Tacrine was the first drug to be approved for Alzheimer's disease (AD) treatment, acting as a cholinesterase inhibitor. The neuropathological hallmarks of AD are amyloid-rich senile plaques, neurofibrillary tangles, and neuronal degeneration. The portfolio of currently approved drugs for AD includes acetylcholinesterase inhibitors (AChEIs) and N-methyl-d-aspartate (NMDA) receptor antagonist. Squaric acid is a versatile structural scaffold capable to be easily transformed into amide-bearing compounds that feature both hydrogen bond donor and acceptor groups with the possibility to create multiple interactions with complementary sites. Considering the relatively simple synthesis approach and other interesting properties (rigidity, aromatic character, H-bond formation) of squaramide motif, we combined this scaffold with different tacrine-based derivatives. In this study, we developed 21 novel dimers amalgamating squaric acid with either tacrine, 6-chlorotacrine or 7-methoxytacrine representing various AChEIs. All new derivatives were evaluated for their anti-cholinesterase activities, cytotoxicity using HepG2 cell line and screened to predict their ability to cross the blood-brain barrier. In this contribution, we also report in silico studies of the most potent AChE and BChE inhibitors in the active site of these enzymes.

Biomedical Research Centre University Hospital Hradec Kralove Sokolska 581 500 05 Hradec Kralove Czech Republic

Centro de Química Orgánica Lora Tamayo C Juan de la Cierva 3 28006 Madrid Spain

Department of Toxicology and Military Pharmacy Faculty of Military Health Sciences Trebesska 1575 500 01 Hradec Kralove Czech Republic

Laboratory of Medicinal Chemistry Institute of General Organic Chemistry Juan de la Cierva 3 28006 Madrid Spain

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Ferri C.P., Prince M., Brayne C., Brodaty H., Fratiglioni L., Ganguli M., Hall K., Hasegawa K., Hendrie H., Huang Y., et al. Global prevalence of dementia: a Delphi consensus study. Lancet. 2005;366:2112–2117. doi: 10.1016/S0140-6736(05)67889-0. PubMed DOI PMC

Long J.Z., Cravatt B.F. The Metabolic Serine Hydrolases and Their Functions in Mammalian Physiology and Disease. Chem. Rev. 2011;111:6022–6063. doi: 10.1021/cr200075y. PubMed DOI PMC

Bartus R.T., Dean R.L., Beer B., Lippa A.S. The cholinergic hypothesis of geriatric memory dysfunction. Science. 1982;217:408–414. doi: 10.1126/science.7046051. PubMed DOI

Zemek F., Drtinova L., Nepovimova E., Sepsova V., Korabecny J., Klimes J., Kuca K. Outcomes of Alzheimer’s disease therapy with acetylcholinesterase inhibitors and memantine. Expert Opin. Drug Saf. 2014;13:759–774. PubMed

Birks J. Cholinesterase inhibitors for Alzheimer’s disease. Cochrane Database Syst. Rev. 2006:CD005593. PubMed PMC

Vickers J.C., Mitew S., Woodhouse A., Fernandez-Martos C.M., Kirkcaldie M.T., Canty A.J., McCormack G.H., King A.E. Defining the earliest pathological changes of Alzheimer’s disease. Curr. Alzheimer Res. 2016;13:281–287. doi: 10.2174/1567205013666151218150322. PubMed DOI PMC

Kim G.H., Kim J.E., Rhie S.J., Yoon S. The Role of Oxidative Stress in Neurodegenerative Diseases. Exp. Neurobiol. 2015;24:325–340. doi: 10.5607/en.2015.24.4.325. PubMed DOI PMC

Wang R., Reddy P.H. Role of Glutamate and NMDA Receptors in Alzheimer’s Disease. J. Alzheimers Dis. JAD. 2017;57:1041–1048. doi: 10.3233/JAD-160763. PubMed DOI PMC

Budimir A. Metal ions, Alzheimer’s disease and chelation therapy. Acta Pharm. Zagreb Croat. 2011;61:1–14. doi: 10.2478/v10007-011-0006-6. PubMed DOI

Heneka M.T., Carson M.J., El Khoury J., Landreth G.E., Brosseron F., Feinstein D.L., Jacobs A.H., Wyss-Coray T., Vitorica J., Ransohoff R.M., et al. Neuroinflammation in Alzheimer’s Disease. Lancet Neurol. 2015;14:388–405. doi: 10.1016/S1474-4422(15)70016-5. PubMed DOI PMC

