Planta Med 2009; 75(3): 195-204
DOI: 10.1055/s-0028-1088397
Pharmacology
Original Paper
© Georg Thieme Verlag KG Stuttgart · New York

In silico Target Fishing for Rationalized Ligand Discovery Exemplified on Constituents of Ruta graveolens

Judith M. Rollinger1 , Daniela Schuster2 , Birgit Danzl1 , Stefan Schwaiger1 , Patrick Markt2 , Michaela Schmidtke3 , Jürg Gertsch4 , Stefan Raduner4 , Gerhard Wolber2 , Thierry Langer2 , Hermann Stuppner1
  • 1Institute of Pharmacy, Pharmacognosy, and Center for Molecular Biosciences, University of Innsbruck, 6020 Innsbruck, Austria
  • 2Institute of Pharmacy, Pharmaceutical Chemistry, and Center for Molecular Biosciences, University of Innsbruck, Innsbruck, Austria
  • 3Institute of Virology and Antiviral Therapy, Friedrich Schiller University, Jena, Germany
  • 4Department of Chemistry and Applied Biosciences, ETH Zurich, Zürich, Switzerland
Further Information

Publication History

Received: July 3, 2008 Revised: October 8, 2008

Accepted: October 27, 2008

Publication Date:
18 December 2008 (online)

Abstract

The identification of targets whose interaction is likely to result in the successful treatment of a disease is of growing interest for natural product scientists. In the current study we performed an exemplary application of a virtual parallel screening approach to identify potential targets for 16 secondary metabolites isolated and identified from the aerial parts of the medicinal plant Ruta graveolens L. Low energy conformers of the isolated constituents were simultaneously screened against a set of 2208 pharmacophore models generated in-house for the in silico prediction of putative biological targets, i. e., target fishing. Based on the predicted ligand-target interactions, we focused on three biological targets, namely acetylcholinesterase (AChE), the human rhinovirus (HRV) coat protein and the cannabinoid receptor type-2 (CB2). For a critical evaluation of the applied parallel screening approach, virtual hits and non-hits were assayed on the respective targets. For AChE the highest scoring virtual hit, arborinine, showed the best inhibitory in vitro activity on AChE (IC50 34.7 μM). Determination of the anti-HRV-2 effect revealed 6,7,8-trimethoxycoumarin and arborinine to be the most active antiviral constituents with IC50 values of 11.98 μM and 3.19 μM, respectively. Of these, arborinine was predicted virtually. Of all the molecules subjected to parallel screening, one virtual CB2 ligand was obtained, i. e., rutamarin. Interestingly, in experimental studies only this compound showed a selective activity to the CB2 receptor (Ki of 7.4 μM) by using a radioligand displacement assay. The applied parallel screening paradigm with constituents of R. graveolens on three different proteins has shown promise as an in silico tool for rational target fishing and pharmacological profiling of extracts and single chemical entities in natural product research.

