INCIDENCE OF HARVEY RAS ONCOGENE POINT MUTATIONS AND THEIR EXPRESSION IN METHYLBENZYLNITROSAMINE-INDUCED ESOPHAGEAL TUMORIGENESIS

Activation of the ras oncogene was investigated in esophageal tumors induced by methylbenzylnitrosamine (MBN) in the Sprague - Dawley rat. DNA was extracted from grossly visible carcinogen-induced tumors. H-ras and K-ras gene sequences were then amplified by the polymerase chain reaction. Point mutations in the ras genes were then identified by selective hybridization to allele-specific oligonucleotide probes. A guanine to adenine transition at the 35th nucleotide in the H-ras coding sequence (GGA to GAA in the 12 codon) was observed in 67% (10 of 15) of the papillomas examined. This mutation codes for glutamate instead of glycine as the 12th amino acid of the ras p21 protein. No other H-ras or K-ras mutations were observed. To determine the distribution of this H-ras mutation in esophageal tissues, histological sections of MBN-treated esophagi were stained with a monoclonal antibody (E184) which selectively recognizes the mutated ras p-21 with glutamate substituted for glycine as the 12th amino acid. Expression of the mutant ras p21 protein was observed in 20% of the squamous papillomas, 13.6% of hyperplastic lesions and 10% of dysplastic lesions. Thus, activation of the H-ras oncogene as a result of guanine to adenine point mutation is a frequent event in esophageal tumors induced by MBN, occurring in 67% of squamous papillomas, but expression of the corresponding mutant ras p21 protein is observed in a much smaller proportion of the tumors in this animal model.