Sonkusare S.K., Kaul C.L., Ramarao P. Dementia of Alzheimer’s disease and other neurodegenerative disorders--memantine, a new hope. Pharmacol. Res. 2005;51:1–17. doi: 10.1016/j.phrs.2004.05.005. PubMed DOI

Crismon M.L. Tacrine: first drug approved for Alzheimer’s disease. Ann. Pharmacother. 1994;28:744–751. doi: 10.1177/106002809402800612. PubMed DOI

Horak M., Holubova K., Nepovimova E., Krusek J., Kaniakova M., Korabecny J., Vyklicky L., Kuca K., Stuchlik A., Ricny J., et al. The pharmacology of tacrine at N-methyl-d-aspartate receptors. Prog. Neuropsychopharmacol. Biol. Psychiatry. 2017;75:54–62. doi: 10.1016/j.pnpbp.2017.01.003. PubMed DOI

Soukup O., Jun D., Zdarova-Karasova J., Patocka J., Musilek K., Korabecny J., Krusek J., Kaniakova M., Sepsova V., Mandikova J., et al. A resurrection of 7-MEOTA: a comparison with tacrine. Curr. Alzheimer Res. 2013;10:893–906. doi: 10.2174/1567205011310080011. PubMed DOI

Lahiri D.K., Lewis S., Farlow M.R. Tacrine alters the secretion of the beta-amyloid precursor protein in cell lines. J. Neurosci. Res. 1994;37:777–787. doi: 10.1002/jnr.490370612. PubMed DOI

Lahiri D.K., Farlow M.R., Sambamurti K. The secretion of amyloid beta-peptides is inhibited in the tacrine-treated human neuroblastoma cells. Brain Res. Mol. Brain Res. 1998;62:131–140. doi: 10.1016/S0169-328X(98)00236-8. PubMed DOI

Watkins P.B., Zimmerman H.J., Knapp M.J., Gracon S.I., Lewis K.W. Hepatotoxic effects of tacrine administration in patients with Alzheimer’s disease. JAMA. 1994;271:992–998. doi: 10.1001/jama.1994.03510370044030. PubMed DOI

Zeiger E., Erexson G., Mortelmans K., Thilagar A. Genetic toxicity studies of 1,2,3,4-tetrahydro-9-acridinamine (tacrine) Mutat. Res. 1997;393:189–197. doi: 10.1016/S1383-5718(97)00096-X. PubMed DOI

Misik J., Nepovimova E., Pejchal J., Kassa J., Korabecny J., Soukup O. Cholinesterase Inhibitor 6-Chlorotacrine—In Vivo Toxicological Profile and Behavioural Effects. [(accessed on 2 August 2019)]; Available online: http://www.eurekaselect.com/158225/article. PubMed

Pang Y.P., Quiram P., Jelacic T., Hong F., Brimijoin S. Highly potent, selective, and low cost bis-tetrahydroaminacrine inhibitors of acetylcholinesterase. Steps toward novel drugs for treating Alzheimer’s disease. J. Biol. Chem. 1996;271:23646–23649. doi: 10.1074/jbc.271.39.23646. PubMed DOI

Recanatini M., Cavalli A., Belluti F., Piazzi L., Rampa A., Bisi A., Gobbi S., Valenti P., Andrisano V., Bartolini M., et al. SAR of 9-amino-1,2,3,4-tetrahydroacridine-based acetylcholinesterase inhibitors: synthesis, enzyme inhibitory activity, QSAR, and structure-based CoMFA of tacrine analogues. J. Med. Chem. 2000;43:2007–2018. doi: 10.1021/jm990971t. PubMed DOI

Carlier P.R., Han Y.F., Chow E.S., Li C.P., Wang H., Lieu T.X., Wong H.S., Pang Y.P. Evaluation of short-tether bis-THA AChE inhibitors. A further test of the dual binding site hypothesis. Bioorg. Med. Chem. 1999;7:351–357. doi: 10.1016/S0968-0896(98)00213-2. PubMed DOI

Korábečný J. Prokognitivní Potenciál Bis(7)-takrinu Jako Zvažovaného Terapeutika Neurodegenerativních Onemocnění. MMSL. 2018;87:34–44. doi: 10.31482/mmsl.2018.006. DOI

