Skip to main content
SpringerLink
Log in

Iloprost-Containing Liposomes for Aerosol Application in Pulmonary Arterial Hypertension: Formulation Aspects and Stability

  • Research Paper
  • Published:
Pharmaceutical Research Aims and scope Submit manuscript

Abstract

Purpose

Pulmonary arterial hypertension (PAH) is a severe and progressive disease. The prostacyclin analogue iloprost is effective against PAH, but requires six to nine inhalations per day. The feasibility of liposomes to provide a sustained release formulation to reduce inhalation frequency is evaluated from a technological point of view.

Methods

Liposomal formulations consisting of di-palmitoyl-phosphatidyl-choline (DPPC), cholesterol (CH) and polyethyleneglycol-di-palmitoyl-phosphatidyl-ethanolamine (DPPE-PEG) were prepared. Their physico-chemical properties were investigated using dynamic light scattering, atomic force microscopy and differential scanning calorimetry. Stability of liposomes during aerosolization using three different nebulizers (air-jet, ultrasonic and vibrating mesh) was investigated with respect to drug loading and liposome size, pre- and post-nebulization.

Results

The phospholipid composition affected the diameters of liposomes only slightly in the range of 200–400 nm. The highest iloprost loading (12 μg/ml) and sufficient liposome stability (70% drug encapsulation post-nebulization) was observed for the DPPC/CH (70:30 molar ratio) liposomes. The formulation’s stability was confirmed by the relatively high phase transition temperature (53°C) and unchanged particle sizes. The incorporation of DPPE-PEG in the liposomes (DPPC/CH/DPPE-PEG, 50:45:5 molar ratio) resulted in decreased stability (20–50% drug encapsulation post-nebulization) and a phase transition temperature of 35°C. The vibrating mesh nebulizer offered a number of significant advantages over the other nebulizers, including the production of small aerosol droplets, high output, and the lowest deleterious physical influence upon all investigated liposomes.

Conclusion

Iloprost-loaded liposomes containing DPPC and CH components yield formulations which are well suited to aerosolization by the vibrating mesh nebulizer. The investigation of sustained release effects for the treatment of PAH in ex vivo and in vivo models is under way.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5
Fig. 6

Similar content being viewed by others

Abbreviations

AFM:

atomic force microscopy

CH:

cholesterol

COF:

5(6)-carboxyfluorescein

DE :

encapsulated drug in the liposomal dispersion

DNE :

non-encapsulated drug in the liposomal dispersion

Dstart :

total drug concentration utilized for drug loading during liposome preparation

DT :

total drug concentration in the liposomal dispersion

DLS:

dynamic light scattering

DPPE-PEG:

polyethyleneglycol-dipalmitoyl-phosphatidylethanolamine

DPPC:

dipalmitoyl-phosphatidylcholine

DSC:

differential scanning calorimeter

EE:

encapsulation efficiency

GSD:

geometric standard deviation

ILO:

iloprost

LDA:

laser Doppler anemometry

MMD:

mass median diameter

PAH:

pulmonary arterial hypertension

PDI:

polydispersity index

PEG:

poly(ethylene glycol)

SD:

standard deviation

Tc:

phase transition temperature

References

  1. H. Olschewski and W. Seeger. Pulmonary Hypertension: Pathophysiology, Diagnosis, Treatment, and Development of a Pulmonary-Selective Therapy, UNI-MED Verlag AG, Bremen, 2002.

    Google Scholar 

  2. S. Archer and S. Rich. Primary pulmonary hypertension: a vascular biology and translational research “work in progress”. Circulation 102:2781–2791 (2000).

    PubMed  CAS  Google Scholar 

  3. D. B. Badesch, S. H. Abman, G. S. Ahearn, R. J. Barst, D. C. McCrory, G. Simonneau, and V. V. McLaughlin. Medical therapy for pulmonary arterial hypertension—ACCP evidence-based clinical practice guidelines. Chest 126:35S–62S (2004).