Abbreviations

AChE:acetylcholinesterase

CD2:cannabinoid receptor type-2

CC50:50 % cytotoxic concentration

CPE:cytopathic effect

3D:three dimensional

GNT:galanthamine

HRV:human rhinovirus

PDB:protein databank

VS:virtual screening

References

  • 1 Ekins S, Mestres J, Testa B. In silico pharmacology for drug discovery: Methods for virtual ligand screening and profiling.  Br J Pharmacol. 2007;  152 9-20
  • 2 Ekins S, Mestres J, Testa B. In silico pharmacology for drug discovery: Applications to targets and beyond.  Br J Pharmacol. 2007;  152 21-37
  • 3 Kirchmair H, Distinto S, Schuster D, Spitzer G, Langer T, Wolber T. Enhancing drug discovery through in-silico screening: Strategies to increase true positives retrieval rates.  Curr Med Chem. 2008;  51 7021-40
  • 4 Rollinger J M, Langer T, Stuppner H. Integrated in silico tools to exploit the natural products’ bioactivity.  Planta Med. 2006;  72 671-8
  • 5 Rollinger J M, Stuppner H, Langer T. Virtual screening for the discovery of bioactive natural products. In: Petersen F, Amstutz R, editors. Natural compounds as drugs, Vol. I.  Basel: Birkhäuser. Verlag;  2008 212-49
  • 6 Steindl T M, Schuster D, Wolber G, Laggner C, Langer T. High-throughput structure-based pharmacophore modelling as a basis for successful parallel virtual screening.  J Comput Aid Mol Des. 2006;  20 703-15
  • 7 Steindl T M, Schuster D, Laggner C, Langer T. Parallel screening: a novel concept in pharmacophore modeling and virtual screening.  J Chem Inf Model. 2006;  46 146-57
  • 8 Langer T, Hoffmann R D, Mannhold R, Kubinyi H. Pharmacophores and pharmacophore searches (Methods and Principles in Medicinal Chemistry). Weinheim; Wiley-VCH 2006
  • 9 Moldenke H M, Moldenke A L. Plants of the Bible. Waltham (Massachusettes); Chronica Botanica Co 1952: 208
  • 10 Foster S, Tyler V E. Tyler’s honest herbal: a sensible guide to the use of herbs and related remedies, 4th edition. Binghampton; The Haworth Herbal Press 1999: 325-6
  • 11 Rollinger J M, Mock P, Zidorn C, Ellmerer E P, Langer T, Stuppner H. Application of the in combo screening approach for the discovery of non-alkaloid acetylcholinesterase inhibitors from Cichorium intybus. .  Curr Drug Discov Technol 2005; 2 : 185 – 93 (erratum:. 2006;  3 89)
  • 12 Rollinger J M, Schuster D, Baier E, Ellmerer E P, Langer T, Stuppner H. Taspine: bioactivity-guided isolation and molecular ligand-target insight of a potent acetylcholinesterase inhibitor from Magnolia × soulangiana. .  J Nat Prod. 2006;  69 1341-6
  • 13 Wermuth C G, Ganellin C R, Lindberg P, Mitscher L A. Glossary of terms used in medicinal chemistry (IUPAC recommendations 1997).  Ann Rep Med Chem. 1998;  33 385-95
  • 14 Ellman G L, Courtney D, Andres V, Featherstone R M. A new and rapid calorimetric determination of acetylcholinesterase activity.  Biochem Pharmacol. 1961;  7 88-95
  • 15 Ingkaninan K, de Best C M, van der Heijden R, Hofte A JP, Karabatakb B, Irth H. High-performance liquid chromatography with on-line coupled UV, mass spectrometric and biochemical detection for identification of acetylcholinesterase inhibitors from natural products.  J Chromatogr A. 2000;  872 61-73
  • 16 Rollinger J M, Hornick A, Langer T, Stuppner H, Prast H. Acetylcholinesterase inhibitory activity of scopolin and scopoletin discovered by virtual screening of natural products.  J Med Chem. 2004;  47 6248-54
  • 17 Wolber G, Langer T. LigandScout: 3 D pharmacophores derived from protein-bound ligands and their use as virtual screening filters.  J Chem Inf Model. 2005;  45 160-9
  • 18 Schmidtke M, Schnittler U, Jahn B, Dahse H M, Stelzner A. A rapid assay for evaluation of antiviral activity against Coxsackie virus B3, influenza virus A, and Herpes simplex virus type 1.  J Virolog Methods. 2001;  95 133-43
  • 19 Makarov V A, Riabova O B, Granik V G, Wutzler P, Schmidtke M. Novel [(biphenyloxy)propyl]isoxazole derivatives for inhibition of human rhinovirus 2 and Coxsackievirus B3 replication.  J Antimicrob Chemother. 2005;  55 483-8
  • 20 Boyd D R, Sharma N D, Barr S A, Carroll J G, Mackerracher D, Malone J F. Synthesis and absolute stereochemistry assignment of enantiopure dihydrofuro- and dihydropyrano-quinoline alkaloids. J Chem Soc [Perkin I] 2000: 3397-405
  • 21 Novak I, Buzas G, Minker E, Koltai M, Szendrei K. Active constituents of Ruta graveolens. .  Pharmazie. 1965;  20 738
  • 22 Novak I, Buzas G, Minker E, Koltai M, Szendrei K. Active principles of Ruta graveolens. .  Planta Med. 1965;  13 226-33
  • 23 Novak I, Buzas G, Minker E, Koltai M, Szendrei K. Isolation of some components from the herb of Ruta graveolens. .  Acta Pharm Hung. 1967;  37 131-42
