MUTAGENESIS OF THE K-RAS PROTOONCOGENE IN MOUSE LUNG-TUMORS INDUCED BY N-ETHYL-N-NITROSOUREA OR N-NITROSODIETHYLAMINE

The role of ras gene activation in the development of lung tumors induced by N-ethyl-N-nitrosourea (ENU) and N-nitrosodiethylamine (DEN) was evaluated in the A/J mouse, a strain susceptible to chemically induced lung tumors. DNAs isolated from both ENU- and DEN-induced lung tumors were screened for activating mutations in the K-ras gene by utilizing the polymerase chain reaction (PCR) and direct sequence analysis. Mutations in the K-ras gene were detected in 11 of 11 ENU-induced tumors and 23 of 28 DEN-induced tumors. In ENU-induced tumors, there were three GC --> AT transitions in the second base of codon 12, and seven AT --> GC transitions and one AT --> TA transversion in the second base of codon 61. A similar spectrum of K-ras mutations was observed in DEN-induced lung tumors: five GC --> AT transitions and two GC --> TA transversions in the second base of codon 12, and sixteen AT --> GC transitions at the second base of codon 61. Ninety-one percent (31/34) of the observed mutations are consistent with the formation of the promutagenic O4-ethylthymine and O6-ethylguanine adducts in DNA. Therefore, lung tumors from the A/J mouse induced by DEN and ENU could be initiated by the interaction of reactive metabolites with specific sites in the K-ras gene. This is the first clear example of activation of the K-ras gene by ethylating agents in a rodent lung tumor system.