Discovery, asymmetric synthesis and biological evaluation of new bisbenzylisoquinoline-orexin receptor antagonists for insomnia treatment.

The development of new orexin receptor (OXR) antagonists represents a crucial avenue for addressing the urgent need for safe and effective anti-insomnia drugs in clinical practice. Our group discovered neferine, a bisbenzylisoquinoline alkaloid isolated from Nelumbinis Plumula, exhibited a predicted binding affinity for OXR through virtual screening. Herein, we report the asymmetric synthesis of neferine and its isomers using a novel CuBr•Me2S/picolinic acid-catalyzed arylation method. Biological assay results indicated that (R,S)-neferine demonstrated superior OXR antagonistic activity than suvorexant, with good binding affinities of KD (OX1R) 0.68 and (OX2R) 0.81 nM. In vivo pharmacodynamic studies showed that (R,S)-neferine reduced spontaneous activities in mice more effectively than daridorexant and significantly improved sleep in insomnia mice. Furthermore, (R,S)-neferine can restore the significant downregulation of PER1 and PER2, along with the upregulation of BMAL1 caused by insomnia, and decrease the expression of OX1R and OX2R, thereby improving core circadian rhythm disorders. (R,S)-neferine carried outstanding pharmacokinetic (F = 63.7 %) and safety profiles (LD50 > 500 mg/kg), no mortality or histopathological changes were observed during the acute toxicity test. Overall, this work highlights the therapeutic potential of the OXR antagonist (R,S)-neferine, providing candidates and viable drug development strategies for the treatment of insomnia.