T Liu et al/ Pharmacology, Biochemistry and Behavior 104(2013)138-143 markedly weakened(Supplementary information), because the al- Castro A. Martinez A Targeting beta-amyloid pathogenesis through acetylcholinesterase kane chain length of Bis(6)-(-)-meptazinol was 6 methylene groups. Cavalli A, Bolognesi ML Minarini A, Rosini M, Tumiatti V, Recanatini M, et al. which was appropriate to form the spherical conformation. Multi-target-directed ligands to combat neurodegenerative diseases. J Med Chem After the exploration of the inhibition of Bis-Mep on AChE activity in vitro, we used a model to evaluate the therapeutic effects of Chen y, shoham E, constantini s. weinstocK M Rivastigmine, a brain-sele tive acen Bis-Mep. Scop is known to induce cognitive impairment by ed-head injury in the mouse. J Neurotrauma 1998: 15: 231-7 ade of central muscarinic receptors, and Scop-treated mice have de Bruin N, Pouzet B. Beneficial effects of galantamine on performance in the object been widely used as a pharmacological in vivo model for screening th the retention interval pharmacol biochem belay 2006: 85 253-60 drugs for the amelioration of cognitive symptoms in AD(Buccafusco Du DM, Carlier PR Development of bivalent acetylcholinesterase inhibitors as potentia et al, 2008). A variety of therapeutic AChE inhibitors, such as tacrine eutic drugs for Alzheimers disease. Curr Pharm Des 2004: 10: 3141-56. (Shutske et al, 1989), rivastigmine(Bejar et al, 1999), donepezil Ellis M cholinesterase inhibitors in the treatment of dementia. JAm Osteopath Assoc (Ogura et al, 2000), and Hup A (Xiong et al, 1995)have been evalu- Ellman GL, Courtney KD, Andres jr V, Featherstone RM. A new and rapid colormetric s.c.)significantly ameliorated Scop-induced cognition deficits in mice. Farlow MR Salloway s, Tarochol yardieyersus standard(0m如p组m ated using Scop-treated mice model. Our results showed that Bis-Mep determination of The bell-shaped dose-response manner was also observed which is s disease: a 24-week, randomized, double-blind garded as the typical mode of AChE inhibitors( Chen et al, 1998 ng et al, 1999). Furthermore, significant inhibition of Bis-Mep on Frolich LA Review of the first transdermal treatment for Alzheimers disease-the possesses strong inhibitory effect on AChE activity in the brain and its Ge xoxastigmine patch Eur Neurol Rev 2008: 1: 33 AChE activity in hippocampus provided direct evidence that Bis-Mep Wang x The peripheral side effects are the common reason for many o study. J Chromatogr 2012: 881: 126-30mnetry high BBB permeability chromatography-tandem mass spectrometry: application to pharmacokinetics patients to withdraw their AChEl treatment(Ellis, 2005). However. tt HM, Guillou C, Guenard D, Argaman A, Botti S, Badet B, et al. The complex of a bivalent derivative of galanthamine with torpedo acetylcholinesterase displays no obvious peripheral side effect was found in the various effective doses of Bis-Mep. The maximal cognitive amelioration of Bis-Mep Kryger G, Silman L, Sussman JL Structure of acetylcholinesterase complexed with E2020 100 ng/kg) was much lower than that reported of rivastigmine (Aricept): implications for the design of new anti-Alzheimer drugs. Structure 2 mg/kg)(Chen et al, 1998)and galantamine (10 mg/kg)(de bruin and Pouzet, 2006)in Scop-treated mice model. The powerful efficacy Lee LH. Shineman DW, Fillit HM A diverse portfolio of novel drug discovery efforts of Bis-Mep implied that Bis-Mep might bring more benefits with Alzheimers Drug Discovery. Alzheimers Res Ther 2010: 2: 33- fewer side effects than others in long-time treatment for AD patients. Massoud F. Gauthier S Upda the pharmacological treatment of Alzheimer However, specific long-term toxicological studies are required to