amine position and Series II at the primary amine sub- 6. Greenblatt HM, Kryger G, Lewis T, Silman I, Sussman JL stituted position. Both enantiomers were combined suc (1999 FEBS Lett463:321-326 cessionally to consider the chiral influence of each 7. Zhang RW, Tang XC, Han YY, Sang GW, Zhang YD, Ma YX, Zhang CL, Yang RM (1991) Acta Pharmacol Sin 12: 250-252 monomer fully Spacer lengths of the carbon bridge ranged 8.Ford JM, Truman CA, Wilcock GK, Roberts CJC(1993)Clin from two to 14-carbons The conformation of ache was harmacol Ther 53: 691-69 assumed to be open to allow the entrance of these bulky 9 Hoskin P), Hanks gw(991) Drugs 41: 326-344 bis-ligands with long chain linkages Pharmacol 79: 191-199 e Table an sti ths shils coar ter tethys bi MEp I. ER -S7 Haroun E, Latimer N (19%6) J Pharm Pharmacol derivatives. Remarkably improved score values, the top 12. Chen Y(2004)Studies on the synthesis, resolution and optical wo of which almost exceed that of E2020. suggested the somers of meptazinol. Dissertation, Shanghai, Fudan Uni- potential optimization of potency. Interaction analysis 13. Kuntz ID(1992) Science 257: 1078-1082 (pictures omitted) showed that bis-MEP derivatives made 14. Drews J(2000) Science 287: 1960-196 good hydrophobic interactions with the residue Trp279 at 15. Morris GM, Goodsell DS, Halliday RS, Huey R, Hart We the PAs. Comparing the chirality of the two series, we Belew RK, Olson AJ(1998)J Comput Chem 19: 1639-16 found that compounds derived from()-S-MEP with the s, 16. Kuntz ID, Blaney JM, Oatley SJ, Langidge R, Ferrin TE (1982) J Mol Biol 161: 269-288 S-conformation of Series I and the SR, RS-conformation of 17. Rarey M, Kramer B, Lengauer T, Klebe GA(1996)J Mol Biol Series Il dominated at the top of the list, which conformed 261:470489 mEP derivatives with a spacer of two to seven showed high Mainz DT, Repasky MP. Knoll EH, Shel ry求k,Shaw 1739-1749 score values, among which 3-and 7-methylene compounds 19. Jones G. Willett P, Glen RC, Leach AR, R(1997)J Mol were found to be the favored candidates. Under the Biol 267: 727-748 guidance of these results, synthesis of the corresponding 20. Hu x, Balaz s, Shelver WH(2004)JMol Graph Model compounds is underway 21. Muegge I, Martin YC(1999)J 2. Kontoyianni M, McClellan LM s Chem Conclusions Greenblatt HM, Dvir H, Silman I, Sussman JL (2003)J Mol Neurosci 20: 369-383 In this paper, we have established a useful docking method 24. Berman HM, Westbrook J, Feng Z, Gilliland G, Bhat TN to predict the binding poses of AChE inhibitors. Using the essig H, shindyalov IN, Bourne Pe (2000) Nucleic Acids Res28:235-242 GOLD docking protocol, the binding conformation and 25 Kryger G, Sliman L, Sussman JL(1999)Structure 7: 297- orientation of()-MEP with TcAChE was illuminated and 26 Bartolucci C, Perola e, Pilger C, Lamba D the potentially preferable bis-MEP derivative candidates screenin To 27. Felder CE, Harel M, Silman 1, Sussman JL (2002)Acta his is the first exploration of the mechanism of action of 28. Ravelli RBG, Raves ML, Ren Z, Bourgeois D, Roth M, Kroon MEP on AChE. Synthesis and biological evaluation of J, Siman I, Sussman JL(1998)Acta Crystallogr Sect these selected analogues are currently underway and the results will be reported in the due course. Preliminary :9. Raves ML, Harel M, Pang YP, Silman I, Kozikowski AP usman JL (1997) Nat Struct Biol 4: 57-63 iological results implied that part of the candidates 30 Dvir H, Wong DM, Harel M, Barril X, Orozco M, Munoz- showed higher AChE inhibition than the core molecule Torrero FJ, Luque D, Camps P, Rosenberry TL, Silman L, )MEP. Although further biological results are needed to Sussman JL (2002) Biochemistry 41: 2970-2981 attest the actual predictive power of this method, the 31 Harel M. Schalk L Ehret-Sabatier L, Bouet F, Goeldner M present study provides an altemate tool for structural Acad Sci90:9031-9035 optimization of (-)-MEP as new AChE inhibitors 32 SYBYL, version 6.9(2002) Tripos Inc, St. Louis, MO, USA 33. GOLD, version 2.1.