Chauhan P., Mahajan S., Kaya U., Hack D., Enders D. Bifunctional Amine-Squaramides: Powerful Hydrogen-Bonding Organocatalysts for Asymmetric Domino/Cascade Reactions. Adv. Synth. Catal. 2015;357:253–281. doi: 10.1002/adsc.201401003. DOI

Zhao B.-L., Li J.-H., Du D.-M. Squaramide-Catalyzed Asymmetric Reactions. Chem. Rec. 2017;17:994–1018. doi: 10.1002/tcr.201600140. PubMed DOI

Karahan S., Tanyeli C. Squaramide catalyzed α-chiral amine synthesis. Tetrahedron Lett. 2018;59:3725–3737. doi: 10.1016/j.tetlet.2018.08.034. DOI

Brandão P., Burke A.J. Recent advances in the asymmetric catalytic synthesis of chiral 3-hydroxy and 3-aminooxindoles and derivatives: Medicinally relevant compounds. Tetrahedron. 2018;74:4927–4957. doi: 10.1016/j.tet.2018.06.015. DOI

Kinney W.A., Abou-Gharbia M., Garrison D.T., Schmid J., Kowal D.M., Bramlett D.R., Miller T.L., Tasse R.P., Zaleska M.M., Moyer J.A. Design and Synthesis of [2-(8,9-Dioxo-2,6-diazabicyclo[5.2.0]non-1(7)-en-2-yl)- ethyl]phosphonic Acid (EAA-090), a Potent N-Methyl-d-aspartate Antagonist, via the Use of 3-Cyclobutene-1,2-dione as an Achiral α-Amino Acid Bioisostere. J. Med. Chem. 1998;41:236–246. doi: 10.1021/jm970504g. PubMed DOI

Dwyer M.P., Yu Y., Chao J., Aki C., Chao J., Biju P., Girijavallabhan V., Rindgen D., Bond R., Mayer-Ezel R., et al. Discovery of 2-Hydroxy-N,N-dimethyl-3-{2-[[(R)-1-(5- methylfuran-2-yl)propyl]amino]-3,4-dioxocyclobut-1-enylamino}benzamide (SCH 527123):  A Potent, Orally Bioavailable CXCR2/CXCR1 Receptor Antagonist. J. Med. Chem. 2006;49:7603–7606. doi: 10.1021/jm0609622. PubMed DOI

Storer R.I., Aciro C., Jones L.H. Squaramides: physical properties, synthesis and applications. Chem. Soc. Rev. 2011;40:2330–2346. doi: 10.1039/c0cs00200c. PubMed DOI

Marín C., Ximenis M., Ramirez-Macías I., Rotger C., Urbanova K., Olmo F., Martín-Escolano R., Rosales M.J., Cañas R., Gutierrez-Sánchez R., et al. Effective anti-leishmanial activity of minimalist squaramide-based compounds. Exp. Parasitol. 2016;170:36–49. doi: 10.1016/j.exppara.2016.07.013. PubMed DOI

Ribeiro C.J.A., Espadinha M., Machado M., Gut J., Gonçalves L.M., Rosenthal P.J., Prudêncio M., Moreira R., Santos M.M.M. Novel squaramides with in vitro liver stage antiplasmodial activity. Bioorg. Med. Chem. 2016;24:1786–1792. doi: 10.1016/j.bmc.2016.03.005. PubMed DOI

Martín-Escolano R., Marín C., Vega M., Martin-Montes Á., Medina-Carmona E., López C., Rotger C., Costa A., Sánchez-Moreno M. Synthesis and biological evaluation of new long-chain squaramides as anti-chagasic agents in the BALB/c mouse model. Bioorg. Med. Chem. 2019;27:865–879. doi: 10.1016/j.bmc.2019.01.033. PubMed DOI

Fu H., Li W., Luo J., Lee N.T.K., Li M., Tsim K.W.K., Pang Y., Youdim M.B.H., Han Y. Promising anti-Alzheimer’s dimer bis(7)-tacrine reduces β-amyloid generation by directly inhibiting BACE-1 activity. Biochem. Biophys. Res. Commun. 2008;366:631–636. doi: 10.1016/j.bbrc.2007.11.068. PubMed DOI

Li C., Carlier P.R., Ren H., Kan K.K.W., Hui K., Wang H., Li W., Li Z., Xiong K., Clement E.C., et al. Alkylene tether-length dependent γ-aminobutyric acid type A receptor competitive antagonism by tacrine dimers. Neuropharmacology. 2007;52:436–443. doi: 10.1016/j.neuropharm.2006.07.039. PubMed DOI

