As realized by this RFA, there is a need to identify novel molecular targets for cancer drug discovery. Among these will be molecules that directly regulate the intrinsic apoptotic pathway. We recently discovered that the large GTPase dynamin-2 (dyn2) signals through p53 to trigger apoptosis, specifically in rapidly dividing cells. As little as five-fold overexpression of endogenous wild-type dyn2 will activate p53-dependent apoptosis and this is absolutely dependent on GTP binding. A mutation defective in dynamin's intrinsic GAP activity is even more potent. Thus, compounds that inhibit dynamin's GTPase activity or its intrinsic GAP may be effective proapoptotic, anti-cancer drugs. To identify these compounds we have developed a simple and robust high-throughput assay for dynamin GTPase activity and established a collaboration with Barry Sharpless, a leading chemist who has pioneered a new azido-based method of """"""""click chemistry"""""""" that facilitates the generation of complex libraries of related compounds, allows for rapid modular SAR of identified leads and most importantly and uniquely allows the assembly of functional moieties to be selected for on the surface of the target molecule to generate customized, high-affinity, high-specificity inhibitors. We propose the following Specific Aims that will allow us to apply click chemistry to identify potential anti-cancer drugs targeted against the regulatory GTPase, dynamin. 1. To identify, characterize and optimize high affinity, high specificity compounds that inhibit dynamin's basal GTPase activity. 2. To identify, characterize and optimize high affinity, high specificity compounds that selectively inhibit dynamin's assembly-stimulated GTPase activity. 3. To apply click chemistry for in situ selection of very high affinity, very high specificity inhibitors on dynamin, itself.
| Ramachandran, Rajesh; Surka, Mark; Chappie, Joshua S et al. (2007) The dynamin middle domain is critical for tetramerization and higher-order self-assembly. EMBO J 26:559-66 |
| Soulet, Fabienne; Schmid, Sandra L; Damke, Hanna (2006) Domain requirements for an endocytosis-independent, isoform-specific function of dynamin-2. Exp Cell Res 312:3539-45 |
