Genetic and molecular mechanisms of the in vitro transformation of Syrian hamster embryo cells by the carcinogen N-ethyl-N-nitrosourea II. Correlation of morphological transformation, enhanced fibrinolytic activity, gene mutations, chromosomal alterations and lethality to specific carcinogen-induced DNA lesions.

The stability of N-ethyl-N-nitrosourea (ENU)-induced DNA damage in Syrian hamster embryo (SHE) cells was determined to study correlations with ENU-induced mutation and transformation. Confluent cultures were treated with ENU and after 0, 3 or 6 days holding in low serum medium to inhibit cell proliferation, the extent of ethylation at different sites in the DNA was determined with h.p.l.c. The amounts of dTp(Et)dT-triester, and O4- and O2-EtThy remained constant during the 6-day period. O6-EtGua slowly decreased (t1/2: 14 days); the initial level was lower than expected from in vitro data, suggesting that Syrian hamster embryo cells contain alkyl transferase. Evidence for active removal was also obtained in the case of 7-EtGua (t1/2: 59 h), O2-EtCyt (t1/2: 96 h) and possibly 3-EtGua (t1/2: 102 h). As expected, the promutagenic O6-EtGua was found to correlate with gene mutations. In addition, however, we have found that the likewise promutagenic O4- and O2-EtThy also correlate with gene mutations. Furthermore, our data suggest that sister chromatid exchange (SCE) and cytotoxicity (clonal survival) have a similar molecular basis. Both correlate with O2-EtCyt and 3-EtGua which are located in the narrow groove and are therefore expected to block DNA replication. Chromatid aberrations and micronuclei could not be correlated to specific DNA lesions, but were found to correlate mainly to N-ethylations. The same holds for morphological transformation, but in this case there is also a small contribution of DNA O-ethylation. In contrast, enhanced fibrinolytic activity did correlate only with stable O-ethylations, including O6-EtGua.