Ros E., Aleu J., Gomez De Aranda I., Cantí C., Pang Y.-P., Marsal J., Solsona C. Effects of Bis(7)-Tacrine on Spontaneous Synaptic Activity and on the Nicotinic ACh Receptor of Torpedo Electric Organ. J. Neurophysiol. 2001;86:183–189. doi: 10.1152/jn.2001.86.1.183. PubMed DOI

Minarini A., Milelli A., Tumiatti V., Rosini M., Simoni E., Bolognesi M.L., Andrisano V., Bartolini M., Motori E., Angeloni C., et al. Cystamine-tacrine dimer: A new multi-target-directed ligand as potential therapeutic agent for Alzheimer’s disease treatment. Neuropharmacology. 2012;62:997–1003. doi: 10.1016/j.neuropharm.2011.10.007. PubMed DOI

Han Y.-F., Wu D.-C., Xiao X.-Q., Chen P.M.Y., Chung W., Lee N.T.K., Pang Y.-P., Carlier P.R. Protection against ischemic injury in primary cultured astrocytes of mouse cerebral cortex by bis(7)-tacrine, a novel anti-Alzheimer’s agent. Neurosci. Lett. 2000;288:95–98. doi: 10.1016/S0304-3940(00)01198-8. PubMed DOI

Zhao Y., Li W., Chow P.C.Y., Lau D.T.K., Lee N.T.K., Pang Y., Zhang X., Wang X., Han Y. Bis(7)-tacrine, a promising anti-Alzheimer’s dimer, affords dose- and time-dependent neuroprotection against transient focal cerebral ischemia. Neurosci. Lett. 2008;439:160–164. doi: 10.1016/j.neulet.2008.05.007. PubMed DOI

Li J., Lu Z., Xu L., Wang Q., Zhang Z., Fang J. Neuroprotective effects of bis(7)-tacrine in a rat model of pressure-induced retinal ischemia. Cell Biochem. Biophys. 2014;68:275–282. doi: 10.1007/s12013-013-9707-4. PubMed DOI

Xiao X.Q., Lee N.T., Carlier P.R., Pang Y., Han Y.F. Bis(7)-tacrine, a promising anti-Alzheimer’s agent, reduces hydrogen peroxide-induced injury in rat pheochromocytoma cells: comparison with tacrine. Neurosci. Lett. 2000;290:197–200. doi: 10.1016/S0304-3940(00)01357-4. PubMed DOI

Li W., Xue J., Niu C., Fu H., Lam C.S.C., Luo J., Chan H.H.N., Xue H., Kan K.K.W., Lee N.T.K., et al. Synergistic Neuroprotection by Bis(7)-tacrine via Concurrent Blockade of N-Methyl-d-aspartate Receptors and Neuronal Nitric-Oxide Synthase. Mol. Pharmacol. 2007;71:1258–1267. doi: 10.1124/mol.106.029108. PubMed DOI

Li W., Lee N.T.K., Fu H., Kan K.K.W., Pang Y.-P., Li M., Tsim K.W.K., Li X., Han Y. Neuroprotection via inhibition of nitric oxide synthase by bis(7)-tacrine. NeuroReport. 2006;17:471–474. doi: 10.1097/01.wnr.0000209014.09094.72. PubMed DOI

Li W., Pi R., Chan H.H.N., Fu H., Lee N.T.K., Tsang H.W., Pu Y., Chang D.C., Li C., Luo J., et al. Novel dimeric acetylcholinesterase inhibitor bis7-tacrine, but not donepezil, prevents glutamate-induced neuronal apoptosis by blocking N-methyl-D-aspartate receptors. J. Biol. Chem. 2005;280:18179–18188. doi: 10.1074/jbc.M411085200. PubMed DOI

Nepovimova E., Korabecny J., Dolezal R., Babkova K., Ondrejicek A., Jun D., Sepsova V., Horova A., Hrabinova M., Soukup O., et al. Tacrine–Trolox Hybrids: A Novel Class of Centrally Active, Nonhepatotoxic Multi-Target-Directed Ligands Exerting Anticholinesterase and Antioxidant Activities with Low In Vivo Toxicity. J. Med. Chem. 2015;58:8985–9003. doi: 10.1021/acs.jmedchem.5b01325. PubMed DOI

