L., Kee H.L., Bocian D.F., Holten D., Lindsey J.S., Hamblin M.R. Imidazole metalloporphyrins as photosensitizers for photodynamic therapy: role of molecular charge, central metal and hydroxyl radical production. Cancer Lett 2009; 282(1): 63–76,
  2. Furre I.E., Shahzidi S., Luksiene Z., Moller M.T., Borgen E., Morgan J., Tkacz-Stachowska K., Nesland J.M., Peng Q. Targeting PBR by hexaminolevulinate-mediated photodynamic therapy induces apoptosis through translocation of apoptosis-inducing factor in human leukemia cells. Cancer Res 2005; 65(23): 11051–11060,
  3. Akhlynina T.V., Jans D.A., Rosenkranz A.A., Statsyuk N.V., Balashova I.Y., Toth G., Pavo I., Rubin A.B., Sobolev A.S. Nuclear targeting of chlorin e6 enhances its photosensitizing activity. J Biol Chem 1997; 272(33): 20328–20331.
  4. Krieg R.C., Messmann H., Schlottmann K., Endlicher E., Seeger S., Scholmerich J., Knuechel R. Intracellular localization is a cofactor for the phototoxicity of protoporphyrin IX in the gastrointestinal tract: in vitro study. Photochem Photobiol 2003; 78(4): 393–399,
  5. Feofanov A., Grichine A., Karmakova T., Kazachkina N., Pecherskih E., Yakubovskaya R., Luќyanets E., Derkacheva V., Egret-Charlier M., Vigny P. Chelation with metal is not essential for antitumor photodynamic activity of sulfonated phthalocyanines. Photochem Photobiol 2002; 75(5): 527–533,
  6. Tong Z., Singh G., Rainbow A.J. Sustained activation of the extracellular signal-regulated kinase pathway protects cells from photofrin-mediated photodynamic therapy. Cancer Res 2002; 62(19): 5528–5535.
  7. Gijsens A., Derycke A., Missiaen L., De Vos D., Huwyler J., Eberle A., de Witte P. Targeting of the photocytotoxic compound AlPcS4 to HeLa cells by transferrin conjugated peg-liposomes. Int J Cancer 2002; 101(1): 78–85,
  8. Zhukova O.S. In vitro models for scrinning of antitumor compounds of different nature. Rossiyskiy bioterapevticheskiy zhurnal 2004; 3(3): 12–18.
  9. Berg K., Moan J. Lysosomes as photochemical targets. Int J Cancer 1994; 59(6): 814–822.
  10. Shirmanova M.V., Balalaeva I.V., Lekanova N.Yu., Mysyagin S.A., Brilkina A.A., Klapshina L.G., Zagaynova E.V. Development of a new photosensitizer based on ytterbium porphyrazine complex. Biofizika 2011; 56(6): 1117–1124.
  11. Wang X., Wang P., Tong W., Liu Q. Comparison of pharmacokinetics, intracellular localizations and sonodynamic efficacy of endogenous and exogenous protoporphyrin IX in sarcoma 180 cells. Ultrasonics 2010; 50(8): 803–810,
  12. Chan W.S., Marshall J.F., Svensen R., Bedwell J., Hart I.R. Effect of sulfonation on the cell and tissue distribution of the photosensitizer aluminum phthalocyanine. Cancer Res 1990; 50(15): 4533–4538.
  13. Juzenas P., Juzeniene A., Rotomskis R., Moan J. Spectroscopic evidence of monomeric aluminium phthalocyanine tetrasulphonate in aqueous solutions. J Photochem Photobiol B 2004; 75(1–2): 107–110,
  14. Sakamoto K., Ohno-Okumura E. Syntheses and functional properties of phthalocyanines. Materials 2009; 2(3): 1127–1179,
  15. Meerovich I.G., Jerdeva V.V., Meerovich G.A., Derkacheva V.M., Savitsky A.P. High-throughput screening system for the study of phototoxicity of photosensitizers in vitro. SPIE Proc 2003; 4952: 203–208,
  16. Stranadko E.F., Armichev A.V., Geynits A.V. Light sources for photodynamic therapy. Lazernaya meditsina 2011; 15(3): 63–69.
