Discovery of novel 5-oxa-2,6-diazaspiro[3.4]oct-6-ene derivatives as potent, selective, and orally available somatostatin receptor subtype 5 (SSTR5) antagonists for treatment of type 2 diabetes mellitus.

Somatostatin receptor subtype 5 (SSTR5) has emerged as a novel attractive drug target for type 2 diabetes mellitus. Starting from N-benzyl azetidine derivatives 1 and 2 as in-house hit compounds, we explored the introduction of a carboxyl group into the terminal benzene of 1 to enhance SSTR5 antagonistic activity by the combination of the substituents at the 3-position of the isoxazoline. Incorporation of a carboxyl group at the 4-position of the benzene ring resulted in a significant enhancement in potency, however, the 4-benzoic acid derivative 10c exhibited moderate human ether-a-go-go related gene (hERG) inhibitory activity. A subsequent optimization study revealed that replacement of the 4-benzoic acid with an isonipecotic acid dramatically reduced hERG inhibition (5.6% inhibition at 30µM) by eliminating π-related interaction with hERG K+ channel, which resulted in the identification of 1-(2-((2,6-diethoxy-4'-fluorobiphenyl-4-yl)methyl)-5-oxa-2,6-diazaspiro[3.4]oct-6-en-7-yl)piperidin-4-carboxylic acid 25a (hSSTR5/mSSTR5 IC50=9.6/57nM). Oral administration of 25a in high-fat diet fed C57BL/6J mice augmented insulin secretion in a glucose-dependent manner and lowered blood glucose concentration.

Betagamma subunits of G(i/o) suppress EGF-induced ERK5 phosphorylation, whereas ERK1/2 phosphorylation is enhanced.

Extracellular signal-regulated kinases (ERKs) play important physiological roles in proliferation, differentiation and gene expression. ERK5 is twice the size of ERK1/2, the amino-terminal half contains the kinase domain that shares the homology with ERK1/2 and TEY activation motif, whereas the carboxy-terminal half is unique. In this study, we examined the cross-talk mechanism between G-protein-coupled receptors (GPCRs) and receptor tyrosine kinases, focusing on ERK1/2 and 5. The pretreatment of rat pheochromocytoma cells (PC12) with pertussis toxin (PTX) specifically enhanced epidermal growth factor (EGF)-induced ERK5 phosphorylation. In addition, lysophosphatidic acid (LPA) attenuated the EGF-induced ERK5 phosphorylation in LPA(1) receptor- and G(i/o)-dependent manners. On the other hand, LPA alone activated ERK1/2 via Gbetagamma subunits and Ras and potentiated EGF-induced ERK1/2 phosphorylation at late time points. These results suggest G(i/o) negatively regulates ERK5, while it positively regulates ERK1/2. LPA did not affect cAMP levels after EGF treatment, and the reagents promoting cAMP production such as forskolin and cholera toxin also attenuated the EGF-induced ERK5 phosphorylation, indicating that the inhibitory effect of LPA on ERK5 inhibition via G(i/o) is not due to inhibition of adenylyl cyclase by Galpha(i/o). However, the inhibitory effect of LPA on ERK5 was abolished in PC12 cells stably overexpressing C-terminus of GPCR kinase2 (GRK2), and overexpression of Gbeta(1) and gamma(2) subunits also suppressed ERK5 phosphorylation by EGF. In response to LPA, Gbetagamma subunits interacted with EGF receptor in a time-dependent manner. These results strongly suggest that LPA negatively regulates the EGF-induced ERK5 phosphorylation through Gbetagamma subunits.

Isolation and synthesis of a new aromatic compound, brefelamide, from dictyostelium cellular slime molds and its inhibitory effect on the proliferation of astrocytoma cells.

[structure: see text] We have explored the diversity of secondary metabolites produced by cellular slime molds to examine the possible use of such cellular slime molds as a resource for novel drug development. A new aromatic amide, brefelamide (1), was isolated from methanol extracts of the fruiting bodies of Dictyostelium brefeldianum and D. giganteum. The structure of 1 was determined by spectral means including EIMS and (1)H and (13)C NMR. The total synthesis of 1 was carried out to confirm the structure and obtain sufficient samples for performing biological evaluation. Interestingly, compound 1 inhibited the cellular proliferation of 1321N1 human astrocytoma cells.