The goal of this proposal is to identify cellular genes that are critical for growth or survival of neoplastically transformed cells and to design prototype """"""""genetic drugs"""""""" targeting such genes. The target genes will be identified through the selection of cDNA fragments that inhibit the function of the gene from which they are derived. These fragments are called growth-inhibiting genetic suppressor elements (GSEs). GSEs that have a cytostatic or cytotoxic effect on neoplastically transformed cells will be isolated using inducible retroviral vectors and selection protocols already developed during the previous period of this project. Three types of selections will be carried out. The first selection will use a random fragment library prepared from a mixture of cDNA clones of different human genes implicated in positive regulation of cell growth. This will be used to isolate GSEs that would be inhibitory for a human fibrosarcoma cell line. The second selection will be carried out in a transformed mouse embryo fibroblast (MEF) cell line, using a library of random cDNA fragments from genes that are overexpressed in the transformed line (relative to untransformed MEF). The third selection will utilize a library of random fragments of normalized cDNA corresponding to all genes expressed in the transformed MEF line; this library will be used to isolate GSEs that are specially cytostatic or cytotoxic to neoplastically transformed MEF cells (in relation to the untransformed MEF cells). Full-length human sequences of unknown genes giving rise to the selected GSGs will be cloned and their expression patterns will be characterized. The biological activity of the GSEs isolated through all selection types will be tested on cell lines derived from different types of mouse or human tumors and on normal cells. The most efficient GSEs will be tested in the form of retroviral vectors or chemically synthesized peptides or oligonucleotides. The proposed studies should lead to the identification of new targets and prototype genetic drugs with potential for the treatment of cancer.
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