Spilovska K., Korabecny J., Kral J., Horova A., Musilek K., Soukup O., Drtinova L., Gazova Z., Siposova K., Kuca K. 7-Methoxytacrine-adamantylamine heterodimers as cholinesterase inhibitors in Alzheimer’s disease treatment--synthesis, biological evaluation and molecular modeling studies. Molecules. 2013;18:2397–2418. doi: 10.3390/molecules18022397. PubMed DOI PMC

Ellman G.L., Courtney K.D., Andres V., Feather-Stone R.M. A new and rapid colorimetric determination of acetylcholinesterase activity. Biochem. Pharmacol. 1961;7:88–95. doi: 10.1016/0006-2952(61)90145-9. PubMed DOI

Pohanka M., Jun D., Kuca K. Improvement of acetylcholinesterase-based assay for organophosphates in way of identification by reactivators. Talanta. 2008;77:451–454. doi: 10.1016/j.talanta.2008.06.007. PubMed DOI

Sepsova V., Karasova J.Z., Korabecny J., Dolezal R., Zemek F., Bennion B.J., Kuca K. Oximes: inhibitors of human recombinant acetylcholinesterase. A structure-activity relationship (SAR) study. Int. J. Mol. Sci. 2013;14:16882–16900. doi: 10.3390/ijms140816882. PubMed DOI PMC

Pohanka M., Karasova J.Z., Kuca K., Pikula J., Holas O., Korabecny J., Cabal J. Colorimetric dipstick for assay of organophosphate pesticides and nerve agents represented by paraoxon, sarin and VX. Talanta. 2010;81:621–624. doi: 10.1016/j.talanta.2009.12.052. PubMed DOI

Bolognesi M.L., Cavalli A., Valgimigli L., Bartolini M., Rosini M., Andrisano V., Recanatini M., Melchiorre C. Multi-Target-Directed Drug Design Strategy: From a Dual Binding Site Acetylcholinesterase Inhibitor to a Trifunctional Compound against Alzheimer’s Disease. J. Med. Chem. 2007;50:6446–6449. doi: 10.1021/jm701225u. PubMed DOI

Nordberg A., Ballard C., Bullock R., Darreh-Shori T., Somogyi M. A review of butyrylcholinesterase as a therapeutic target in the treatment of Alzheimer’s disease. Prim. Care Companion CNS Disord. 2013;15 doi: 10.4088/PCC.12r01412. PubMed DOI PMC

Greig N.H., Utsuki T., Ingram D.K., Wang Y., Pepeu G., Scali C., Yu Q.-S., Mamczarz J., Holloway H.W., Giordano T., et al. Selective butyrylcholinesterase inhibition elevates brain acetylcholine, augments learning and lowers Alzheimer beta-amyloid peptide in rodent. Proc. Natl. Acad. Sci. USA. 2005;102:17213–17218. doi: 10.1073/pnas.0508575102. PubMed DOI PMC

Nepovimova E., Uliassi E., Korabecny J., Peña-Altamira L.E., Samez S., Pesaresi A., Garcia G.E., Bartolini M., Andrisano V., Bergamini C., et al. Multitarget drug design strategy: quinone-tacrine hybrids designed to block amyloid-β aggregation and to exert anticholinesterase and antioxidant effects. J. Med. Chem. 2014;57:8576–8589. doi: 10.1021/jm5010804. PubMed DOI

Spilovska K., Korabecny J., Horova A., Musilek K., Nepovimova E., Drtinova L., Gazova Z., Siposova K., Dolezal R., Jun D., et al. Design, synthesis and in vitro testing of 7-methoxytacrine-amantadine analogues: a novel cholinesterase inhibitors for the treatment of Alzheimer’s disease. Med. Chem. Res. 2015;24:2645–2655. doi: 10.1007/s00044-015-1316-x. DOI

Muckova L., Pejchal J., Jost P., Vanova N., Herman D., Jun D. Cytotoxicity of acetylcholinesterase reactivators evaluated in vitro and its relation to their structure. Drug Chem. Toxicol. 2019;42:252–256. doi: 10.1080/01480545.2018.1432641. PubMed DOI

Di L., Kerns E.H., Fan K., McConnell O.J., Carter G.T. High throughput artificial membrane permeability assay for blood-brain barrier. Eur. J. Med. Chem. 2003;38:223–232. doi: 10.1016/S0223-5234(03)00012-6. PubMed DOI

Li C., Wainhaus S., Uss A.S., Cheng K.-C. High-Throughput Screening Using Caco-2 Cell and PAMPA Systems. In: Ehrhardt C., Kim K.-J., editors. Drug Absorption Studies: In Situ, In Vitro and In Silico Models. Springer US; Boston, MA, USA: 2008. pp. 418–429. Biotechnology: Pharmaceutical Aspects.

