A Detailed Investigation on Conformation, Permeability and PK Properties of Two Related Cyclohexapeptides

  • Ian Lewis
  • Michael Schaefer
  • Trixie Wagner
  • Lukas Oberer
  • Emine Sager
  • Peter Wipfli
  • Thomas Vorherr


This contribution provides detailed insights regarding permeability properties of published cyclohexapeptides related to their conformation. The study includes data on oral bioavailability and PK parameters including application of a known enhancer. Interestingly, the latter did not show an effect, neither on the cyclic peptide exhibiting low oral uptake nor on the similar molecule showing a 30 % oral bioavailability in mice. These results are discussed in the context of exposed polar surface area for the preferred conformation and structural data obtained by NMR and X-ray crystallography.

Graphical Abstract


Permeability Pharmacokinetics Cyclohexapeptides Conformation 





Bioavailability on p.o. administration




In-vitro or in vivo clearance


Cyslosporine A




DIPEA diisopropyl ethylamine


Dimethyl formamide


Dodecylmaltoside enhancer


Lithium t-butoxide


Nuclear Overhauser effect spectroscopy


Parallel artificial membrane permeability assay


Madin Darby canine kidney cells


Plasma–protein binding


Root mean squared deviation


Per os


Hydrodynamic radius


NOE in the rotating frame


Reversed-phase high performance liquid chromatography


Solvent-accessible polar surface area


Supercritical fluid chromatography


Trifluoroacetic acid


Thin layer chromatography



The excellent support of Sandrine Desrayaud, Valerie Cordier, Elisabeth Braun, Laurent Hoffmann and Francis Risser is gratefully acknowledged. We thank Armin Widmer ( for the molecular modeling program Witnotp.

Conflict of interest

Ian Lewis, Michael Schaefer, Trixie Wagner, Lukas Oberer, Emine Sager, Peter Wipfli and Thomas Vorherr declare that they have no conflict of interest concerning Human and Animal Rights and Informed Consent.

Supplementary material

10989_2014_9447_MOESM1_ESM.pdf (662 kb)
Supplementary material 1 (PDF 661 kb)


