Asymmetric syntheses identify preferred stereochemistry in small molecule allosteric modulators of the neuropeptide Y4 receptor
Weaver, C. David
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PURPOSE: The neuropeptide Y4 receptor (Y4) is a GPCR belonging to a family of five receptors that bind ligands neuropeptide Y (NPY), peptide YY (PYY), and pancreatic polypeptide (PP). These ligands are hormones that play important roles in the regulation of feeding behavior and energy homeostasis. Small molecule ligands that selectively activate the Y4 receptor are potential therapeutics for obesity. Currently, there is a lack of non-peptide Y4-selective ligands available for studying Y4. Having recently identified multiple small molecule Y4 ligands via HTS, optimization of these hits is ongoing. The objective of the work described here is to develop asymmetric syntheses of two confirmed small molecule Y4 ligands, enabling determination of the preferred stereochemistry for Y4 activity and facilitating further optimization efforts. METHODS: Compounds were prepared via solution phase chemistry. Microwave reactions were run in an Anton Paar Monowave 200. Flash chromatography was carried out on either a CombiFlash® EZ Prep or CombiFlash® Rf+ system and utilized RediSep® Rf normal phase disposable columns. Preparative HPLC was carried out on a CombiFlash® EZ Prep system using a RediSep® Prep C18 10 x 250 mm, 100Å, 5 μm HPLC column from Teledyne ISCO. Compounds were characterized via NMR on a Bruker Fourier 300HD NMR spectrometer and via LCMS on an Agilent 6230 Accurate-Mass TOF LC/MS. Compound pharmacology was assessed in a two-addition protocol via Ca2+ flux assays in COS7 cells stably expressing Y4 and a chimeric G-protein. RESULTS: Four diastereomers of Y4 PAM tBPC were synthesized to [greater than] 87% d.e. utilizing a synthetic route employing an asymmetric alkylation, chelation controlled addition, and a Grubbs RCM reaction for the formation of key bonds. (1S,2S)-tBPC was found to be the preferred diastereomer for Y4 activity. Two enantiomers of Y4 NAM VU0637120 were synthesized to [greater than] 99% e.e. by employing commercially available chiral starting materials. (S)-VU0637120 was highly preferred for Y4 activity compared to (R)-VU0637120. A library based on (S)-VU0637120 was subsequently synthesized. CONCLUSIONS: Identification of the preferred stereoisomers for these Y4 ligands were successfully accomplished through asymmetric syntheses. These results enable the further optimization of these compounds in the context of the preferred stereochemistry, enhancing the probability of identifying optimized tools for studying the Y4 receptor.