Analysis of DNA modifications

Abstract

Approximately 90% of the chemicals considered carcinogenic in humans lead, in animal experiments, either spontaneously or after metabolic activation, to covalently bound modifications of the bases of DNA, so called DNA adducts [1]. DNA adducts are the precursors of mutations, because they or their immediate neighbors are possible sites for modifications (substitutions or deletion of bases and frameshift) during replication. Such damage occurs only at very low concentrations in vivo, however, and is largely eliminated by DNA-repair processes. Despite these effective repair systems, ten to one thousand adducts can normally be found in a cell. Thus the adduct concentration in a diploid cell with approximately 6×10 nucleotides is in the range of one adduct in 10 to 10 unmodified nucleotides [2]. Knowing to what extent a chemical forms DNA adducts can contribute important information about causes of cancer and the mechanism of its development. DNA adducts are therefore an internal and individual dosimeter of the exposure of an organism to genotoxic compounds and ultimately describe the biologically effective dose of a DNA-damaging chemical. They are thus suitable biomarkers of exposure, so that they become a focus of molecular epidemiological studies for estimating the cancer risks arising from lifestyle [3]. An applicable method for analyzing DNA adducts must satisfy several requirements. First, it must be able to detect DNA adducts in a realistic concentration range (femtomolar range, one adduct in 10 to 10 unmodified nucleotides) with great precision. Second, it should guarantee simultaneous determination of greatly varied adduct classes with a high sample throughput. No existing method fulfils these requirements. It therefore seems useful to classify DNA modifications into two groups, exogenous and endogenous DNA adducts. Exogenous modifications are often bulky DNA adducts (modification by aromatic or polycyclic aromatic metabolites) and their concentration range is one adduct in 10 to 10 unmodified nucleotides. Endogenous DNA adducts are formed by small modifications, for example by oxygen radicals, and are normally found at concentrations of one adduct in 10 to 10 unmodified nucleotides. Important methods have been used to determine exogenous and endogenous DNA adducts.