con- Munoz- Torrero D. Acetylcholinesterase inhibitors as disease-modifying therapies for firm this latter view Izheimer's disease Curr Med Chem 2008: 15: 2433-55. In conclusion, Bis-Mep as a mixed type AChE inhibitor exhibits Ogura H, Kosasa T, Kuriya Y, Yamanishi Y Donepezil, a centrally acting acetylcholine- nificant efficacy in ameliorating Scop-induced cognitive impairment thibitor, alleviates learning deficits in hypocholinergic models in rats. and inhibiting brain AChE in mice. These current evidences suggested D Williams that it might be a potentially promising dual-binding AChE inhibitor for AD therapeutics rine produces a dramatic rearrangement in the active-site gorge. Supplementarydatatothisarticlecanbefoundonlineathttp:// amacokinetic behavior in plasma, cerebrospinal fluid dx. doi.org/10.1016/pbb201211.009. ral cortex after intranasal administration of hydrochloride meptazinol Acknowledgments disease: do we need new inhibitors. Expert Opin Emerg Drugs 2005: 10: 817-25 velopment Project(No 2009ZX09103 and No 2012ZX09312-003), Na- Terry r Av. Buccafusco I. The cholinergic hypothesis of age and Alzheimer's disease-related cognitive deficits: recent challenges and their implications for novel tional Natural Science Foundation of China(No. 30801393, 30801435 drug development. Pharmacol Exp Ther 2003: 306: 821-7. and 30973509), and Shanghai Municipal Science and Technology Com- TSuno N. thee 20099. 591-8 ng the molecular basis of memory failure inAlzheimer's of patients with Alzheimers disease. Expert Rev mission(No.10431902700) Walsh DM, Selkoe D). Decipherin References le PR, Ho wL, Lee N, Pang YP, Han YF Attenuation of scopolamine-induced rats by bis(7)-tacrine a novel dimeric AChE inhibitor. Zhongguo Yao li Xue Bao 1999: 20: 211 BartoliniM, Bertucci C, Cavrini V, Andrisano V beta-Amyloid aggregation induced by xie Q, Wang H, Xia Zh, Liu MY, Wang X, Fu W,et aL Bis-(-)-nor-meptazinols as novel uman acetylcholinesterase: inhibition studies. Biochem Pharmacol 2003: 65 407-16 with high inhibitory potency on Amyloid-B olamine-induced mem Xiong ZQ, Tang XC, Lin JL Zhu DY. Effects of isovanihuperzine hts into ligand interactions at the polamine-induced memory impairment. Zhongguo Yao Li Xue Bao 1995: 16: Buccafusco I, Terry Jr AV, Webster S). Martin D, Hohnadel EJ, Bouchard KA, et aL IK, Cui YY, Wang H, Zhu L Niu YY, et al Differential omputer-assisted operant-conditioning memory tasks for screening drug candi- anisodamine versus scopolamine. Neurosci Lett 2008: 443: 241-5 dates. Psychopharmacology(Berl)2008 199: 481-94.markedly weakened (Supplementary information), because the al￾kane chain length of Bis(6)-(−)-meptazinol was 6 methylene groups, which was appropriate to form the spherical conformation. After the exploration of the inhibition of Bis-Mep on AChE activity in vitro, we used a model to evaluate the therapeutic effects of Bis-Mep. Scop is known to induce cognitive impairment by the block￾ade of central muscarinic receptors, and Scop-treated mice have been widely used as a pharmacological in vivo model for screening drugs for the amelioration of cognitive symptoms in AD (Buccafusco et al., 2008). A variety of therapeutic AChE inhibitors, such as tacrine (Shutske et al., 1989), rivastigmine (Bejar et al., 1999), donepezil (Ogura et al., 2000), and Hup A (Xiong et al., 1995) have been evalu￾ated using Scop-treated mice model. Our results showed that Bis-Mep (s.c.) significantly ameliorated Scop-induced cognition deficits in mice. The bell-shaped dose–response manner was also observed which is regarded as the typical mode of AChE inhibitors (Chen et al., 1998; Wang et al., 1999). Furthermore, significant inhibition of Bis-Mep on AChE activity in hippocampus provided direct evidence that Bis-Mep