  17. Mang T.S. Lasers and light sources for PDT: past, present and future. Photodiagnosis and Photodynamic Therapy 2004; 1(1): 43–48.
  18. Stranadko E.F., Yashunskiy D.V., Khatuntseva E.A., Ustyuzhanina N.E., Ryabov M.V., Ibragimov T.M., Nifant’ev N.E., Gosh R. Looking for new photosensitizers with excitation wavelength in a long-wavelength spectrum. Lazernaya meditsina 2009; 13(1): 29–34.
  19. Theodossiou T., MacRobert A.J. Comparison of the photodynamic effect of exogenous photoprotoporphyrin and protoporphyrin IX on PAM 212 murine keratinocytes. Photochem Photobiol 2002; 76(5): 530–537,
  20. Butler M.C., Itotia P.N., Sullivan J.M. A high-throughput biophotonics instrument to screen for novel ocular photosensitizing therapeutic agents. Invest Ophthalmol Vis Sci 2010; 51(5): 2705–2720,
  21. Zheng G., Chen J., Stefflova K., Jarvi M., Li H., Wilson B.C. Photodynamic molecular beacon as an activatable photosensitizer based on protease-controlled singlet oxygen quenching and activation. Proc Natl Acad Sci USA 2007; 104(21): 8989–8994,
  22. Bachor R., Shea C.R., Gillies R., Hasan T. Photosensitized destruction of human bladder carcinoma cells treated with chlorin e6-conjugated microspheres. Proc Natl Acad Sci USA 1991; 88(4): 1580–1584.
  23. Pogue B.W., Ortel B., Chen N., Redmond R.W., Hasan T. A photobiological and photophysical-based study of phototoxicity of two chlorins. Cancer Res 2001; 61(2): 717–724.
  24. Vrouenraets M.B., Visser G.W.M., Stigter M., Oppelaar H., Snow G.B., van Dongen G.A.M.S. Targeting of aluminum (III) phthalocyanine tetrasulfonate by use of internalizing monoclonal antibodies. Cancer Res 2001; 61(5); 1970–1975.
  25. Song K., Li J., Li L., Zhang P., Geng F., Dong R., Yang Q., Qu X., Kong B. Intracellular metabolism, subcellular localization and phototoxicity of HMME/HB in ovarian cancer cells. Anticancer Res 2011; 31(10): 3229–3235.
  26. Kotel’nikov A.I., Rybkin A.Yu., Goryachev N.S., Belik A.Yu., Kornev A.B., Troshin P.A. Photodynamic activity of hybrid nanostructure based on polycationic derivative of fullerene and phthalocyanine dye Photosense. Doklady Akademii nauk 2013; 452(4): 408–412.
  27. Yakimansky A.V., Meleshko T.K., Ilgach D.M., Bauman M.A., Anan’eva T.D., Klapshina L.G., Lermontova S.A., Balalaeva I.V., Douglas W.E. Novel regular polyimide-graft-(polymethacrylic acid) brushes: synthesis and possible applications as nanocontainers of cyanoporphyrazine agents for photodynamic therapy. Journal of Polymer Science Part A: Polymer Chemistry 2013; 51(20): 4267–4281,
  28. Freshni R. Kultura zhivotnykh kletok [Living cell culture]. Moscow: Binom. Laboratoriya znaniy; 2010. 691 p.
Shilyagina N.Y., Plekhanov V.I., Shkunov I.V., Shilyagin P.А., Dubasova L.V., Brilkina А.А., Sokolova E.A., Turchin I.V., Balalaeva I.V. LED Light Source for in vitro Study of Photosensitizing Agents for Photodynamic Therapy. Sovremennye tehnologii v medicine 2014; 6(2): 15–24

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