Our broad goals include delineating how the cell division cycle is regulated, determining how cancer cellsderail cell cycle regulatory pathways, and ultimately using this information for diagnosing and treatinghuman cancers. We propose to focus our efforts on the Cdc25A protein phosphatase as this enzyme is akey regulator of the cell division cycle in mammals and is overproduced in a wide range of human tumors.Stable cell lines and mouse colonies will be generated that enable Cdc25A regulation to be studied in cellsand living mice using molecular imaging technologies. In particular, cell lines that inducibly express a fusionprotein between human Cdc25A and firefly luciferase (Cdc25A-FLuc) will be used in a high throughputscreen to identify the serine/threonine protein phosphatase holoenzymes that regulate the stability ofCdc25A in vivo. In principal the proteins identified in this study could provide novel targets for imagingagents and therapeutic intervention in cancer treatment. In addition, knock-in mice will be generated thatexpress a fusion protein between endogenous Cdc25A and click beetle red luciferase (Cdc25A-CBRLuc)from the Cdc25A locus. In this way endogenous Cdc25A protein levels can be monitored non-invasivelyand repetitively in living mice under steady state conditions and in response to DMAdamaging agents anddrugs targeting cell cycle checkpoints. In addition, molecular imaging strategies will be applied to mousemodels of breast cancer to validate target specificity of rational anti-cancer therapeutic regimens incombination therapies. The reagents and results obtained in these studies will be useful in future clinicaltrials that combine DNA damaging agents with novel Chk1 inhibitors or novel checkpoint abrogators. Theproposed studies are expected to enhance our understanding of basic principles of cell cycle control inmammals and may impact future therapeutic strategies for cancer treatment.
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