possesses strong inhibitory effect on AChE activity in the brain and its high BBB permeability. The peripheral side effects are the common reason for many patients to withdraw their AChEI treatment (Ellis, 2005). However, no obvious peripheral side effect was found in the various effective doses of Bis-Mep. The maximal cognitive amelioration of Bis-Mep (100 ng/kg) was much lower than that reported of rivastigmine (2 mg/kg) (Chen et al., 1998) and galantamine (10 mg/kg) (de Bruin and Pouzet, 2006) in Scop-treated mice model. The powerful efficacy of Bis-Mep implied that Bis-Mep might bring more benefits with fewer side effects than others in long-time treatment for AD patients. However, specific long-term toxicological studies are required to con- firm this latter view. In conclusion, Bis-Mep as a mixed type AChE inhibitor exhibits sig￾nificant efficacy in ameliorating Scop-induced cognitive impairment and inhibiting brain AChE in mice. These current evidences suggested that it might be a potentially promising dual-binding AChE inhibitor for AD therapeutics. Supplementary data to this article can be found online at http:// dx.doi.org/10.1016/j.pbb.2012.11.009. Acknowledgments Funding for this study was supported by the National Basic Research Program of China (No. 2010CB529806), National Innovative Drug De￾velopment Project (No. 2009ZX09103 and No. 2012ZX09312-003), Na￾tional Natural Science Foundation of China (No. 30801393, 30801435 and 30973509), and Shanghai Municipal Science and Technology Com￾mission (No. 10431902700). References Bartolini M, Bertucci C, Cavrini V, Andrisano V. beta-Amyloid aggregation induced by human acetylcholinesterase: inhibition studies. Biochem Pharmacol 2003;65: 407–16. Bejar C, Wang RH, Weinstock M. Effect of rivastigmine on scopolamine-induced mem￾ory impairment in rats. Eur J Pharmacol 1999;383:231–40. Bourne Y, Taylor P, Radic Z, Marchot P. Structural insights into ligand interactions at the acetylcholinesterase peripheral anionic site. EMBO J 2003;22:1-12. Buccafusco JJ, Terry Jr AV, Webster SJ, Martin D, Hohnadel EJ, Bouchard KA, et al. The scopolamine-reversal paradigm in rats and monkeys: the importance of computer-assisted operant-conditioning memory tasks for screening drug candi￾dates. Psychopharmacology (Berl) 2008;199:481–94. Castro A, Martinez A. Targeting beta-amyloid pathogenesis through acetylcholinesterase inhibitors. Curr Pharm Des 2006;12:4377–87. Cavalli A, Bolognesi ML, Minarini A, Rosini M, Tumiatti V, Recanatini M, et al. Multi-target-directed ligands to combat neurodegenerative diseases. J Med Chem 2008;51:347–72. Chen Y, Shohami E, Constantini S, Weinstock M. Rivastigmine, a brain-selective acetyl￾cholinesterase inhibitor, ameliorates cognitive and motor deficits induced by closed-head injury in the mouse. J Neurotrauma 1998;15:231–7. de Bruin N, Pouzet B. Beneficial effects of galantamine on performance in the object recognition task in Swiss mice: deficits induced by scopolamine and by prolonging the retention interval. Pharmacol Biochem Behav 2006;85:253–60. Du DM, Carlier PR. Development of bivalent acetylcholinesterase inhibitors as potential therapeutic drugs for Alzheimer's disease. Curr Pharm Des 2004;10:3141–56. Ellis JM. Cholinesterase inhibitors in the treatment of dementia. J Am Osteopath Assoc 2005;1053:145–58. Ellman GL, Courtney KD, Andres Jr V, Featherstone RM. A new and rapid colormetric determination of acetylcholinesterase activity. Biochem Pharmacol 1961;7:88–95. Farlow MR, Salloway S, Tariot PN, Yardley J, Moline ML, Wang Q, et al. Effectiveness and tolerability of high-dose (23 mg/d) versus standard-dose (10 mg/d) donepezil in moderate to severe Alzheimer's disease: a 24-week, randomized, double-blind study. Clin Ther 