Sharon L. Ricketts, Ph.D.

Dissertation research performed under the direction of William B. Coleman

ABSTRACT
    Several distinct regions of human chromosome 11 demonstrate loss of heterozygosity or confer tumor suppression in chromosome transfer studies in specific types of human tumors (including hepatocellular carcinoma), suggesting the presence of multiple tumor suppressor loci on this chromosome.  Our laboratory developed a functional model for the identification of human liver tumor suppressor genes in which human chromosome 11 was introduced via microcell mediated chromosome transfer into the GN6TF rat liver epithelial tumor cell line, producing MCH cell lines that exhibit suppression of tumorigenicity.  Chromosome deletion mapping localized the tumor suppressor locus to a 950-kb region of 11p11.2-p12 between chromosomal markers D11S1361 and D11S1357, suggesting that this region harbors one or more genes with liver tumor suppressor function.  Examination of 181 EST and gene markers among the suppressed MCH cell lines localized twenty-three to the liver tumor suppressor region.  RT-PCR analysis (using the twenty-three markers) of gene expression identified five candidate genes, including three known genes (PIG11, PRDM11, and MTCH2) and two ESTs (stSG29748 and stSG10014) corresponding to cDNAs from novel human genes.  The eventual conclusion that one or more of these genes play a causal role in the molecular pathogenesis of human HCC required investigation of human liver tumors and derived cell lines.  We explored (i) gene expression and methylation-dependent gene silencing in the human HCC cell lines and (ii) LOH in primary human HCC tumors.  The five candidate transcripts were used to screen total RNA from a series of human HCC cell lines (n=19); 3 of 5 candidate genes demonstrated loss of expression in subsets of these cell lines.  These results indicate the expression of candidate transcripts, identified in a rat-human MCH model system, is deficient in human HCC cell lines, consistent with the activity of a liver tumor suppressor gene.  The evaluation of the five candidate transcripts in human HCC primary tumors and cell lines reinforces the conclusion that they play a role in the molecular pathogenesis of HCC.  Further characterization of genes in this study will contribute greater understanding of the molecular lesions and events that form the mechanistic basis for neoplastic transformation of human liver.