Monitoring the (photo)genotoxicity of photosensitizer drugs: direct quantitation of single-strand breaks in deoxyribonucleic acid using an oligonucleotide chip.
Oligonucleotide chip-based assays can be a sample-thrifty, time-saving, routine tool for evaluation of chemical-induced DNA strand breaks. This article describes a novel approach using an oligonucleotide chip to determine photosensitizer-induced DNA single-strand breaks. Surface coverage of fluorophore-labeled oligonucleotides on silicon dioxide chip surfaces was determined on alkaline phosphatase digestion. Fluorescence maxima (at 520 nm) of the solutions were converted to molar concentrations of the fluorescein-modified oligonucleotide by interpolation from a predetermined standard linear calibration curve. The photosensitizing activity of chlorpromazine and triflupromazine toward DNA single-strand breaks was then studied at different drug doses and also as a function of photoirradiation time. Photoinduced single-strand breaks calculated using the method described here agreed with values predicted by theoretical extrapolation of the single-strand breaks obtained for plasmid DNAs from agarose gel electrophoresis, and thereby indirectly validated the chip-based assays. Under UV irradiation (>or=93.6 kJ/m2) chlorpromazine (>or=0.08 mM) was found to have significant photogenotoxicity. However, triflupromazine did not exhibit any (photo)genotoxicity over the concentration range studied (0.04-0.20mM). The method developed will be useful for quantitative screening of drug genotoxicity in terms of induction of breaks in DNA.
Kim
Min Jung
MJ
Research Institute of Pharmaceutical Sciences and College of Pharmacy, Seoul National University, 599 Gwanangno, Gwanak-gu, Seoul, Republic of Korea.
Pal
Sukdeb
S
Naoghare
Pravin K
PK
Song
Joon Myong
JM
D001483:Base Sequence; D002746:Chlorpromazine / Q000633:toxicity; D004247:DNA / Q000235:genetics / Q000378:metabolism*; D053904:DNA Breaks, Single-Stranded / Q000187:drug effects* / Q000528:radiation effects; D004305:Dose-Response Relationship, Drug; D004307:Dose-Response Relationship, Radiation; D009153:Mutagens / Q000633:toxicity*; D020411:Oligonucleotide Array Sequence Analysis; D017319:Photosensitizing Agents / Q000633:toxicity*; D013499:Surface Properties; D013997:Time Factors; D014273:Triflupromazine / Q000633:toxicity; D014466:Ultraviolet Rays
D009153:Mutagens; D017319:Photosensitizing Agents; D004247:DNA; D014273:Triflupromazine; D002746:Chlorpromazine