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Me-Talnetant and Osanetant Interact within Overlapping but Not Identical Binding Pockets in the Human Tachykinin Neurokinin 3 Receptor Transmembrane Domains

Psychiatry Disease Area (P.M., A.M., C.K., M.-T.Z., J.G.W., W.S.) and Chemistry Discovery (C.B., H.R., C.R.), F. Hoffmann-La Roche Ltd., Basel, Switzerland

Recent clinical trials have indicated that neurokinin 3 receptor antagonists (S)-(+)-N-{{3-[1-benzoyl-3-(3,4-dichlorophenyl)-piperidin-3-yl]prop-1-yl}-4-phenylpiperidin-4-yl}-N-methylacetamine (SR142801; osanetant) and (S)-(-)-N-(-ethylbenzyl)-3-hydroxy-2-phenylquinoline-4-carboxamide (SB223412; talnetant) may treat symptoms of schizophrenia. Using site-directed mutagenesis, rhodopsin-based modeling, [³H](S)-(-)-N-(-ethylbenzyl)-3-methoxy-2-phenylquinoline-4-carboxamide (Me-talnetant) and [³H]osanetant binding, and functional Schild analyses, we have demonstrated the important molecular determinants of neurokinin B (NKB), Me-talnetant, and osanetant binding pockets. The residues Asn138^2.57, Asn142^2.61, Leu232^45.49, Tyr315^6.51, Phe342^7.39, and Met346^7.43 were found to be crucial for the NKB binding site. We observed that the M134^2.53A, V169^3.36M, F342^7.39M, and S341^7.38I/F342^7.39M mutations resulted in the complete loss of [³H]Metalnetant and [³H]osanetant binding affinities and also abolished their functional potencies in an NKB-evoked accumulation of [³H]inositol phosphates assay, whereas the mutations V95^1.42A, N142^2.61A, Y315^6.51F, and M346^7.43A behaved differently between the interacting modes of two antagonists. V95^1.42A and M346^7.43A significantly decreased the affinity and potency of Me-talnetant. Y315^6.51F, although not affecting Me-talnetant, led to a significant decrease in affinity and potency of osanetant. The mutation N142^2.61A, which abolished the potency and affinity of osanetant, led to a significant increase in the affinity and potency of Me-talnetant. The proposed docking mode was further validated using (S)-2-(3,5-bis-trifluoromethyl-phenyl)-N-[4-(4-fluoro-2-methyl-phenyl)-6-((S)-4-methanesulfonyl-3-methyl-piperazin-1-yl)-pyridin-3-yl]-N-methyl-isobutyramide (RO49085940), from another chemical class. It is noteworthy that the mutation F342^7.39A caused an 80-fold gain of RO4908594 binding affinity, but the same mutation resulted in the complete loss of the affinity of Me-talnetant and partial loss of the affinity of osanetant. These observations show that the binding pocket of Me-talnetant and osanetant are overlapping, but not identical. Taken together, our data are consistent with the proposed docking modes where Me-talnetant reaches deeply into the pocket formed by transmembrane (TM)1, -2, and -7, whereas osanetant fills the pocket TM3, -5, and -6 with its phenyl-piperidine moiety.

Address correspondence to: Dr. Pari Malherbe, F. Hoffmann-La Roche Ltd., Bldg. 69/333, CH-4070 Basel, Switzerland. E-mail: parichehr.malherbe{at}roche.com