The objective of this project is the identification of recurrent genetic alterations that are relevant to neoplastic development and to provide markers for early detection and prognostic assessment of cancer, particularly in solid tumors (project Site Visit July 1997). This objective is consistent with the major initiative launched by the NCI to decipher the molecular anatomy of various cancers. The research focuses primarily on human- and mouse-induced hepatocellular carcinoma (HCC) that were investigated using a battery of the most advanced molecular cytogenetic procedures. In human HCC, by comparative genomic hybridization (CGH), DNA copy-number imbalances involving multiple sites were localized. Seventeen genetic imbalances are novel in HCC, thus extending significantly the map of genetic changes and providing a starting point for the isolation of new genes relevant in pathogenesis of liver neoplasia, as well as providing molecular probes for both diagnosis and monitoring treatment of the disease. The analysis of individual and combined CGH profiles based on hepatitis B-virus status and the geographic origin of the primary tumors showed a similar pattern of chromosome imbalances. In mouse HCC from transgenic mice overexpressing c-myc and transforming growth factor-alpha, nonrandom chromosomal alterations were identified among which a translocation involving chromosomes 5 and 6 is the first balanced alteration reported in mouse or human HCC. The breakpoint of this translocation is located near a tumor susceptibility gene and the site of myc transgene integration. More particularly, preferential sites of genetic changes were identified as the breakpoints in recurrent stable rearrangements of several chromosomes involved the sites of damage in early dysplastic liver lesions. Overall, the identification of novel recurrent genomic alterations and genes relevant to the development of HCC provides new insight in the molecular basis of this neoplasia and serve as a framework for future investigations toward the completion of genomic anatomy of liver cancer. Combined CGH and spectral karyotype is the most powerful approach for detection of genomic imbalances and structural reorganization across the entire tumor genome. This approach was applied to elucidate the disputed origin of clonally abnormal chromosomes in cervical carcinoma HeLa cells. The complete derivation of all complex abnormalities underlines the current level resolution of molecular cancer chromosome analysis. Furthermore, the detection of dispersed copies of papillomavirus-18 at the sites of cryptic translocations undetectable by other techniques demonstrated that viral integration triggered genomic instability, a critical step in tumor development. Two newly discovered recurrent unbalanced translocations identified in our studies of human HCC involved regions of genomic fragility and recombination. Fragile histidine triad (FHIT), a tumor suppressor gene spanning the region 3p14.2 of high fragility, is frequently altered in several cancers, particularly those of epithelial cell origin and associated with known carcinogenic agents. HCC is closely related with carcinogenic agents such as hepatitis B and C virus infections, dietary aflatoxin, alcohol consumption, and exposure to chemical carcinogens. We identified chromosome rearrangements of the short arm of chromosome 3 in 61% of the HCC cell lines, a decreased or absent FHIT mRNA expression in 66% of the lines, and intragenic deletions and absence of protein expression in 50% of primary tumors, indicating that FHIT alterations are pathologically relevant to the development of HCC. Several new genes isolated in our and other laboratories were chromosomally localized by FISH and include: human GFR alpha 3, Stat 6 cyclin I, palmitol-protein, thioesterase genes and mouse arylalkylamine N-acetyltransferase (AANAT), cyclin G2, cyclin I, and Bog genes. In addition, a repetitive sequence belonging to a subfamily of satellite 1 DNA specific sequences was isolated from human cells transformed with SV40 virus and was assigned uniquely to the centromere of chromosome 4. These localizations will facilitate the identification of loci in genetic diseases of unknown etiology mapping at the same chromosomal regions and their involvement in neoplasia. - Chromosome alterations, Comparative genomic hybridization, Deletion translocations, Fluorescence in situ hybridization,
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