3-Amido pyrrolopyrazine JAK kinase inhibitors: development of a JAK3 vs JAK1 selective inhibitor and evaluation in cellular and in vivo models.

The Janus kinases (JAKs) are involved in multiple signaling networks relevant to inflammatory diseases, and inhibition of one or more members of this class may modulate disease activity or progression. We optimized a new inhibitor scaffold, 3-amido-5-cyclopropylpyrrolopyrazines, to a potent example with reasonable kinome selectivity, including selectivity for JAK3 versus JAK1, and good biopharmaceutical properties. Evaluation of this analogue in cellular and in vivo models confirmed functional selectivity for modulation of a JAK3/JAK1-dependent IL-2 stimulated pathway over a JAK1/JAK2/Tyk2-dependent IL-6 stimulated pathway.

ErbB3 inhibitory surrobodies inhibit tumor cell proliferation in vitro and in vivo.

ErbB3 is an important regulator of tumorigenesis and is implicated in development of resistance to several currently used oncology drugs. We have identified ErbB3 inhibitors based on a novel biologic scaffold termed a surrobody. Two of these inhibitors appear to work by a previously unrecognized mechanism of action.

Biochemical characterization of the inhibition of the dengue virus RNA polymerase by beta-d-2'-ethynyl-7-deaza-adenosine triphosphate.

Dengue virus (DENV), an emerging pathogen from the Flaviviridae family with neither vaccine nor antiviral treatment available, causes a serious worldwide public health threat. In theory, there are several ways by which small molecules could inhibit the replication cycle of DENV. Here, we show that the nucleoside analogue beta-d-2'-ethynyl-7-deaza-adenosine inhibits representative strains of all four serotypes of DENV with an EC(50) around or below 1microM.

Structural insights for design of potent spleen tyrosine kinase inhibitors from crystallographic analysis of three inhibitor complexes.

Spleen tyrosine kinase is considered an attractive drug target for the treatment of allergic and antibody mediated autoimmune diseases. We have determined the co-crystal structures of spleen tyrosine kinase complexed with three known inhibitors: YM193306, a 7-azaindole derivative and R406. The cis-cyclohexyldiamino moiety of YM193306 is forming four hydrophobically shielded polar interactions with the spleen tyrosine kinase protein and is therefore crucial for the high potency of this inhibitor.

Molecular mechanism of the Syk activation switch.

Many immune signaling pathways require activation of the Syk tyrosine kinase to link ligation of surface receptors to changes in gene expression. Despite the central role of Syk in these pathways, the Syk activation process remains poorly understood. In this work we quantitatively characterized the molecular mechanism of Syk activation in vitro using a real time fluorescence kinase assay, mutagenesis, and other biochemical techniques.

The crystal structure of JNK2 reveals conformational flexibility in the MAP kinase insert and indicates its involvement in the regulation of catalytic activity.

c-Jun N-terminal kinase (JNK) 2 is a member of the mitogen-activated protein (MAP) kinase group of signaling proteins. MAP kinases share a common sequence insertion called "MAP kinase insert", which, for ERK2, has been shown to interact with regulatory proteins and, for p38alpha, has been proposed to be involved in the regulation of catalytic activity. We have determined the crystal structure of human JNK2 complexed with an indazole inhibitor by applying a high-throughput protein engineering and surface-site mutagenesis approach.

Steady state kinetics of spleen tyrosine kinase investigated by a real time fluorescence assay.

Spleen tyrosine kinase (Syk) is a cytoplasmic tyrosine kinase that plays an important signaling role in several types of immune cells. To improve our understanding of the enzymology and activation mechanism of Syk, we characterized the steady state kinetics of Syk substrate phosphorylation. A new real time fluorescence kinase assay was employed that utilizes a nonnatural amino acid in the peptide substrate which undergoes an enhancement in fluorescence following phosphorylation.