  • 24 Reisch J, Novak I, Szendrei K, Minker E. Chemistry of natural substances. V. Isoimperatorin, a component of Ruta graveolens. .  Pharmazie. 1966;  21 628-9
  • 25 Reisch J, Novak I, Szendrei K, Minker E. Chemistry of natural substances. XIX. Additional C3-substituted coumarin derivatives from Ruta graveolens: daphnoretin and its methyl ether.  Planta Med. 1968;  16 372-6
  • 26 Reisch J, Adesina S K, Bergenthal D. Constituents of Zanthoxylum leprieurii fruit pericarps. Part 103: Natural product chemistry.  Pharmazie. 1985;  40 811-2
  • 27 Steck W, Bailey B K, Shyluk J P, Gamborg O. Coumarins and alkaloids from cell cultures of Ruta graveolens. .  Phytochemistry. 1971;  10 191-4
  • 28 Wolters B, Eilert U. Antimicrobial substances in callus cultures of Ruta graveolens. .  Planta Med. 1981;  43 166-74
  • 29 Del Castillo J B, Rodriguez L F, Secundino M. Angustifolin, a coumarin from Ruta angustifolia. .  Phytochemistry. 1984;  23 2095-6
  • 30 Puricelli L, Innocenti G, Delle Monache G, Caniato R, Filippini R, Cappelletti E M. In vivo and in vitro production of alkaloids by Haplophyllum patavinum. .  Nat Prod Lett. 2002;  16 95-100
  • 31 Gunawardana Y AGP, Cordell G A, Ruangrungsi N, Chomya S, Tantivatana P. Traditional medicinal plants of Thailand. VII. Alkaloids of Evodia lepta and Evodia gracilis. .  J Sci Soc Thailand. 1987;  13 107-12
  • 32 Chen J J, Chang Y L, Teng C M, Su C C, Chen I S. Quinoline alkaloids and anti-platelet aggregation constituents from the leaves of Melicope semecarpifolia.  Planta Med. 2002;  68 790-3
  • 33 Razakova D M, Bessonova I A, Yunusov S Y. Alkaloids from Haplophyllum perforatum. .  Khim Prir Soedin. 1976;  5 682
  • 34 Akhmedzhanova V I, Bessonova I A, Yunusov S Y. Alkaloids of Haplophyllum foliosum. .  Khim Prir Soed. 1980;  6 803-5
  • 35 Schöpf C, Wüst W. The absolute configuration of optically active β-hydroxy-β-phenylpropionic acids.  Ann. 1959;  626 150-4
  • 36 Chlouchi A, Girard C, Tillequin F, Bevalot F, Waterman P G, Muyard F. Coumarins and furoquinoline alkaloids from Philotheca deserti var. deserti (Rutaceae).  Biochem Syst Ecol. 2006;  34 71-4
  • 37 Chen C C, Huang Y L, Huang F I, Wang W, Ou J C. Water-soluble glycosides from Ruta graveolens. .  J Nat Prod. 2001;  64 990-2
  • 38 Spencer G F, Desjardins A E, Plattner R D. 5-(2-Carboxylethyl)-6-hydroxy-7-methoxybenzofuran, a fungal metabolite of xanthotoxin.  Phytochemistry. 1990;  29 2495-7
  • 39 Relkin N R. Beyond symptomatic therapy: a re-examination of acetylcholinesterase inhibitors in Alzheimer′s disease.  Exp Rev Neurother. 2007;  7 735-48
  • 40 Mukherjee P K, Kumar V, Mal M, Houghton P J. Acetylcholinesterase inhibitors from plants.  Phytomedicine. 2007;  14 289-300
  • 41 Patick A K. Rhinovirus chemotherapy.  Antivir Res. 2006;  71 391-6
  • 42 Iwamura H, Suzuki H, Kaya T, Inaba T. In vitro and in vivo pharmacological characterization of JTE-907, a novel selective ligand for cannabinoid CB2 receptor.  J Pharmacol Exp Ther. 2001;  296 420-5
  • 43 Ibrahim M M, Porreca F, Lai J, Albrecht P J, Rice F L, Khodorova A. CB2 cannabinoid receptor activation produces antinociception by stimulating peripheral release of endogenous opioids.  Proc Natl Acad Sci USA. 2005;  102 3093-8
  • 44 Steindl T M, Crump C E, Hayden F G, Langer T. Pharmacophore modeling, docking, and principal component analysis based clustering: Combined computer-assisted approaches to identify new inhibitors of the human rhinovirus coat protein.  J Med Chem. 2005;  48 6250-60
  • 45 Rollinger J M, Steindl T M, Schuster D, Kirchmair J, Anrain K, Ellmerer E P. Structure-based virtual screening for the discovery of natural inhibitors for human rhinovirus coat protein.  J Med Chem. 2008;  51 842-51
  • 46 Markt P. Discovery of novel CB2 receptor ligands by a pharmacophore-based virtual screening workflow. J Med Chem, in press
  • 47 Raduner S, Majewska A, Chen J Z, Xie X Q, Hamon J, Faller B. Alkylamides from Echinacea are a new class of cannabinomimetics: cannabinoid type 2 receptor-dependent and -independent immunomodulatory effects.  J Biol Chem. 2006;  281 14 192-206
  • 48 Gertsch J, Leonti M, Raduner S, Racz I, Chen J Z, Xie X Q. Beta-caryophyllene is a dietary cannabinoid.  Proc Natl Acad Sci USA. 2008;  105 9099-104

A. Univ.-Professor Mag. pharm. Dr. Judith Maria Rollinger

Institute of Pharmacy, Pharmacognosy and Center for Molecular Biosciences

University of Innsbruck

Innrain 52c

A-6020 Innsbruck

Austria

Phone: +43-512-507-5308

Fax: +43-512-507-2939

Email: judith.rollinger@uibk.ac.at