    Article  PubMed  Google Scholar 

  4. R. T. Schermuly, C. Inholte, H. A. Ghofrani, H. Gall, N. Weissmann, A. Weidenbach, W. Seeger, and F. Grimminger. Lung vasodilatory response to inhaled iloprost in experimental pulmonary hypertension: amplification by different type phosphodiesterase inhibitors. Respir. Res. 6:76 (2005).

    Article  PubMed  Google Scholar 

  5. V. V. McLaughlin, A. Shillington, and S. Rich. Survival in primary pulmonary hypertension—the impact of epoprostenol therapy. Circulation 106:1477–1482 (2002).

    Article  PubMed  CAS  Google Scholar 

  6. S. Rich and V. V. McLaughlin. The effects of chronic prostacyclin therapy on cardiac output and symptoms in primary pulmonary hypertension. J. Am. Coll. Cardiol. 34:1184–1187 (1999).

    Article  PubMed  CAS  Google Scholar 

  7. H. A. Ghofrani, F. Rose, R. T. Schermuly, H. Olschewski, R. Wiedemann, N. Weissmann, C. Schudt, H. Tenor, W. Seeger and F. Grimminger. Amplification of the pulmonary vasodilatory response to inhaled iloprost by subthreshold phosphodiesterase types 3 and 4 inhibition in severe pulmonary hypertension. Crit. Car. Med. 30:2489–2492 (2002).

    Article  CAS  Google Scholar 

  8. R. T. Schermuly, H. Yilmaz, H. A. Ghofrani, K. Woyda, S. Pullamsetti, A. Schulz, T. Gessler, R. Dumitrascu, N. Weissmann, F. Grimminger, and W. Seeger. Inhaled iloprost reverses vascular remodeling in chronic experimental pulmonary hypertension. Am. J. Respir. Crit. Care Med. 172:358–363 (2005).

    Article  PubMed  Google Scholar 

  9. H. Olschewski, H. A. Ghofrani, T. Schmehl, J. Winkler, H. Wilkens, M. M. Höper, J. Behr, F. X. Kleber, and W. Seeger. Inhaled iloprost to treat severe pulmonary hypertension. Ann. Intern. Med. 132:435–443 (2000).

    PubMed  CAS  Google Scholar 

  10. S. Conti, L. Polonelli, R. Frazzi, M. Artusi, R. Bettini, D. Cocconi, and P. Colombo. Controlled delivery of biotechnological products. Curr. Pharm. Biotechnol. 1:313–323 (2000).

    Article  PubMed  CAS  Google Scholar 

  11. A. I. Bot, T. E. Tarara, D. J. Smith, S. R. Bot, C. M. Woods, and J. G. Weers. Novel lipid-based hollow-porous microparticles as a platform for immunoglobulin delivery to the respiratory tract. Pharm. Res. 17:275–283 (2000).

    Article  PubMed  CAS  Google Scholar 

  12. K. M. Taylor and J. M. Newton. Liposomes for controlled delivery of drugs to the lung. Thorax 47:257–259 (1992).

    Article  PubMed  CAS  Google Scholar 

  13. J. Fiegel, J. Fu, and J. Hanes. Poly(ether-anhydride) dry powder aerosols for sustained drug delivery in the lungs. J. Control. Release 96:411–423 (2004).

    Article  PubMed  CAS  Google Scholar 

  14. R. L. Juliano and H. N. McCullough. Controlled delivery of an antitumor drug: localized action of liposome encapsulated cytosine arabinoside administered via the respiratory system. J. Pharmacol. Exp. Ther. 214:381–387 (1980).

    PubMed  CAS  Google Scholar 

  15. D. A. Thomas, M. A. Myers, B. Wichert, H. Schreier, and R. J. Gonzalez-Rothi. Acute effects of liposome aerosol inhalation on pulmonary function in healthy human volunteers. Chest 99:1268–1270 (1991).

