Cellular photodestruction induced by hypericin in AY-27 rat bladder carcinoma cells

In a recent clinical study we showed that hypericin accumulates selectively in urothelial lesions following intravesical administration of the compoun d to patients. In the present study the efficacy of hypericin as a photoche motherapeutic tool against urinary bladder carcinoma was investigated using the AY-27 cells (chemically induced rat bladder carcinoma cells). The upta ke of hypericin by the cells increased by prolonging the incubation time an d increasing the extracellular hypericin concentration. Photodynamic treatm ent of the cells incubated with 0.8 and 1.6 muM hypericin concentrations re sulted in remarkable cytotoxic effects the extent of which depended on the fluence rates. Photoactivation of 1.6 muM hypericin by 0.5, 1.0 or 2.0 mW/c m(2) for 15 min resulted in 3, 30 and 95% of the antiproliferative effect, respectively. Increasing the photoactivating light dose from 0.45 to 3.6 J/ cm(2) resulted in a five-fold increase in hypericin photodynamic activity. Irrespective of the fluence rates and irradiation times incubation of the c ells with 10 muM hypericin induced rapid and extensive cell death in all co nditions. The type of cell death (apoptosis or necrosis) induced by photoac tivated hypericin depended largely on the hypericin concentration and the p ostirradiation time. At lower hypericin concentrations and shorter postirra diation times apoptosis was the prominent mode of cell death; increasing th e hypericin concentration and/or prolonging the postirradiation time result ed in increased necrotic cell death. Cell pretreatment with the singlet oxy gen quencher histidine, but not with the free-radical quenchers, significan tly protected the cells from photoactivated hypericin-induced apoptosis, at least when a relatively low concentration (1.25 muM) was used. This result suggests the involvement of a Type-II photosensitization pro-cess. However , cells treated with higher hypericin concentrations (2.5-5 muM) were inade quately protected by histidine. Since hypericin is thus shown to be a poten t and efficient photosensitizer, and since the conditions used were the sam e as when hypericin is used clinically to locate early-stage urothelial car cinoma lesions, hypericin may well become very important for the photodynam ic treatment of superficial bladder carcinoma.