Comparative studies of alkylating and N-heterocyclic compounds on different genetic endpoints with special emphasis on amplification of minisatellite sequences

Detta är en avhandling från Stockholm : Stockholm University

Sammanfattning: Several mutational events are involved in the multistage process of cancer. Genetic instability of tumor DNA is shown in a variety of genetic changes such as point mutations, chromosomal aberrations, aneuploidy, DNA amplification and somatic recombination. Highly repetitive sequences, minisatellite DNA, have been changed in tumors, most probably by DNA amplification.To better predict carcinogens all these genetic changes should be considered. The present investigation has focused on the study of genetic changes especially DNA amplification with the use of genetic instability of minisatellites. DNA alterations in human bladder tumors were examined. A new in vitro test system was developed to study DNA amplification.In this study DNA fingerprint analysis, with the use of minisatellite DNA, of 22 bladder tumor patients is performed. DNA alterations were determined in approximately 50% of the examined bladder tumor patients. In eight of these ten patients a loss of bands was demonstrated.To study DNA amplifications an in vitro test system was developed. A haploid yeast strain was constructed, the TR(MS1)-1, which carries a chromosomal integration of the human minisatellite sequence MSI. The spontaneous frequency of new MSI length alleles was approximately 30%. Both amplifications and deamplifications can be detected in this system without selection. A plasmid pop-out frequency can also be meausured.A number of model compounds was used during this study, namely three alkylating agents, a nongenotoxic carcinogen and several nitrogen heterocyclic compounds. To compare genotoxicity and the mechanism of action, these compounds are tested in several in vitro test systems. Ethylene oxide (EO) was demonstrated to be more potent than propylene oxide (PO) in several of the measured endpoints in yeast. EO induced changes in the amplification spectrum of new MSI length alleles in TR(MS1)-1 while PO increased the plasmid pop-out frequency. 2,3,7,8-Tetrachlorodibenzo(p)dioxin (TCDD) was also seen to induce changes in the amplification spectrum of new MSI length alleles, thus demonstrating an effect of TCDD at the DNA level. Among the N-heterocyclic compounds, camptothecin increased the plasmid pop-out frequency in TR(MS1)-1. Two tryptophan photoproducts, "284" and "312", were shown to bind to the A A-receptor. They were demonstrated to be nongenotoxic and antimutagenic in bacteria and slightly genotoxic in yeast. These two compounds are antimutagenic by inhibiting the cytochrome P450IA1. "284" increased cell survival or cell division. These two tryptophan photoproducts, ”284" and "312", are postulated to be biological signal substances by being the endogenous ligand to the Ah -receptor.