    Article  PubMed  CAS  Google Scholar 

  16. Y. Darwis and I. W. Kellaway. Nebulisation of rehydrated freeze-dried beclomethasone dipropionate liposomes. Int. J. Pharm. 215:113–121 (2001).

    Article  PubMed  CAS  Google Scholar 

  17. M. Saari, M. T. Vidgren, M. O. Koskinen, V. M. H. Turjanmaa, and M. M. Nieminen. Pulmonary distribution and clearance of two beclomethasone liposome formulations in healthy volunteers. Int. J. Pharm. 181:1–9 (1999).

    Article  PubMed  CAS  Google Scholar 

  18. T. R. Desai, R. E. Hancock, and W. H. Finlay. A facile method of delivery of liposomes by nebulization. J. Control. Release 84:69–78 (2002).

    Article  PubMed  CAS  Google Scholar 

  19. R. J. Gonzalez-Rothi, S. Suarez, G. Hochhaus, H. Schreier, A. Lukyanov, H. Derendorf, and T. D. Costa. Pulmonary targeting of liposomal triamcinolone acetonide phosphate. Pharm. Res. 13:1699–1703 (1996).

    Article  PubMed  CAS  Google Scholar 

  20. S. Suarez, R. Gonzalez-Rothi, H. Schreier, and G. Hochhaus. Effect of dose and release rate on pulmonary targeting of liposomal triamcinolone acetonide phosphate. Pharm. Res. 15:461–465 (1998).

    Article  PubMed  CAS  Google Scholar 

  21. C. Khanna, J. C. Waldrep, P. M. Anderson, R. W. Weischelbaum, D. E. Hasz, E. Katsanis, and J. S. Klausner. Nebulized interleukin 2 liposomes: aerosol characteristics and biodistribution. J. Pharm. Pharmacol. 49:960–971 (1997).

    Article  PubMed  CAS  Google Scholar 

  22. R. M. Ten, P. M. Anderson, N. N. Zein, Z. Temesgen, M. L. Clawson, and W. Weiss. Interleukin-2 liposomes for primary immune deficiency using the aerosol route. Int. Immunopharm. 2:333–334 (2001).

    Article  Google Scholar 

  23. T. L. Yaksh, J. C. Provencher, M. L. Rathbun, R. R. Myers, H. Powell, P. Richter, and F. R. Kohn. Safety assessment of encapsulated morphine delivered epidurally in a sustained-release multivesicular liposome preparation in dogs. Drug Deliv. 7:27–36 (2000).

    Article  PubMed  CAS  Google Scholar 

  24. O. R. Hung, E. M. Sellers, H. L. Kaplan, and M. K. Romach. Phase IB clinical trial of aerosolized liposome encapsulated fentanyl (AeroLEF (TM). Clin. Pharmacol. Ther. 75:P4 (2004).

    Article  Google Scholar 

  25. J. L. Rau. Design principles of liquid nebulization devices currently in use. Respir. Care 47:1257–1275 (2002).

    PubMed  Google Scholar 

  26. O. N. M. McCallion, K. M. G. Taylor, M. Thomas, and A. J. Taylor. Nebulization of fluids of different physicochemical properties with air-jet and ultrasonic nebulizers. Pharm. Res. 12:1682–1688 (1995).

    Article  PubMed  CAS  Google Scholar 

  27. J. B. Fink, D. Schmidt and J. Power. Comparison of a nebulizer using a novel aerosol generator with a standard ultrasonic nebulizer designed for use during mechanical ventilation. In I. Aerogen (ed.), ATS 2001, 2001.

  28. P. A. Bridges and K. M. G. Taylor. An investigation of some of the factors influencing the jet nebulisation of liposomes. Int. J. Pharm. 204:69–79 (2000).

