The plan of research in this proposal is aimed: (1) at elucidating the significance of somatic mutations affecting the structure of c-myc genes and quantitative abnormalities of c-myc gene expression commonly found in tumors; and (2) at studying the role of different oncogenes in malignant transformation of various types of normal cells in vitro and in carcinogenesis in vivo. To address the first question, a co-transformation assay has been adopted in which the EJ ras oncogene and various cloned human and chicken myc genes are co-transfected into rat embryo cells, with the appearance of foci indicating the presence of a co-transformationally active myc gene. Using this assay, it is possible to determine whether truncation of the c-myc genes and nucleotide alterations present in the genes isolated from tumors are important for myc co-transformational activity. Normal c-myc genes, as well as tumor-derived myc alleles that are inactive in this assay in their native form, will be placed under the transcriptional control of retroviral long terminal repeats to see whether enhanced transcription of these genes renders them co-transformationally active. By placing the various myc alleles under the control of inducible promoters, such as the human metallothionein promoter and the long terminal repeat of mouse mammary tumor virus, it will be possible to correlate the level of expression of various mutant and normal myc genes with co-transformational activity. An approach to the second major aim requires the development of replication-defective murine retroviral vectors which can deliver various oncogenes to many different types of murine and rat cells at high efficiency. Such vectors, derived from Moloney murine leukemia virus, will be used to introduce myc, ras, B-lym, and other oncogenes into normal and established lymphoid, hematopoietic, and fibroblastic cells. The impact of the presence of active oncogenes in these cells will be studied in vitro and after introduction of the cells into host animals. Particular attention will be paid to: (1) the development of the transformed phenotype and tumors, both immediately and after the introduction of additional oncogenes; (2) any alterations in the activity of host cellular genes with the introduction of different oncogenes; (3) the tissue specificity of the effects of the various oncogenes; and (4) the functional and differentiated characteristics of the cells transformed by the oncogene-bearing retroviruses. These experiments will explore and attempt to recreate the multistep path towards the development of malignancies of various types of tissues in animals. In the process, a better definition of the role played by different oncogenes will be obtained, as well as a better understanding of the """"""""pre-malignant"""""""" state that exists prior to the development of overt malignancy. (X)
