Part I. Design of a novel class of reversible non-covalent small molecule inhibitors for human Granzyme B (hBrB) Part II. Curcumin and mimics as proteasome inhibitors Part III. Design of novel coactivator binding inhibitors (CBIs) for the estrogen recept

Part I. Design of a novel class of reversible non-covalent small molecule inhibitors for human Granzyme B (hGrB): Grazyme B, lymphocyte serine protease, plays a critical role in controlling graft versus leukemia and graft-versus-host diseases. A key aim of this study is to design non-covalent small molecule inhibitors using a computational model and screening approach: 1) computational solvent mapping to identify "hot spots" in the active site; 2) virtual screening with three constraints based on the solvent mapping results; 3) measure enzyme activity and selectivity; 4) validate by modeling known covalent blockers. As a result, novel classes of hGrB inhibitors have been identified. To extend the pool of scaffolds, 'scaffold hopping' has been carried out to perform shape-based searching.;Part II. Curcumin and mimics as proteasome inhibitors: The proteasome is an important target of curcumin, and several groups have reported that inhibition of proteasome activity by curcumin and chalcone-based derivatives cause apoptosis in human cancer cell lines. In those studies, DFT calculations and in silico docking studies were performed to suggest binding poses in the CT-like subunit. However, these studies introduce critical and most-likely incorrect assumptions: 1) the LUMOs were calculated for the unstable diketo form; 2) the LUMO characteristics were interpreted to suggest that the carbonyl carbons should be the site of the terminal Thr1 nucleophilic attack instead of the β-carbon of the enone; 3) an incorrect cis-structure for the chalcone analogs was employed for the docking. Thus, the specific aim of this study is to identify the correct binding poses of curcumin and chalcone-based derivatives and to explore the binding pockets of the three active β subunits to explain why curcumin potently inhibits the CT-like activity.;Part III. Design of novel coactivator binding inhibitors (CBIs) for the estrogen receptor α: break the 1μM barrier: Novel classes of coactivator binding inhibitors against estrogen receptor α have been developed. However, they fail to deliver IC50 values below 1μM in the reporter gene assay. To understand these observations and break the 1μM barrier, we performed MD simulations to examine solvent-based entropic contributions to the free energy of ligand binding. The coactivator is far more effective at expelling water molecules from the binding site than the CBIs. These observations strongly suggest that the next generation of small molecule CBIs should span more of the peptide space, particularly on the shelf adjacent to the binding groove.