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Exploitation of the Dose/Time-Response Relationship for a New Measure of DNA Repari in the Single-Cell Gel Electrophoresis (Comet) Assay  

Kim, Byung-Soo (Department of Applied Statistics, Yonsei University)
Edler, Lutz (Biostatistics Unit R0700, German Cancer Research Center)
Park, Jin-Joo (Division of Interaction of Carcinogens with Biological Macromolecules, German Cancer Research Center)
Fournier, Dietrich Von (Gynecological Radiology, Radiological Clinic, University of Heidelberg)
Haase, Wulf (Clinic of Radiotherapy and Radiological Oncology, St. Vincentius Clinics)
Sautter-Bihl, Mare-Luise (Clinic of Radiotherapy, Municipal Clinics gGmbH Karlsruhe)
Hagmuller, Egbert (Surgical Department of the Heilbronn Clinics)
Gotzes, Florian (Division of Interaction of Carcinogens with Biological Macromolecules, German Cancer Research Center)
Thielmann, Heinz Walter (Division of Interaction of Carcinogens with Biological Macromolecules, German Cancer Research Center)
Publication Information
Toxicological Research / v.20, no.2, 2004 , pp. 89-100 More about this Journal
Abstract
The comet assay (also called the single-cell gel electrophoresis assay) has been widely used for detecting DNA damage and repair in individual cells. Since the conventional methods of evaluating comet assay data using frequency statistics are unsatisfactory we developed a new quantitative measure of DNA damage/repair that is based on all information residing in the dose/time-response curves of a comet experiment. Blood samples were taken from 25 breast cancer patients before undergoing radiotherapy. The comet assay was performed under alkaline conditions using isolated lymphocytes. Tail DNA, tail length, tail moment and tail inertia of the comet were measured for each patient at four doses of $\gamma$-rays (0, 2, 4 and 8 Gy) and at four time points after irradiation (0, 10, 20 and 30 min) using 100 cells each. The resulting three-dimensional dose-time response surface was modeled by multiple regression, and the second derivative, termed 2D, on dose and time was determined. A software module was programmed in SAS/AF to compute 2D values. We applied the new method successfully to data obtained from cancer patients to be assessed for their radiation sensitivity. We computed the 2D values for the four damage measures, i.e., tail moment, tail length, tail DNA and tail inertia, and examined the pairwise correlation coefficients of 2D both on the log scale and the unlogged scale. 2D values based on tail moment and tail DNA showed a high correlation and, therefore, these two damage measures can be used interchangeably as far as DNA repair is concerned. 2D values based on tail inertia have a correlation profile different from the other 2D values which may reflect different facets of DNA damage/repair. Using the dose-time response surface, other statistical models, e.g., the proportional hazards model, become applicable for data analysis. The 2D approach can be applied to all DNA repair measures, Le., tail moment, tail length, tail DNA and tail inertia, and appears to be superior to conventional evaluation methods as it integrates all data of the dose/time-response curves of a comet assay.
Keywords
Quantitation of comet assay data; Dose-time-response surface; Second derivative; $\gamma$-radiation; Radiation sensitivity of cancer patients; Human lymphocytes.;
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