2010;32:1234–51. Frölich LA. Review of the first transdermal treatment for Alzheimer's disease—the Rivastigmine patch. Eur Neurol Rev 2008;1:33–5. Ge XX, Wang XL, Jiang P, Xie Y, Jiang T, Rong ZX, et al. Determination of Bis(9)-(−)-Meptazinol, a bis-ligand for Alzheimer's disease, in rat plasma by liquid chromatography–tandem mass spectrometry: application to pharmacokinetics study. J Chromatogr 2012;881:126–30. Greenblatt HM, Guillou C, Guénard D, Argaman A, Botti S, Badet B, et al. The complex of a bivalent derivative of galanthamine with torpedo acetylcholinesterase displays drastic deformation of the active-site gorge: implications for structure-based drug design. J Am Chem Soc 2004;126:15405–11. Kryger G, Silman I, Sussman JL. Structure of acetylcholinesterase complexed with E2020 (Aricept): implications for the design of new anti-Alzheimer drugs. Structure 1999;7:297–307. Lee LH, Shineman DW, Fillit HM. A diverse portfolio of novel drug discovery efforts for Alzheimer's disease: meeting report from the 11th International Conference on Alzheimer's Drug Discovery. Alzheimers Res Ther 2010;2:33–6. Massoud F, Gauthier S. Update on the pharmacological treatment of Alzheimer's disease. Curr Neuropharmacol 2010;8:69–80. Muñoz-Torrero D. Acetylcholinesterase inhibitors as disease-modifying therapies for Alzheimer's disease. Curr Med Chem 2008;15:2433–55. Ogura H, Kosasa T, Kuriya Y, Yamanishi Y. Donepezil, a centrally acting acetylcholines￾terase inhibitor, alleviates learning deficits in hypocholinergic models in rats. Methods Find Exp Clin Pharmacol 2000;22:89–95. Rydberg EH, Brumshtein B, Greenblatt HM, Wong DM, Shaya D, Williams LD, et al. Com￾plexes of alkylenelinked tacrine dimers with Torpedo californica acetylcholinesterase: binding of bis(5)-tacrine produces a dramatic rearrangement in the active-site gorge. J Med Chem 2006;49:5491–500. Shi ZQ, Zhang QZ, Jiang XG. Pharmacokinetic behavior in plasma, cerebrospinal fluid and cerebral cortex after intranasal administration of hydrochloride meptazinol. Life Sci 2005;77:2574–83. Shutske GM, Pierrat FA, Kapples KJ, Cornfeldt ML, Szewczak MR, Huger FP, et al. 9-Amino-1,2,3,4-tetrahydroacridin-1-ols:synthesis and evaluation as potential Alzheimer's disease therapeutics. J Med Chem 1989;32:1805–13. Small DH. Acetylcholinesterase inhibitors for the treatment of dementia in Alzheimer's disease: do we need new inhibitors. Expert Opin Emerg Drugs 2005;10:817–25. Terry Jr AV, Buccafusco JJ. The cholinergic hypothesis of age and Alzheimer's disease-related cognitive deficits: recent challenges and their implications for novel drug development. J Pharmacol Exp Ther 2003;306:821–7. Tsuno N. Donepezil in the treatment of patients with Alzheimer's disease. Expert Rev Neurother 2009;9:591–8. Walsh DM, Selkoe DJ. Deciphering the molecular basis of memory failure in Alzheimer's disease. Neuron 2004;44:181–93. Wang H, Carlie PR, Ho WL, Lee NT, Pang YP, Han YF. Attenuation of scopolamine-induced deficits in navigational memory performance in rats by bis(7)-tacrine,a novel dimeric AChE inhibitor. Zhongguo Yao Li Xue Bao 1999;20:211–7. Xie Q, Wang H, Xia Zh, Liu MY, Wang X, Fu W, et al. Bis-(−)-nor-meptazinols as novel nanomolar cholinesterase inhibitors with high inhibitory potency on Amyloid-β aggregation. J Med Chem 2008;51:2027–36. Xiong ZQ, Tang XC, Lin JL, Zhu DY. Effects of isovanihuperzine A on cholinesterase and scopolamine-induced memory impairment. Zhongguo Yao Li Xue Bao 1995;16: 21–5. Zhang WW, Song MK, Cui YY, Wang H, Zhu L, Niu YY, et al. Differential neuropsychopharmacological influences of naturally occurring tropane alkaloids anisodamine versus scopolamine. Neurosci Lett 2008;443:241–5. T. Liu et al. / Pharmacology, Biochemistry and Behavior 104 (2013) 138–143 143