Recent studies have shown that a significant proportion of human tumors of different histological characteristics contain activated oncogenes with potential to morphologically transform normal cultured rodent cell lines. Many of the oncogenes detected by the NIH 3T3 transfection assay belong to the ras gene family. The mechanisms of activation of ras genes in these human tumors have been established as point mutations in two preferential codons in their protein coding exons. Although the role that these activated oncogenes could be playing in the genesis of these tumors has not been determined, it is likely that the mutational events which result in the activation of these genes could be an important step in the multistep process of human carcinogenesis. We will develop diagnostic screening procedures for the detection of mutant ras oncogenes in human tumors, using molecular hybridization techniques. First, mutations at codons 12 and 61 of ras genes which generate restriction endonuclease polymorphisms will be detected by Southern blot experiments using fragments of the previously cloned ras oncogenes as radioactive probes. Second, synthetic oligonucleotides of defined sequence will be used in Southern blot hybridization experiments to detect those mutations at codons 12 and 61 which do not create or destroy restriction endonuclease sites at these positions of ras genes. These studies will also reveal the possible presence of amplified ras oncogenes in human tumors, and could also detect mutant ras genes at positions other than 12 and 61. Third, the possible existence of other types of activation of ras genes in human tumors will be searched by Northern blot experiments of tumor's RNA. Finally, we will try to find a correlation between the presence of these mutant ras oncogenes and the histopathological characteristics and clinical behaviors of the corresponding tumors. These studies should have application as diagnosis and eventually prognosis for the specific tumors containing mutant ras oncogenes. (6)
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