New and emerging drug treatments for severe asthma.

Asthma is a common chronic inflammatory condition of the airways affecting over 300 million people world-wide. In 5%-10% of cases, it is severe, with disproportionate healthcare resource utilization including costs associated with frequent exacerbations and the long-term health effects of systemic steroids. Characterization of inflammatory pathways in severe asthma has led to the development of targeted biological and small molecule therapies which aim to achieve disease control while minimizing corticosteroid-associated morbidity. Herein, we review currently licensed agents and those in development, and speculate how drug therapy for severe asthma might evolve and impact on clinical outcomes in the near future.

A versatile synthesis of substituted benzimidazolium salts by an amination/ring closure sequence.

[reaction: see text]. A new method to produce benzimidazolium salts based on a successive Buchwald-Hartwig amination and ring closure is reported. A variety of different benzimidazolium salts can be prepared using this procedure. Amines that bear an alpha-chiral group undergo the reaction to furnish chiral benzimidazolium salts. The salts that lack a C2 substituent on the heterocycle are readily deprotonated to give nucleophilic carbenes.

Expanded scope in ethylene-alkyne cross-metathesis: coordinating heteroatom functionality at the propargylic position.

The Grubbs 1,3-dimesityl-4,5-dihydroimidazol-2-ylidene-substituted ruthenium complex 1 catalyzed ethylene-alkyne cross-metathesis and was shown to tolerate free hydroxyl groups and coordinating functionality at the propargylic and homopropargylic positions. Hindered and enantiomerically enriched 1-substituted alkynes also react efficiently under the reported conditions.

Proximity and orientation underlie signaling by the non-receptor tyrosine kinase ZAP70.

Signaling by the antigen receptor of T lymphocytes initiates different developmental transitions, each of which require the tyrosine kinase ZAP70. Previous studies with agonist and antagonist peptides have indicated that ZAP70 might respond differently to different structures of the TCR-CD3 complex induced by bound peptides. The roles of membrane proximity and orientation in activation of ZAP70 signaling were explored using synthetic ligands and their binding proteins designed to produce different architectures of membrane-bound complexes composed of ZAP70 fusion proteins. Transient membrane recruitment of physiological levels of ZAP70 with the membrane-permeable synthetic ligand FK1012A leads to rapid phosphorylation of ZAP70 and activation of the ras/MAPK and Ca2+/calcineurin signaling pathways. ZAP70 SH2 domains are not required for signaling when the kinase is artifically recruited to the membrane, indicating that the SH2 domains function solely in recruitment and not in kinase activation. Using additional synthetic ligands and their binding proteins that recruit ZAP70 equally well but orient it at the cell membrane in different ways, we define a requirement for a specific presentation of ZAP70 to its downstream targets. These results provide a mechanism by which ZAP70, bound to the phosphorylated receptor, could discriminate between conformational changes induced by the binding of different MHC-peptide complexes to the antigen receptor and introduce an approach to exploring the role of spatial orientation of signaling complexes in living cells.

Inducible gene expression and protein translocation using nontoxic ligands identified by a mammalian three-hybrid screen.

The natural product rapamycin has been used to provide temporal and quantitative control of gene expression in animals through its ability to interact with two proteins simultaneously. A shortcoming of this approach is that rapamycin is an inhibitor of cell proliferation, the result of binding to FKBP12-rapamycin-associated protein (FRAP). To overcome this limitation, nontoxic derivatives of rapamycin bearing bulky substituents at its C16-position were synthesized, each in a single step. The isosteric isopropoxy and methallyl substituents with the nonnatural C16-configuration abolish both binding to FRAP and inhibition of T cell proliferation. Binding proteins for these derivatives were identified from libraries of cDNAs encoding mutants of the FKBP12-rapamycin-binding (FRB) domain of FRAP by using a mammalian three-hybrid transcription assay. Targeting of the mutations was guided by the structure of the FKBP12-rapamycin-FRB ternary complex. Three compensatory mutations in the FRB domain, all along one face of an alpha-helix in a rapamycin-binding pocket, were identified that together restore binding of the rapamycin derivatives. Using this mutant FRB domain, one of the nontoxic rapamycin derivatives induced targeted gene expression in Jurkat T cells with an EC50 below 10 nM. Another derivative was used to recruit a cytosolic protein to the plasma membrane, mimicking a process involved in many signaling pathways.