Neuroprotective properties of c-Rafi inhibitors

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DESCRIPTION (provided by applicant): Neurological diseases disrupt the quality of the lives of patients, puts a tremendous burden on family caregivers, and cost society billions of dollars annually. A common feature of neurological diseases is the degeneration of neurons by apoptosis. Drugs that inhibit neuronal apoptosis could thus be candidates for therapeutic intervention in neurodegenerative disorders. Moreover, identifying the molecular targets of such neuroprotective drugs and understanding the signal transduction pathways that are utilized in their action would lead to the development of more effective therapeutic strategies. Working with a cell culture paradigm of neuronal apoptosis that uses rat cerebellar granule neurons we have identified a drug, GW5074 {5-Iodo-3-[(3,5-dibromo-4-hydroxyphenyl)methylene]-2-indolinone} that completely inhibits neuronal apoptosis. GW5074 is a specific and potent inhibitor of c-Raf when tested in vitro. Paradoxically, however, treatment of cultured neurons with GW5074 leads to the accumulation of activating modifications on c-Raf. Moreover, GW5074 treatment stimulates B-Raf activity. Among the molecules affected downstream of c-Raf in neurons treated with GW5074 are the antiapoptotic molecule NF-kappa b. GW5074 also inhibits the proapoptotic transcription factor, c-jun. Although GW5074 is the most effective, neuroprotection is also observed by two other chemical inhibitors of c-Raf. The utility of small molecule inhibitors of c-Raf as neuroprotective agents has not been described previously. The overall goal of this proposal is to use GW5074 to understand the molecular mechanism by which inhibitors of c-Raf exert their antiapoptotic effect and to more thoroughly investigate the potential of GW5074 as a neurotherapeutic agent. Our specific goals are to: (1) better understand the effect of GW5074 on c-Raf and B-Raf understand the effect of GW5074 on c-Raf, (2) analyze the downstream mechanisms by which GW5074 exerts its neuroprotective effect, and (3) examine whether the molecular effects of GW5074 in cultured cerebellar granule neurons are also observed in an animal model of neurodegeneration. It is our hope that GW5074 (or other c-Raf inhibitors like it) will emerge as a highly effective and versatile therapeutic agent that could proceed towards clinical trials for the treatment of neurological diseases in the near future.