The gut endocrine system is emerging as a central player in the control of appetite and glucose homeostasis, and as a rich source of peptides with therapeutic potential in the field of diabetes and obesity. In this study we have explored the physiology of insulin-like peptide 5 (Insl5), which we identified as a product of colonic enteroendocrine L-cells, better known for their secretion of glucagon-like peptide-1 and peptideYY. i.p. Insl5 increased food intake in wild-type mice but not mice lacking the cognate receptor Rxfp4. Plasma Insl5 levels were elevated by fasting or prolonged calorie restriction, and declined with feeding. We conclude that Insl5 is an orexigenic hormone released from colonic L-cells, which promotes appetite during conditions of energy deprivation.

The antibacterial activity of TAK-083 was tested against 54 clinical isolates of Helicobacter pylori and was compared with those of amoxicillin, clarithromycin, and metronidazole. The growth-inhibitory activity of TAK-083 was more potent than that of amoxicillin, clarithromycin, or metronidazole (the MICs at which 90% of the strains are inhibited were 0.031, 0.125, 64, and 8 g/ml, respectively). The antibacterial activity of TAK-083 was highly selective against H. pylori; there was a >30-fold difference between the concentration of TAK-083 required to inhibit the growth of H. pylori and that required to inhibit the growth of common aerobic and anaerobic bacteria. Exposure of H. pylori strains to TAK-083 at the MIC or at a greater concentration resulted in an extensive loss of viability. When four H. pylori strains were successively subcultured in the medium containing subinhibitory concentrations of TAK-083, no significant change in the MICs of this compound was observed. TAK-083 strongly inhibited the formation of tryptophanyl-tRNA in H. pylori while exhibiting little effect on the same system in eukaryotes. TAK-083 was efficacious in the treatment of gastric infection caused by H. pylori in Mongolian gerbils. The results presented here indicate that TAK-083 is a promising candidate for the treatment of H. pylori infection.

A novel class of therapeutic drug candidates for heart failure, highly potent and selective GRK2 inhibitors, exhibit potentiation of β-adrenergic signaling in vitro studies. Hydrazone derivative 5 and 1,2,4-triazole derivative 24a were identified as hit compounds by HTS. New scaffold generation and SAR studies of all parts resulted in a 4-methyl-1,2,4-triazole derivative with an N-benzylcarboxamide moiety with highly potent activity toward GRK2 and selectivity over other kinases. In terms of subtype selectivity, these compounds showed enough selectivity against GRK1, 5, 6, and 7 with almost equipotent inhibition to GRK3. Our medicinal chemistry efforts led to the discovery of 115h (GRK2 IC = 18 nM), which was obtained the cocrystal structure with human GRK2 and an inhibitor of GRK2 that potentiates β-adrenergic receptor (βAR)-mediated cAMP accumulation and prevents internalization of βARs in β2AR-expressing HEK293 cells treated with isoproterenol. Therefore, 115h appears to be a novel class of therapeutic for heart failure treatment.

Poly(ADP-ribose) polymerases (PARPs) use nicotinamide adenine dinucleotide (NAD) as a co-substrate to transfer ADP-ribose when it releases nicotinamide as the metabolized product. Enzymes of the PARP family play key roles in detecting and repairing DNA, modifying chromatin, regulating transcription, controlling energy metabolism, and inducing cell death. PARP14, the original member of the PARP family, has been reported to be associated with the development of inflammatory diseases and various cancer types, making it a potential therapeutic target. In this study, we purified the macrodomain-containing PARP14 enzyme and established an assay for detecting the auto-ribosylation activity of PARP14 using RapidFire high-throughput mass spectrometry and immunoradiometric assay using [H]NAD. Subsequently, we performed high-throughput screening using the assays and identified small-molecule hit compounds, which showed NAD-competitive and PARP14-selective inhibitory activities. Co-crystal structures of PARP14 with certain hit compounds revealed that the inhibitors bind to the NAD-binding site. Finally, we confirmed that the hit compounds interacted with intracellular PARP14 by a cell-based protein stabilization assay. Thus, we successfully identified primary candidate compounds for further investigation.

An accurate immunoassay system was developed for the quantitative analysis of linear alkylbenzene sulfonates (LAS), the most widely used anionic surfactants among domestic detergents. To generate LAS-specific monoclonal antibodies (mAb), hybridoma cells were produced by the fusion of mouse myeloma cells and spleen cells from mice immunized with sulfophenyl-5-valeric acid coupled to bovine serum albumin. After screening using a competitive enzyme-linked immunosorbent assay (ELISA), a mAb with a high binding affinity for LAS was selected and used in the development of a sensitive competitive direct ELISA. The detection range is between 20 and 500 ppb LAS. The ELISA assay was optimized and validated by comparison with conventional methods for the analysis of LAS and anionic surfactants, such as high-performance liquid chromatog raphy (HPLC) and methylene blue active substances (MBAS) methods, in river samples. The correlation coef ficients between the assay values obtained using the ELISA and the HPLC and MBAS were 0.98 and 0.97, respectively. It is anticipated that this immunoassay system will be a useful monitoring technique for the detection of LAS in environmental water samples.