CELL AND MOLECULAR BIOLOGY OF RAT HEPATOCARCINOGENESIS

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This research program is directed toward an understanding of the regulation of genetic expression in normal, preneoplastic, and neoplastic liver in the rat. The approach utilized will be to study the fine structure of regulation of several genes whose expression in preneoplastic and/or neoplastic liver is significantly different from normal, to characterize more completely the stage of progression, and to characterize hepatocarcinogenesis in a transgenic model as compared to multi-stage carcinogenesis induced by chemicals in the rat. The structure and function of the regulatory components of the serine dehydratase (SDH), aldehyde dehydrogenase (Type 3) (ALDH) and gamma- glutamyl transpeptidase (GGT) genes, all of whose expression is abnormal during hepatocarcinogenesis, will be characterized. Characterization will include the determination of the specific nucleotide sequences involved in the regulation of the expression of these genes in vivo, the identification and characterization of protein(s) that bind to the regulatory sequences of these genes, the levels of such proteins in nuclei of normal, preneoplastic and malignant hepatocytes, and functioning of the regulatory sequences of these genes in their expression during preneoplasia and overt neoplasia. Utilizing a model developed in this laboratory for the study of multi- stage hepatocarcinogenesis in rat liver, we plan to develop better methods to quantitate the development of the stage of progression in hepatocarcinogenesis, to characterize this stage at both the cellular and molecular levels and to develop quantitative methods for the identification of progressor agents. Characterization of the stage will involve a study of the regulation of the expression of several known and potential "marker" genes utilizing cytochemical methods. Chromosomal abnormalities which develop early during the formation of hepatocellular carcinoma in the rat in the stage of progression and the specificity of such abnormalities for the evolution of this stage will be determined. Potential tumor suppressor genes, especially the p53 gene, will be characterized as to their chromosomal localization and any mutation in their sequences, the latter initially in relation to the p53 gene. We will also use a colony of transgenic rats carrying the SV40 T antigen transgene under the control of the mouse albumin promoter/enhancer to compare genetically programmed hepatocarcinogenesis with chemically induced hepatocarcinogenesis. The stages of initiation, promotion, and progression will be delineated in the transgenic model and compared with these stages occurring during the chemical induction of hepatocarcinogenesis. The effect of environmental factors including hormones, diet, and xenobiotics on the natural history of genetically programmed hepatocarcinogenesis will be determined. Characterization of both neoplastic and morphologically normal hepatocytes from the transgenic animals will be carried out in vitro. Together these studies should enable us to define more accurately the critical genetic characteristics of multi-stage hepatocarcinogenesis in the rat, especially during the stage of progression, and to gain a better understanding of the mechanisms of the abnormalities in genetic expression characteristic of the stages of promotion and progression.