Novel methodology for predicting photogenotoxic risk of pharmaceutical substances based on reactive oxygen species (ROS) and DNA-binding assay.
Drug-induced phototoxic skin responses involves photoirritation, photoallergy, and photogenotoxicity. In the present study, we attempted to develop effective analytical tools to predict the photogenotoxic potential of pharmaceutical substances on the basis of their photobiological and DNA-binding properties. Photocleavage of plasmid DNA was assessed by capillary gel electrophoresis (CGE), and generation of reactive oxygen species (ROS) from photoirradiated photosensitizers was also assessed by spectrophotometrical determination. The affinity of drugs with DNA was determined by fluorescent titration or competitive binding assay using ethidium bromide. Structural transition from supercoiled form to open circular form of plasmid DNA were observed in the presence of nine irradiated-photosensitizers, however DNA damage by six phototoxic and five nonphototoxic drugs were negligible or weak even though some chemicals exhibited significant generation of ROS. DNA photocleavers tended to interact with DNA, binding constants of which were found to be less than 1 microM, whereas most nonphotogenotoxic chemicals failed to bind with DNA. The differences in DNA-binding properties might be attributed to a data discrepancy between ROS-generation and DNA photocleaving activity. The combination use of ROS assay and DNA-binding assay could be fast and reliable prediction for the photogenotoxic potential of a large number of drug candidates at the drug-discovery stage.
Onoue
Satomi
S
Department of Pharmacokinetics and Pharmacodynamics and Global Center of Excellence (COE) Program, School of Pharmaceutical Sciences, University of Shizuoka, Shizuoka 422-8526, Japan. onoue@u-shizuoka-ken.ac.jp
Seto
Yoshiki
Y
Oishi
Ami
A
Yamada
Shizuo
S
D002942:Circular Dichroism; D004247:DNA / Q000737:chemistry / Q000378:metabolism* / Q000528:radiation effects; D004249:DNA Damage; D004278:DNA, Superhelical / Q000737:chemistry / Q000187:drug effects / Q000528:radiation effects; D017484:Dermatitis, Phototoxic; D064420:Drug-Related Side Effects and Adverse Reactions; D019075:Electrophoresis, Capillary; D004996:Ethidium / Q000737:chemistry; D008027:Light; D009153:Mutagens / Q000737:chemistry*; D017319:Photosensitizing Agents / Q000737:chemistry / Q000633:toxicity*; D010957:Plasmids / Q000737:chemistry; D011237:Predictive Value of Tests; D017382:Reactive Oxygen Species / Q000737:chemistry*; D013050:Spectrometry, Fluorescence
D004278:DNA, Superhelical; D009153:Mutagens; D017319:Photosensitizing Agents; D017382:Reactive Oxygen Species; D004247:DNA; D004996:Ethidium