Cheung J., Rudolph M.J., Burshteyn F., Cassidy M.S., Gary E.N., Love J., Franklin M.C., Height J.J. Structures of human acetylcholinesterase in complex with pharmacologically important ligands. J. Med. Chem. 2012;55:10282–10286. doi: 10.1021/jm300871x. PubMed DOI

Nachon F., Carletti E., Ronco C., Trovaslet M., Nicolet Y., Jean L., Renard P.-Y. Crystal structures of human cholinesterases in complex with huprine W and tacrine: elements of specificity for anti-Alzheimer’s drugs targeting acetyl- and butyryl-cholinesterase. Biochem. J. 2013;453:393–399. doi: 10.1042/BJ20130013. PubMed DOI

Trott O., Olson A.J. AutoDock Vina: Improving the speed and accuracy of docking with a new scoring function, efficient optimization and multithreading. J. Comput. Chem. 2010;31:455–461. doi: 10.1002/jcc.21334. PubMed DOI PMC

Hepnarova V., Korabecny J., Matouskova L., Jost P., Muckova L., Hrabinova M., Vykoukalova N., Kerhartova M., Kucera T., Dolezal R., et al. The concept of hybrid molecules of tacrine and benzyl quinolone carboxylic acid (BQCA) as multifunctional agents for Alzheimer’s disease. Eur. J. Med. Chem. 2018;150:292–306. doi: 10.1016/j.ejmech.2018.02.083. PubMed DOI

Rydberg E.H., Brumshtein B., Greenblatt H.M., Wong D.M., Shaya D., Williams L.D., Carlier P.R., Pang Y.-P., Silman I., Sussman J.L. Complexes of alkylene-linked tacrine dimers with Torpedo californica acetylcholinesterase: Binding of Bis5-tacrine produces a dramatic rearrangement in the active-site gorge. J. Med. Chem. 2006;49:5491–5500. doi: 10.1021/jm060164b. PubMed DOI

Bajda M., Więckowska A., Hebda M., Guzior N., Sotriffer C.A., Malawska B. Structure-Based Search for New Inhibitors of Cholinesterases. Int. J. Mol. Sci. 2013;14:5608–5632. doi: 10.3390/ijms14035608. PubMed DOI PMC

Cavalli A., Bolognesi M.L., Minarini A., Rosini M., Tumiatti V., Recanatini M., Melchiorre C. Multi-target-directed ligands to combat neurodegenerative diseases. J. Med. Chem. 2008;51:347–372. doi: 10.1021/jm7009364. PubMed DOI

León R., Garcia A.G., Marco-Contelles J. Recent advances in the multitarget-directed ligands approach for the treatment of Alzheimer’s disease. Med. Res. Rev. 2013;33:139–189. doi: 10.1002/med.20248. PubMed DOI

Proschak E., Stark H., Merk D. Polypharmacology by Design: A Medicinal Chemist’s Perspective on Multitargeting Compounds. J. Med. Chem. 2019;62:420–444. doi: 10.1021/acs.jmedchem.8b00760. PubMed DOI

Mosmann T. Rapid colorimetric assay for cellular growth and survival: Application to proliferation and cytotoxicity assays. J. Immunol. Methods. 1983;65:55–63. doi: 10.1016/0022-1759(83)90303-4. PubMed DOI

Riss T.L., Moravec R.A., Niles A.L., Duellman S., Benink H.A., Worzella T.J., Minor L. Cell Viability Assays. In: Sittampalam G.S., Coussens N.P., Nelson H., Arkin M., Auld D., Austin C., Bejcek B., Glicksman M., Inglese J., Iversen P.W., et al., editors. Assay Guidance Manual. Eli Lilly & Company and the National Center for Advancing Translational Sciences; Bethesda, MD, USA: 2004.

Liu B., Wang L., Jin Y.-H. An effective PSO-based memetic algorithm for flow shop scheduling. IEEE Trans. Syst. Man Cybern. Part B Cybern. Publ. IEEE Syst. Man Cybern. Soc. 2007;37:18–27. doi: 10.1109/TSMCB.2006.883272. PubMed DOI

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