    Article  PubMed  CAS  Google Scholar 

  29. L. A. Dailey, T. Schmehl, T. Gessler, M. Wittmar, F. Grimminger, W. Seeger, and T. Kissel. Nebulization of biodegradable nanoparticles: impact of nebulizer technology and nanoparticle characteristics on aerosol features. J. Control. Release 86:131–144 (2003).

    Article  PubMed  CAS  Google Scholar 

  30. R. Abu-Dahab, U. F. Schäfer, and C. M. Lehr. Lectin-functionalized liposomes for pulmonary drug delivery: effect of nebulization on stability and bioadhesion. Eur. J. Pharm. Sci. 14:37–46 (2001).

    Article  PubMed  CAS  Google Scholar 

  31. K. K. M. Leung, P. A. Bridges, and K. M. G. Taylor. The stability of liposomes to ultrasonic nebulisation. Int. J. Pharm. 145:95–102 (1996).

    Article  CAS  Google Scholar 

  32. K. M. G. Taylor and O. N. M. McCallion. Ultrasonic nebulisers for pulmonary drug delivery. Int. J. Pharm. 153:93–104 (1997).

    Article  CAS  Google Scholar 

  33. E. Kleemann, L. A. Dailey, H. G. Abdelhady, T. Gessler, T. Schmehl, C. J. Roberts, M. C. Davies, W. Seeger, and T. Kissel. Modified polyethylenimines as non-viral gene delivery systems for aerosol gene therapy: investigations of the complex structure and stability during air-jet and ultrasonic nebulization. J. Control. Release 100:437–450 (2004).

    Article  PubMed  CAS  Google Scholar 

  34. Y. Barenholz. Liposome application: problems and prospects. Curr. Opin. Colloid Interface Sci. 6:66–77 (2001).

    Article  CAS  Google Scholar 

  35. R. Dhand. New frontiers in aerosol delivery during mechanical ventilation. Respir. Care 49:666–677 (2004).

    PubMed  Google Scholar 

  36. A. M. A. Elhissi and K. M. G. Taylor. Delivery of liposomes generated from proliposomes using air-jet, ultrasonic, and vibrating-mesh nebulisers. J. Drug Del. Sci. Tech. 15:261–265 (2005).

    CAS  Google Scholar 

  37. P. A. Bridges and K. M. G. Taylor. Nebulisers for the generation of liposomal aerosols. Int. J. Pharm. 173:117–125 (1998).

    Article  CAS  Google Scholar 

  38. B. V. Wichert, R. J. Gonzalez-Rothi, L. E. Straub, B. M. Wichert, and H. Schreier. Amikacin liposomes: characterization, aerosolization, and in vitro activity against Mycobacterium avium-intracellulare in alveolar macrophages. Int. J. Pharm. 78:227–235 (1992).

    Article  CAS  Google Scholar 

  39. M. Anderson and A. Omri. The effect of different lipid components on the in vitro stability and release kinetics of liposome formulations. Drug Deliv. 11:33–39 (2004).

    Article  PubMed  CAS  Google Scholar 

  40. T. Schmehl, T. Gessler, and E. Waschkowitz. WO 2003/084507A3 Atomizable liposomes and the use for the pulmonary administration of active substances, 2003.

  41. V. P. Torchilin and V. Weissig. Liposomes—A Practical Approach, Oxford University Press, Oxford, 2003.

    Google Scholar 

  42. R. C. MacDonald, R. I. MacDonald, B. P. M. Menco, K. Takeshita, N. K. Subbarao, and L. Hu. Small-volume extrusion apparatus for preparation of large, unilamellar vesicles. Biochim. Biophys. Acta 1061:297–303 (1991).

    Article  PubMed  CAS  Google Scholar 

  43. K. M. G. Taylor and R. M. Morris. Thermal-analysis of phase-transition behaviour in liposomes. Thermochim. Acta 248:289–301 (1995).

