Enantiomeric Separation, Absolute Configuration by X-ray Crystallographic Analysis, and Functional Evaluation of Enantiomers of the Dual Ligand, SYA0340 at 5-HT and 5-HT Receptors.

We have previously identified 5-chloro-2-methyl-2-(3-(4-(pyridin-2-yl)piperazin-1-yl)propyl)-2,3-dihydro-1-inden-1-one (SYA0340) as a dual 5-HT and 5-HT receptor ligand, and we posited such ligands might find utility in the treatment of various CNS related illnesses including cognitive and anxiolytic impairments. However, SYA0340 has a chiral center and its enantiomers may confound the readouts for their functional characteristics. Thus, in this study, we resynthesized SYA0340, separated the enantiomers, identified the absolute configurations, and evaluated their binding affinities and functional characteristics at both the 5-HT and 5-HT receptors. The results of this study show that the (+)-SYA0340-P1 [specific rotation [α] = +18.4 (deg.mL)/(g.dm)] has a binding affinity constant, = 1.73 ± 0.55 nM at 5-HTR and = 2.20 ± 0.33 nM at 5-HTR and (-)-SYA0340-P2 [specific rotation [α] = -18.2 (deg.mL)/(g.dm)] has = 1.06 ± 0.32 nM (5-HTR) and 4.7 ± 1.1 nM (5-HTR). Using X-ray crystallographic techniques, the absolute configuration of the P2 isomer was identified as the S-enantiomer and, therefore, the P1 isomer as the R-enantiomer. Functionally, both SYA0340-P1 (EC = 1.12 ± 0.41 nM; = 94.6 ± 3.1%) and SYA0340-P2 (EC = 2.21 ± 0.59 nM; = 96.8 ± 5.1%) display similar agonist properties at the 5-HTR while both enantiomers display antagonist properties at the 5-HTR with P1 (IC = 32.1 ± 9.2 nM) displaying over 8 times greater potency as P2 (IC = 277 ± 46 nM). Thus, based on the functional evaluation results, SYA0340-P1 is considered as the eutomer of the pair of enantiomers of SYA0340. It is expected that these enantiomers will serve as new pharmacological probes for the 5-HT and 5-HT receptors.