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Glen E. Kisby, Nancy Springer, and Peter S. Spencer
Portland Environmental Hazards Research Center (PEHRC)
Sulfur and nitrogen mustards are reported to produce neurobehavioral
and neuropathological changes in animals and humans. Since DNA damage
is considered the primary mechanism by which sulfur and nitrogen mustards
produce their cytotoxic effects, we developed a rapid and sensitive isocratic
high-performance liquid chromatography method to detect the principal
monofunctional and bifunctional guanine DNA adducts produced by nitrogen
mustards. The monofunctional mechlorethamine (HN2, nitrogen mustard) DNA
adduct N-(2-hydroxyethyl)-N-[(2-(7-guaninyl)-ethyl)]-methylamine (GMOH)
and the bifunctional cyclophosphamide DNA adduct N,N-bis[2-(7-guaninyl)-ethyl]amine
(G-Nor-G) eluted from a C18 column (tr= 2.7 and 2.1 min, respectively)
using a sodium citrate/acetonitrile buffer (pH 6.5). The DNA adducts were
detected electrochemically (EC). The lowest concentrations of GMOH and
G-Nor-G that could be detected were 63 fmol and 3.5 pmol, respectively.
GMOH was detected in calf thymus DNA treated in vitro with HN2 (~5.1 pmol/µg
DNA) and in cultures of rat granule cell neurons (85 fmol/µg DNA)
and SY5Y neuroblastoma cells (42 fmol/µgDNA) treated for 24h with
10 µM HN2. These studies demonstrate that HPLC/EC is a rapid and
sensitive technique for detecting monofunctional and bifunctional nitrogen
mustard DNA adducts in tissues. The findings indicate that HN2 may target
neurons in vivo, possibly through a mechanism involving DNA damage. They
supplement evidence that nitrogen and sulfur mustards exert neurotoxic
effects in animals and humans exposed to large concentrations of these
cytotoxic agents.
"Keywords:" Mustard DNA adducts Nervous tissue
This work was supported by a grant from the Department of
Veterans Affairs to the PEHRC, a joint project of the Portland Veterans
Affairs Medical Center and the Center for Research on Occupational and
Environmental Toxicology, Oregon Health Sciences University. |
