Numerous studies suggest that inactivation or deletion of one or more tumor suppressor genes may represent early and/or necessary steps in the neoplastic transformation of liver. Furthermore, comparative chromosomal mapping studies suggest that the development of liver tumors in humans and rodents may share a common molecular mechanism that involves inactivation of analogous tumor suppressor genes. Based upon these observations, we have suggested that chromosome transfer studies utilizing human chromosomes and rat liver tumor cell lines may facilitate the facile localization, identification, and cloning of human genes responsible for tumor suppression in liver. To this end, we have established and characterized a model for the functional identification of human tumor suppressor genes in rat liver tumor cell lines. Human chromosome 11, which has been implicated in the pathogenesis of human liver tumors, was introduced into highly aggressive rat liver epithelial tumor cell lines using a microcell-mediated chromosome transfer technique. We have demonstrated that human chromosome 11 suppresses the tumorigenic potential of some rat liver tumor cell lines, providing direct evidence for the presence of a liver tumor suppressor gene on this human chromosome. Further, we have established a microsatellite- based map of the liver tumor suppressor region, localizing the gene(s) to a 950-kb segment of human 11p11.2-p12 and forming the basis for positional cloning of candidate tumor suppressor genes. The studies contained in this proposal seek to identify, isolate, and characterize the human 11p11.2-p12 liver tumor suppressor gene. The goals of this proposal are to (i) establish a PAC contig spanning the liver tumor suppressor region of human 11p11.2-p12, (ii) utilize the isolated PAC clones in the construction of a complete physical map of this chromosomal region, (iii) isolate candidate liver tumor suppressor genes based upon complementarity to PAC clones corresponding to this region, (iv) characterize candidate liver tumor suppressor genes by DNA sequence analysis, and (v) determine the ability of candidate genes to express tumor suppressor activity in transfected rate liver tumor cell lines.
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