The purpose of this exploratory grant proposal is to test the hypothesis that one/several metabolic enzymes with deficiency in a gene of interest to cause cell death in ovarian human tumor cells. The gene of interest is a tumor suppressor gene deficient in the particular ovarian carcinoma. Recently we established both chemical- and genetic-synthetic lethality screens in cultu5red human cells. The technology employed an immortalized human cell line, deficient in a gene of interest, which is complemented by an episomal survival plasmid expressing the wild-type cDNA for the gene of interest, and the use of a novel GFP-based double- label fluorescence system. Selective pressure by either chemicals, or dominant-negative genetic suppressor elements (GSEs), selected from a library expressing short truncate sense and antisense cDNAs for a candidate synthetic lethal gene, prevented the spontaneous loss of the episomal survival plasmid. Retention or loss over time of the survival plasmid could be sensitively detected by measuring the fluorescence levels expressed off the GFP-tagged survival plasmid. This also allowed the isolation of cells retaining the survival plasmid, and the rescue of the putative GSEs. We now propose to apply the genetic synthetic lethality screen t ovarian carcinoma in order to examine whether any one of several candidate metabolic genes is synergistically lethal with a deficiency in the tumor suppressor gene of interest. I successful, it would allow the search of new drugs for ovarian malignancy, aimed at noel target/s whose specificity is insured by its/their synthetic lethality with a mutated tumor suppressor gene of interest.
