The overall emphasis will be to investigate the role of HBV and WHV in hepatic oncogenesis. The significance and specificity of chromosome aberrations and Insulin-like growth factor II activation associated with HCC in hepadna virus carriers will be investigated. Cell culture assays and """"""""in vivo"""""""" hepatocarcinogenesis studies in woodchucks will test mechanisms of HBV and WHV induced carcinogenesis. Specific studies to be conducted include: 1. Analysis of loss or amplification of alleles from specific chromosomes in primary human HCC. In this study we will determine (a) the extent and frequency of deletions in chromosome llp in HCCs using the loss of restriction fragment length polymorphisms (RFLPs) as an assay for loss of alleles, (b) the frequency of loss or amplification of alleles in chromosomes other than 11 and (c) the copy number, hemizygous, homozygous (reduplicated) or amplified, of the polymorphic alleles remaining after loss of their homologue and the relationship (if any) of HBV integration to loss of alleles on chromosomes containing these integrations. 2. Determine the frequency, specificity and mechanisms of transcriptional activation of IGF-II in primary woodchuck HCCs. The abundance and sizes aof IGFD-II RNAs intissues at stages of hepatic oncogenesis, inwoodchucks will be studied along with the specificity of IGF-II activation, and the mechanism of IGF-II transcription by sequencing cDNA clones of IGF-II mRNAs from woodchuck HCCs. 3. Analyze the activity of integrated HBV and WHY DNA using biological assays for tumorigenesis and transcription. Cell lines contain HBV and WHV constructs will be used to address the following questions (a) can HBV and WHV function in a """"""""Hit and Run"""""""" tumorigenesis mechanism and do they affect the mutation rate of cellular genes and (b) how do HBV integrations containing enhancer elements affect transcription of cellular genes flanking HBV. 4. Experimental manipulation of HCC in woodchucks and isolation of woodchuck hepatocyte cell lines. Partial hepatectomy and CC14 and phenobarbitol treatments will be used to manipulate HCC in woodchucks. Preneoplastic nodules and HCCs will be analyzed for WHV integration and IGF-II activation to study events proceeding HCC. Woodchuck hepatocyte cell cultures will be initiated using newly developed methods of Kitagawa to study WHV replication and progression to HCC.
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