Activation of ras oncogenes and expression of tumor‐specific transplantation antigens in methylcholanthrene‐induced murine fibrosarcomas

Abstract

The DNA of 22 fibrosarcomas, newly induced in BALB/c mice by subcutaneous doses of 3-methylcholanthrene (3-MCA), was tested in NIH 3T3 transformation assay. Activation of K-ras and N-ras was found in 7 and 3 cases respectively. No H-ras activation was detected. Polymerase chain reaction and oligonucleotide hybridization performed on the DNA of the 22 sarcomas revealed 5 cases of K-ras mutation at codon 12, 3 at codon 13 and 1 at both codons. One case of K13 mutation was not detectable by transfection. Three cases of mutation at codon 61 of N-ras were also found, one of which was simultaneous with a K12 mutation. Tumor-specific transplantation antigens (TSTA) were assessed in the 22 original tumors. Altogether 16 sarcomas were immunogenic, with the highest frequency of TSTA+ tumors (10/11 and 5/6) in the groups given 1.0 and 0.1 mg of 3-MCA respectively, the lowest (1/5) in that with 0.01 mg of carcinogen; ras mutations occurred in the DNAs of 11 out of the 16 TSTA+ sarcomas, but none of the DNAs of the 6 TSTA- tumors showed ras mutation. The results suggest that 3-MCA-induced transformation of subcutaneous fibroblasts can involve mutations in codons 12, 13 or 61 of K- and N- but not H-ras gene and that such mutation is accompanied by the expression of TSTA.

Multiple molecular alterations in mouse lung tumors

“…To amplify sequences of Ki-ras across codon 61, the oligonucleotide primers previously described [20] were used. PCR reactions and DNA hybridizations to a panel of 19-mer synthetic oligonucleotide probes specific for the normal codon as well as for the activating mutations of codon 61 [2 1 I were performed as previously de-…”

Mouse embryo fibroblasts transformed by activated ras or dominant‐negative p53 express cross‐reactive tumor rejection antigens

“…Primary M E F were chosen for transfection because of concern that chemically transformed or immortalized parental cells would have a high background of immunogenicity due to extensive mutagenesis. These concerns are supported by the observation of a previous study that TRA were not cross-protective between a series of methylcholanthrene-transformed tumors in BALB/c mice, even though some of them carried identical activating rus point mutations (Carbone et a/., 1991). The lack of cross-reactivity suggests that the TRA of those tumors are the products of randomly mutated genes.…”

“…Indirect evidence comes from the observation that the antigenicity of chemically induced tumors is directly proportional to the dose of the mutagen applied (Prehn, 1975;Prehn and Bartlett, 1987;Carbone et al, 1991). More direct evidence has been obtained from the study of highly immunogenic Tum-(regressor) variants of mastocytoma P815 derived by mutagenesis with N-methy-"nitro-N-nitrosoguanidine (Uyttenhove et al, 1980).…”

“…The molecular basis of the heterogeneity of tumor antigens is not understood, but there is some evidence to support the hypothesis that tumor-specific antigens can be the protein products of genes altered randomly by the chemical or radiation mutagenesis used to induce transformation. Indirect evidence comes from the observation that the antigenicity of chemically induced tumors is directly proportional to the dose of the mutagen applied (Prehn, 1975;Prehn and Bartlett, 1987;Carbone et al, 1991). More direct evidence has been obtained from the study of highly immunogenic Tum-(regressor) variants of mastocytoma P815 derived by mutagenesis with N-methy-"nitro-N-nitrosoguanidine (Uyttenhove et al, 1980).…”

Revisiting immunosurveillance and immunostimulation: Implications for cancer immunotherapy

“…A correlation between the dose of carcinogen administrated and the immunogenicity of the subsequent tumor has been reported in mouse models [15-17]. Since human tumors most likely occur as a result of low exposure to carcinogens, incipient tumors would be expected to be poorly immunogenic.…”