Wenxin Gu

Celastrols as inducers of the heat shock response and cytoprotection

Alterations in protein folding and the regulation of conformational states have become increasingly important to the functionality of key molecules in signaling, cell growth, and cell death. Molecular chaperones, because of their properties in protein quality control, afford conformational flexibility to proteins and serve to integrate stress-signaling events that influence aging and a range of diseases including cancer, cystic fibrosis, amyloidoses, and neurodegenerative diseases. We describe here characteristics of celastrol, a quinone methide triterpene and an active component from Chinese herbal medicine identified in a screen of bioactive small molecules that activates the human heat shock response. From a structure/function examination, the celastrol structure is remarkably specific and activates heat shock transcription factor 1 (HSF1) with kinetics similar to those of heat stress, as determined by the induction of HSF1 DNA binding, hyperphosphorylation of HSF1, and expression of chaperone genes. Celastrol can activate heat shock gene transcription synergistically with other stresses and exhibits cytoprotection against subsequent exposures to other forms of lethal cell stress. These results suggest that celastrols exhibit promise as a new class of pharmacologically active regulators of the heat shock response.

Stereospecific total syntheses of proteasome inhibitors omuralide and lactacystin.

Omuralide, a transformation product of the microbial metabolite lactacystin, was the first molecule discovered as a specific inhibitor of the proteasome and is unique in that it specifically inhibits the proteolytic activity of the 20S subunit of the proteasome without inhibiting any other protease activities of the cell. The total syntheses of omuralide and (+)-lactacystin are reported. An important key intermediate is synthesized at an early stage, which allows analogues of these two natural products to be made readily.

Inactivation of mitochondrial monoamine oxidase B by methylthio-substituted benzylamines

Mitochondrial monoamine oxidase was inactivated by o-mercaptobenzylamine (1) and o- (2) and p-methylthiobenzylamine (5). Experiments were carried out to provide evidence for possible mechanisms of inactivation. The corresponding o- (3) and p-hydroxybenzylamine (4) are not inactivators. Four radiolabeled analogues of 2 and 5, having radioactivity at either the methyl or benzyl groups, were synthesized, and all were shown to incorporate multiple equivalents of radioactivity into the enzyme. Inactivation in the presence of an electrophile scavenger decreased the number of molecules incorporated, but still multiple molecules became incorporated; catalase did not further reduce the number of inactivator molecules bound. Two inactivation mechanisms are proposed, one involving a nucleophilic aromatic substitution (SNAr) mechanism and the other a dealkylation mechanism. Evidence for both mechanisms is that inactivation leads to reduction of the flavin (oxidation of the inactivator), but upon denaturation the flavin is reoxidized, indicating that attachment is not at the flavin. A cysteine titration indicates the loss of four cysteines after inactivation and denaturation. Support for the SNAr mechanism was obtained by showing that o- and p-chlorobenzylamine also inactivate MAO. Chemical model studies were carried out that also support both SNAr and dealkylation mechanisms.

Concise route to the chiral pyrrolidine core of selective inhibitors of neuronal nitric oxide.

2-(((3R,4R)-4-(Allyloxy)-1-benzylpyrrolidin-3-yl)methyl)-6-(2,5-dimethyl-1H-pyrrol-1-yl)-4-methylpyridine (2), a key intermediate for the preparation of novel neuronal nitric oxide synthase (nNOS) inhibitors, is synthesized using diisopropyl (R)-(+)-malate as the starting material. The key steps involve a Frater-Seebach diastereoselective alkylation and a fast intramolecular cyclization.