  1. Alex A, Millan DS, Perez M, Wakenhut F, Whitlock GA (2011) Intramolecular hydrogen bonding to improve membrane permeability and absorption in beyond rule of five chemical space. Med Chem Commun 2:669–674CrossRefGoogle Scholar
  2. Bockus AT, McEwen CM, Lokey RS (2013) Form and function in cyclic peptide natural products: a pharmacokinetic perspective. Curr Top Med Chem 13:821–836CrossRefPubMedGoogle Scholar
  3. Brooks BR, Bruccoleri RE, Olafson BD, States DJ, Swaminathan S, Karplus M (1983) CHARMM: a program for macromolecular energy, minimization, and dynamics calculations. J Comput Chem 4:87–217CrossRefGoogle Scholar
  4. Calimet N, Schaefer M, Simonson T (2001) Protein molecular dynamics with the generalized born/ACESolvent model. Proteins: structure. Funct Genet 45:144–158CrossRefGoogle Scholar
  5. Chan WC, White PD (2000) Fmoc solid phase peptide synthesis, the practical approach series. In: Hames BD (ed), Oxford University Press, OxfordGoogle Scholar
  6. Chatterjee J, Gilon C, Hoffman A, Kessler H (2008) N-methylation of peptides: a new perspective in medicinal chemistry. Acc Chem Res 41(10):1331–1342CrossRefPubMedGoogle Scholar
  7. Clark DE (1999) Rapid calculation of polar molecular surface area and its application to the prediction of transport phenomena, 1. Prediction of intestinal absorption. J Pharm Sci 88(8):807–814CrossRefPubMedGoogle Scholar
  8. Connolly ML (1983) Solvent-accessible surfaces of proteins and nucleic acids. Science 221(4612):709–713CrossRefPubMedGoogle Scholar
  9. Daura X, Gademann K, Jaun B, Seebach D, van Gunsteren WF, Mark AE (1999) Peptide folding: when simulation meets experiment. Angew Chem Int Ed 38:236–240CrossRefGoogle Scholar
  10. Deshmukh DD, Nagilla R, Ravis WR, Betageri GV (2010) Effect of dodecylmaltoside (DDM) on uptake of BCS III compounds, tiludronate and cromolyn, in Caco-2 cells and rat intestine model. Drug Deliv 17(3):145–151CrossRefPubMedGoogle Scholar
  11. Di L, Artursson P, Avdeef A, Ecker GF, Faller B, Fischer H, Houston JB, Kansy M, Kerns EH, Kramer SD, Lennernäs H, Kiyohiko S (2012) Evidence-based approach to assess passive diffusion and carrier-mediated drug transport. Drug Discov Today 17(15–16):905–912CrossRefPubMedGoogle Scholar
  12. El Tayar N, Mark AE, Vallat P, Brunne RM, Testa B, van Gunsteren WF (1993) Solvent-dependent conformation and hydrogen bonding capacity of cyclosporin A: evidence from partition coefficients and molecular dynamics simulations. J Med Chem 36:3757–3764CrossRefPubMedGoogle Scholar
  13. Giordanetto F, Kihlberg J (2014) Macrocyclic drugs and clinical candidates: what can medicinal chemists learn from their properties? J Med Chem 57:278–295CrossRefPubMedGoogle Scholar
  14. Goetz GH, Farrell W, Shalaeva M, Sciabola S, Anderson D, Yan J, Philippe L, Shapiro MJ (2014a) High throughput method for the indirect detection of intramolecular hydrogen bonding. J Med Chem 57(7):2920–2929CrossRefPubMedGoogle Scholar
  15. Goetz GH, Philippe L, Shapiro MJ (2014b) EPSA: A novel supercritical fluid chromatography technique enabling the design of permeable cyclic peptides. ACS Med Chem Lett. doi:  10.1021/ml500239m
  16. Golor G, Hu K, Ruffin M, Buchelt A, Bouillaud E, Wang Y, Maldonado M (2012) A first-in-man study to evaluate the safety, tolerability, and pharmacokinetics of pasireotide (SOM230), a multireceptor-targeted somatostatin analog, in healthy volunteers. Drug Des, Dev Therapy 6:71–79CrossRefGoogle Scholar
  17. Hill TA, Lohman R, Hoang HN, Nielsen DS, Scully CCG, Kok WM, Liu L, Lucke AJ, Stoermer MJ, Schroeder CI, Chaousis S, Colless B, Bernhardt PV, Edmonds DJ, Griffith DA, Rotter CJ, Ruggeri RB, Price DA, Liras S, Craik DJ, Fairlie DP (2014) Cyclic penta- and hexaleucine peptides without N-methylation are orally absorbed. ACS Med Chem Lett. doi: 10.1021/ml5002823
  18. Hoveyda HR, Marsault E, Gagnon R, Mathieu AP, Vézina M, Landry A, Wang Z, Benakli K, Beaubien S, Saint-Louis C, Brassard M, Pinault J, Ouellet L, Bhat S, Ramaseshan M, Peng X, Foucher L, Beauchemin S, Bhérer P, Veber DF, Peterson ML, Fraser GL (2011) Optimization of the potency and pharmacokinetic properties of a macrocyclic ghrelin receptor agonist (Part I): development of Ulimorelin (TZP-101) from hit to clinic. J Med Chem 54:8305–8320CrossRefPubMedGoogle Scholar
  19. Hutchinson EG, Thornton JM (1994) A revised set of potentials for beta-turn formation in proteins. Protein Sci 3(12):2207–2216CrossRefPubMedCentralPubMedGoogle Scholar
  20. Kabsch W, Sander C (1983) Dictionary of protein secondary structure: pattern recognition of hydrogen-bonded and geometrical features. Biopolymers 22(12):2577–2637CrossRefPubMedGoogle Scholar
  21. Karplus M (1959) Contact electron-spin coupling of nuclear magnetic moments. J Chem Phys 30(1):11–15CrossRefGoogle Scholar
  22. Kenichiro I, Passioura T, Suga H (2013) Technologies for the synthesis of mRNA-encoding libraries and discovery of bioactive natural product-inspired non-traditional macrocyclic peptides. Molecules 18:3502–3528CrossRefGoogle Scholar
  23. Knipp GT, Vander Velde GD, Siahaan TJ, Borchardt RT (1997) The effect of β-turn structure on the passive diffusion of peptides across Caco-2 cell monolayers. Pharm Res 14:1332–1340CrossRefPubMedGoogle Scholar