(2004) Cambridge Crystallographic Data Centre. Cambridge. UK Acknowledgement We gratefully acknowledge financial support 34 CORINA, version 3.0.(2004)Molecular Networks GmbH, from the National Natural science Foundation of china 35. Gasteiger J, Marsili M(1980) Tetrahedron 36: 3219-3228 36 MDL 5.0(2002)MDL Information Systems Inc, San Leandro, A. USA References 37 Gasteiger J, Rudolph C, Sadowski J(1990)Tetrahedron Comp 1. Marchbanks RM(1982)J Neurochem 39: 9-15 38 Pilger C, Bartolucci C, Lamba D, Tropsha A, Fels G(2001)J Mol Graph Model 19: 288-296 2. Coyle JT, Price DL, Delong MR(1983) Science 219: 1184- 39. Sy 4. Kawakami Y, Inoue A, Kawai T, Wakita M, Sugimoto H 40 Carlier PR, Chow ESH, Han YF, Jing Liu J, EI Yazal J, Pang 3. Davis KL, Powchik P(1995) Lancet 345: 625-630 YP(1999) J Med Chen42:42254231 5.EnzA,Boddeke H, Gray J, Spiegel R(1991)Ann NY Acad Sci 41. Pang YP, Quiram P, Jelacic T, Hong E, Brimijoin S(1996)J Hopfinger AJ(1996) Bioorg Med Chem 4: 1429-144 Biol chen271:23646-23649amine position and Series II at the primary amine sub￾stituted position. Both enantiomers were combined suc￾cessionally to consider the chiral influence of each monomer fully. Spacer lengths of the carbon bridge ranged from two to 14-carbons. The conformation of AChE was assumed to be open to allow the entrance of these bulky bis-ligands with long chain linkages. Table 3 lists the chiral character, methylene group length, and fitness scores of the top 15 bis-MEP derivatives. Remarkably improved score values, the top two of which almost exceed that of E2020, suggested the potential optimization of potency. Interaction analysis (pictures omitted) showed that bis-MEP derivatives made good hydrophobic interactions with the residue Trp279 at the PAS. Comparing the chirality of the two series, we found that compounds derived from (−)-S-MEP with the S, S-conformation of Series I and the SR,RS-conformation of Series II dominated at the top of the list, which conformed with the activity of (−)-S-MEP versus (+)-R-MEP. Bis￾MEP derivatives with a spacer of two to seven showed high score values, among which 3- and 7-methylene compounds were found to be the favored candidates. Under the guidance of these results, synthesis of the corresponding compounds is underway. Conclusions In this paper, we have established a useful docking method to predict the binding poses of AChE inhibitors. Using the GOLD docking protocol, the binding conformation and orientation of (−)-MEP with TcAChE was illuminated and the potentially preferable bis-MEP derivative candidates were picked out by virtual screening. To our knowledge, this is the first exploration of the mechanism of action of MEP on AChE. Synthesis and biological evaluation of these selected analogues are currently underway and the results will be reported in the due course. Preliminary biological results implied that part of the candidates showed higher AChE inhibition than the core molecule (−)MEP. 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