Among the compounds prepared at

Among the compounds prepared at this stage, the cyclopropylmethylenoxy analogue attracted our most attention since it demonstrated particularly good in vitro potency at GlyT1 (16nM), no activity at the GlyT2 isoform up to the highest concentration tested (30μM), good physical properties with moderate lipophilicity: log=1.83, good aqueous solubility: 31μg/mL (pH 6.5), high membrane permeability: Pe (Pampa): 3.2, and good mouse and human microsomal stability. Consequently, was selected for further evaluation. An in vivo pharmacokinetic study in mouse () revealed that , as expected based on its in vitro profile, exhibited low clearance, good plasma exposure, excellent oral bioavailability, and moderate plasma protein binding (PPB). Despite demonstrating a relatively low brain/plasma ratio, could achieve nevertheless a robust exposure in the brain, with a of 373ng/g at the dose of 10mg/kg (po), above its inhibitory activity at the GlyT1 transporter. These favorable data prompted us to evaluate the effect of KPT-330 on the extracellular level of glycine in mouse striatum (). We were delighted to observe that produced at the dose of 10mg/kg administrated orally, a robust and sustained increase of glycine levels: 2.3-fold over basal level at 40min and 1.7-fold at 140min. Encouraged by this result, further in vitro profiling in safety assays was performed. Compound (like all compounds tested in this series) exhibited no significant inhibition of the major drug metabolizing CYP450 enzymes (3A4, 2D6, 2C9, 2C19, 1A2; IC>29.1μM). However, was found to exhibit a fairly high activity at the hERG potassium channel with an IC of 0.6μM measured in a patch-clamp assay. In summary, we report here on the discovery of benzoylpiperazines as a novel chemotype of GlyT1 inhibitors. Starting from the chemically tractable HTS hit , rapid exploration of SAR accompanied by the early assessment of molecular property and metabolic stability of the synthesized derivatives allowed the prompt identification of a highly promising sub-series in which the nitro and morpholine residues in are replaced, respectively, by a methylsulfone and an alkoxy group. Within this ‘alkoxy-methylsulfone-benzoylpiperazine’ sub-series, highly potent and selective GlyT1 inhibitors were identified, which as seen with representative compound demonstrated drug-like properties and in vivo efficacy after oral administration. Results from our subsequent lead optimization work in this sub-series, which was mainly focused on the improvement of brain penetration/in vivo activity as well as of hERG selectivity, will be reported in due course. Acknowledgments
Based on the glutamatergic hypothesis of schizophrenia, glycine transporter type 1 (GlyT1) is a promising target for antipsychotic agents. Several GlyT1 inhibitors have been reported, and some of these agents are now being investigated in clinical trials. We recently isolated a cyclic tetrapeptide, named WSS2220 (), as a potent and selective inhibitor of GlyT1. Compound was produced by sp. TA-0426 that was isolated from a soil sample collected at Henan Province, China. In our successive exploration of GlyT1 inhibitors, we were able to isolate five new tetrapeptides, –, from a fermentation broth of TA-0426. In this paper, we report the isolation, structure determination and biological activity of these new peptides. The preliminary structure–activity relationship (SAR) based on the structures of the isolated compounds and their synthetic analogues is also described. The -BuOH extract (6.0g) of TA-0426 was suspended in distilled water and extracted with EtOAc. The EtOAc extract was then separated by silica gel column chromatography (SCC), eluting with a CHCl–MeOH solvent system. The cyclic peptide-containing material, eluted with CHCl–MeOH 9:1, was further purified by reversed-phase HPLC (column; YMC-Pack ODS-AM 250×10mm; id, 5μm; YMC, flow rate; 2.5mL/min), eluting with CHCN–HO 45:55, to give WSS2219 (, 13.4 mg; rt=15.6min), WSS2221 (, 3.7mg, rt=11.4min), WSS2222 (, 5.7mg, rt=12.0min) and WSS2217 (, 3.5mg, rt=9.7min) as colorless powders. The CHCl–MeOH 19:1 eluent from SCC was purified by reversed-phase HPLC, eluting with CHCN–HO 65:35, to give WSS2218 (, 15.2mg, rt=13.6min) as a colorless powder ().