Pg. Johnson et Ta. Beerman, DAMAGE-INDUCED IN EPISOMAL EBV DNA IN RAJI CELLS BY ANTITUMOR DRUGS AS MEASURED BY PULSED-FIELD GEL-ELECTROPHORESIS, Analytical biochemistry, 220(1), 1994, pp. 103-114
The studies described below were carried out to analyze the damage ind
uced by DNA active drugs to episomal (Epstein-Barr virus, EBV) DNA in
the Raji Burkitt's lymphoma cell line. This work: (i) applies pulsed-f
ield gel electrophoresis (PFGE) techniques to quantify DNA damage on a
large (approximate to 180 kbp), circular target, (ii) investigates th
e DNA strand-scission behavior of different classes of drugs on the EB
V episome, and (iii) compares EBV episomal damage to that generated in
genomic DNA in the Raji cell line. Cells were treated with ionizing r
adiation to induce random strand scission, and the migration of topolo
gical forms of EBV was measured using PFGE. DNA damage induced in the
episome by DNA active drugs was then assayed. Three drugs, acting by d
ifferent types of DNA interactive mechanisms, were used: bleomycin, an
intercalative DNA strand-scission agent; and amsacrine (mAMSA) and te
niposide (VM26), intercalative and nonintercalative topoisomerase II a
ctive drugs, respectively. Rad equivalency of damage was determined by
comparing the drug-induced change in percentage of Forms I and III to
that generated by ionizing radiation. Additionally, single- and doubl
e-strand scission induced in genomic (total cellular) DNA by X-rays, b
leomycin, amsacrine, and teniposide were assayed by high-sensitivity a
lkaline and neutral filter elution techniques. We demonstrate that pul
sed-field gel electrophoresis is a useful technique for measuring form
conversion in large episomal DNA. While all three drugs effect both e
pisomal and genomic DNA strand scission, bleomycin appears to preferen
tially damage the EBV episome. The topoisomerase II active drugs mAMSA
and VM26 show no evidence of episome-directed damage in this system a
nd, in fact, damage genomic DNA at somewhat higher rates. (C) 1994 Aca
demic Press, Inc.