  24. Liu X, Wright M, Hop CECA (2014) Rational use of plasma protein and tissue binding data in drug design. Miniperspective. J Med Chem 57(20):8238–8248Google Scholar
  25. Ma P, Wang Y, Van Der Hoek J, Nedelman J, Schran H, Tran LL, Lamberts SW (2005) Pharmacokinetic-pharmacodynamic comparison of a novel multi ligand somatostatin analog, SOM230, with octreotide in patients with acromegaly. Clin Pharmacol Ther 78(1):69–80CrossRefPubMedGoogle Scholar
  26. Marzinzik AL, Vorherr Th (2013) Towards intracellular delivery of peptides. Chimia 67(12):899–905CrossRefPubMedGoogle Scholar
  27. Miranda E, Nordgren IK, Male AL, Lawrence CE, Hoakwie F, Cuda F, Court W, Fox KR, Townsend PA, Packham GK, Eccles SA, Tavassoli A (2013) A cyclic peptide inhibitor of HIF-1 heterodimerization that inhibits hypoxia signaling in cancer cells. J Am Chem Soc 135(28):10418–10425CrossRefPubMedCentralPubMedGoogle Scholar
  28. Mönkärea J, Hakalab RA, Kovalainena M, Korhonenb H, Herzigc K, Seppäläb JV, Järvinena K (2012) Photocrosslinked poly(ester anhydride)s for peptide delivery: effect of oligomer hydrophobicity on PYY3-36 delivery. Eur J Pharm Biopharm 80(1):33–38CrossRefGoogle Scholar
  29. Nielsen DS, Hoang HN, Lohman R, Diness F, Fairlie DP (2012) Total synthesis, structure, and oral absorption of a thiazole cyclic peptide, sanguinamide A. Org Lett 14(22):5720–5723CrossRefPubMedGoogle Scholar
  30. Okumu FW, Pauletti GM, Vander Velde GD, Siahaan TJ, Borchardt RT (1997) Effect of restricted conformational flexibility on the permeation of model hexapeptides across Caco-2 cell monolayers. Pharm Res 14:169–175CrossRefPubMedGoogle Scholar
  31. Over B, McCarren P, Artursson P, Foley M, Giordanetto F, Grönberg G, Hilgendorf C, Lee MD, Matsson P, Muncipinto G, Pellisson M, Perry MWD, Svensson R, Duvall JR, Kihlberg J (2014) Impact of stereospecific intramolecular hydrogen bonding on cell permeability and physicochemical properties. J Med Chem 57:2746–2754CrossRefPubMedCentralPubMedGoogle Scholar
  32. Poillot C, Bichraoui H, Tisseyre C, Bahemberae E, Andreotti N, Sabatier J, Ronjat M, De Waard M (2012) Small efficient cell-penetrating peptides derived from scorpion toxin maurocalcine. J Biol Chem 287(21):17331–17342CrossRefPubMedCentralPubMedGoogle Scholar
  33. Rafi SB, Hearn BR, Vedantham P, Jacobson MP, Renslo AR (2012) Predicting and improving the membrane permeability of peptidic small molecules. J Med Chem 55(7):3163–3169CrossRefPubMedCentralPubMedGoogle Scholar
  34. Ramachandran GN, Ramakrishnan C, Sasisekharan V (1963) Stereochemistry of polypeptide chain configurations. J Mol Biol 7(1):95–99CrossRefPubMedGoogle Scholar
  35. Rand AC, Leung SSF, Eng H, Rotter CJ, Sharma R, Kalgutkar AS, Zhang Y, Varma MV, Farley KA, Khunte B, Limberakis C, Price DA, Liras S, Mathiowetz AM, Jacobson MP, Lokey RS (2012) Optimizing PK properties of cyclic peptides: the effect of side chain substitutions on permeability and clearance. Med Chem Comm 3(10):1282–1289Google Scholar
  36. Rezai T, Bock JE, Zhou MV, Kalyanaraman C, Lokey RS, Jacobson MP (2006) Conformational flexibility, internal hydrogen bonding, and passive membrane permeability: successful in silico prediction of the relative permeabilities of cyclic peptides. J Am Chem Soc 128:14073–14080CrossRefPubMedGoogle Scholar
  37. Rezai T, Bock JE, Zhou MV, Kalyanaraman C, Lokey RS, Jacobson MP (2011) On-resin N-methylation of cyclic peptides for discovery of orally bioavailable scaffolds conformational flexibility, internal hydrogen bonding, and passive membrane permeability: successful in silico prediction of the relative permeabilities of cyclic peptides. Nat Chem Biol 7:810CrossRefPubMedCentralPubMedGoogle Scholar
  38. Richmond TJ (1984) Solvent accessible surface area and excluded volume in proteins. Analytical equations for overlapping spheres and implications for the hydrophobic effect. J Mol Biol 178(1):63–89CrossRefPubMedGoogle Scholar
  39. Schaefer M, Karplus M (1996) A comprehensive analytical treatment of continuum electrostatics. J Phys Chem B 100:1578–1599CrossRefGoogle Scholar
  40. Schaefer M, Bartels C, Karplus M (1998) Solution conformations and thermodynamics of structured peptides: molecular dynamics simulation with an implicit solvation model. J Mol Biol 284(3):835–848CrossRefPubMedGoogle Scholar
  41. Seidler J, McGovern SL, Doman TN, Shoichet BK (2003) Identification and prediction of promiscuous aggregating inhibitors among known drugs. J Med Chem 46:4477–4486CrossRefPubMedGoogle Scholar
  42. Smith D, Artursson P, Avdeef A, Di L, Ecker GF, Faller B, Houston JB, Kansy M, Kerns EH, Kramer SD, Lennernäs H, van de Waterbeemd H, Sugano K, Testa B (2014) Passive lipoidal diffusion and carrier-mediated cell uptake are both important mechanisms of membrane permeation in drug disposition. Mol Pharm 11(6):1727–1738CrossRefPubMedGoogle Scholar

Copyright information

© Springer Science+Business Media New York 2014

Authors and Affiliations

  • Ian Lewis
    • 1
  • Michael Schaefer
    • 1
  • Trixie Wagner
    • 1
  • Lukas Oberer
    • 1
  • Emine Sager
    • 1
  • Peter Wipfli
    • 1
  • Thomas Vorherr
    • 1
  1. 1.Novartis Institutes for Biomedical ResearchBaselSwitzerland

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