Exocyclic base adducts in human DNA such as the etheno (epsilon) adducts epsilon dA and epsilon dC are formed by environmental and endogenous genotoxins that are generated from oxidative stress, dietary factors and metal storage diseases. A third epsilon-adduct, epsilon dG, has been detected in rodent tissues upon exposure to epsilon-adduct forming chemicals. To assess the role of these promutagenic DNA lesions in human malignant diseases, four representatives of epsilon-adduct generating chemicals (trans-4-hydroxy-2-nonenal,2-chloro-acetaldehyde, chloro-ethylene oxide and vinyl carbamate, a reactive metabolise of urethane) will be investigated by the following approaches:
(i) Quantification of epsilon dA and epsilon dC in cell/tissue DNA of humans, mice, Drosophila and E. cold by ultrasensitive methods, recently developed by study partners; development of a new sensitive method for analysis of epsilon dG in human DNA.
(ii) Studies on diet-related formation of epsilon-adducts in rodents, and the proportionality between epsilon-adducts in DNA of white blood cells vs. those in breast and colon; comparison of DNA epsilon-adduct levels and repair activities in lung tissues and lymphocytes from lung cancer patients.
(iii) Identification and purification of repair enzymes that remove of epsilon dG and epsilon dC in human cells and in E. coli; construction of E. cold repair-deficient strains for defining mutation specificity of epsilon bases;
correlation of the persistence/repair of epsilon-bases with biological endpoints (free radical induced cytotoxicity; cancer) measured in knock-out mice deficient in repair (ADPRT) and tumor supressor (p53) genes.
(iv) Development of methods for analyzing formation/repair of DNA epsilon-adducts at specific target gene (p53) and at nucleotide resolution level.
(v) Dosimetry-based comparisons, particularly at low doses, of SCE's, chromosome aberrations, micronuclei and hprt-mutations in
human hepatoma cells, and of several genetic endpoints in Drosophila; characterization of mutation spectra in hprt (hep-g2) and in vermilion/rosy genes (Drosophila).
(vi) Carcinogenic risk assessment in humans, qualitative and quantitative structure-activity relationships (QSAR's, SAR's).
Funding SchemeCSC - Cost-sharing contracts
2300 RA Leiden