Type 1 neurofibromatosis (NF1) is a prevalent familial cancer syndrome affecting certain numbers of human beings. The abnormal genetic locus in neurofibromatosis is Nf1, a tumor suppressor gene. The protein product of Nf1 (neurofibromin) is a GTPase-activating protein (GAP) that negatively regulates Ras signaling and expresses in a high level in Schwann cells that are the cell origin of NF1 disease. Loss-of- function mutations or deletion of Nf1 are responsible for hyper-activation of Ras pathway in NF1 patients. Our preliminary data demonstrated that mutated Nf1, together with the suppression of protein kinase C (PKC), is synthetically lethal, via the activation of the mitotic exit checkpoint. In this process, Plk1 was activated in a Chk1-dependent fashion. Through binding to cdc14B, Plk1 blocked its functions and further silenced Cdh1 activity, resulting in the increase of cyclin B1 stability and accumulation of Nf1 deficient neurofibromatosis cells in the M phase of the cell cycle. Importantly, the activation of this mitotic exit checkpoint coincided with the induction of apoptosis. In yeast and mammalian cells, mitotic exit involves an intricately ordered series of events, leading from the splitting of sister chromatids at anaphase onset to the completion of cell division by cytokinesis. If insults target late stages of mitosis, mitotic exit checkpoint would be activated and subsequently induce mitotic catastrophe to eliminate unwanted cells or terminates mitosis. Based on our preliminary data and our knowledge of the mitotic exit checkpoint, we hypothesize that Nf1 mutation, together with loss of PKC, is synthetically lethal, which is through the activation of the mitotic exit checkpoint. Identification of critical molecular events in this mitotic crisis will allow efficient therapeutic interventions of neurofibromatosis 1. To test this hypothesis, we will: 1) investigate the mechanism by which Plk1/cdc14B axis regulates the activation of this mitotic exit checkpoint in HMG- treated Nf1 deficient neurofibromatosis 1 cells;2) investigate the role of Cdh1 in the mitotic exit checkpoint induced by PKC suppression in the Nf1 deficient cells;and 3) determine how Ras-mediated pathways trigger the mitotic exit checkpoint in HMG-treated Nf1 deficient cells and further induce apoptosis in vivo. Since Nf1 defect affects a large of human population and the poor prognosis for patients with advanced malignant peripheral nerve sheath tumor (MPNST) emphasize the urgent need for new chemotherapeutic approaches to treat this disease, the outcomes of this proposed study will have translational importance in the design of new therapeutic strategies to manipulate intracellular signaling and specifically re-direct the Nf1 deficient cells to undergo apoptosis, which is fully in accord with the central mission of NIH.
Type 1 neurofibromatosis (NF1) is a prevalent familial cancer syndrome affecting many individuals worldwide. Here, we propose the use of molecular/cellular biological techniques and genetic/chemical inhibitors of cell signaling pathways to explore the importance of the induction synthetic lethality in Nf1 deficient cells. The long-term goal of these experiments is the development of new and rationally designed therapeutic strategies to more effectively treat the diseases caused by Nf1 defect.
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