    Article  CAS  Google Scholar 

  44. H. Olschewski, B. Rohde, J. Behr, R. Ewert, T. Gessler, H. A. Ghofrani, and T. Schmehl. Pharmacodynamics and pharmacokinetics of inhaled iloprost, aerosolized by three different devices, in severe pulmonary hypertension. Chest 124:1294–1304 (2003).

    Article  PubMed  CAS  Google Scholar 

  45. D. A. Groneberg, C. Witt, U. Wagner, K. F. Chung, and A. Fischer. Fundamentals of pulmonary drug delivery. Respir. Med. 97:382–387 (2003).

    Article  PubMed  CAS  Google Scholar 

  46. P. W. Barry and C. O’Callaghan. Inhalational drug delivery from seven different spacer devices. Thorax 51: 835–840 (1996).

    Article  PubMed  CAS  Google Scholar 

  47. O. G. Mouritsen and K. Jorgensen. A new look at lipid-membrane structure in relation to drug research. Pharm. Res. 15:1507–1519 (1998).

    Article  PubMed  CAS  Google Scholar 

  48. D. D. Lasic and D. Needham. The stealth liposomes: a prototypical biomaterial. Chem. Rev. 95:2601–2627 (1995).

    Article  CAS  Google Scholar 

  49. D. D. Lasic. Novel applications of liposomes. Trends Biotechnol. 16: 307–321 (1998).

    Article  PubMed  CAS  Google Scholar 

  50. D. Crommelin and H. Schreier. Liposomes in Colloidal Drug Delivery Systems, Marcel Dekker, New York, 1994.

    Google Scholar 

  51. S. C. Semple, A. Chonn, and P. R. Cullis. Influence of cholesterol on the association of plasma proteins with liposomes. Biochemistry 35:2521–2525 (1996).

    Article  PubMed  CAS  Google Scholar 

  52. M. Grit and D. J. Crommelin. Chemical stability of liposomes: implications for their physical stability. Chem. Phys. Lipids 64:3–18 (1993).

    Article  PubMed  CAS  Google Scholar 

  53. M. Brandl, M. Drechsler, D. Bachmann, C. Tardi, M. Schmidtgen, and K. H. Bauer. Preparation and characterization of semi-solid phospholipid dispersions and dilutions thereof. Int. J. Pharm. 170:187–199 (1998).

    Article  CAS  Google Scholar 

  54. A. Schering. Sicherheitsdatenblatt Iloprost, Berlin, 1997, pp. 1–5.

  55. K. S. Konduri, S. Nandedkar, N. Düzgünes, V. Suzara, J. Artwohl, R. Bunte, and P. R. J. Gangadharam. Efficacy of liposomal budesonide on experimental asthma. J. Allergy Clin. Immunol. 111:321–327 (2003).

    Article  PubMed  CAS  Google Scholar 

  56. K. S. Konduri, S. Nandedkar, D. A. Rickaby, N. Düzgünes, and R. J. Gangadharam. The use of sterically stabilized liposomes to treat asthma. Methods Enzymol. 391:413–427 (2005).

    Article  PubMed  CAS  Google Scholar 

  57. R. N. Niven and H. Schreier. Nebulization of liposomes. I. Effects of lipid composition. Pharm. Res. 7:1127–1133 (1990).

    Article  PubMed  CAS  Google Scholar 

  58. M. A. Myers, D. A. Thomas, L. Straub, D. W. Soucy, R. W. Niven, M. Kaltenbach, C. I. Hood, H. Schreier, and R. J. Gonzalez-Rothi. Pulmonary effects of chronic exposure to liposome aerosols in mice. Exp. Lung Res. 19:1–19 (1993).

    Article  PubMed  CAS  Google Scholar 

  59. J. C. Waldrep, B. E. Gilbert, C. M. Knight, M. B. Black, P. W. Scherer, V. Knight, and W. Eschenbacher. Pulmonary delivery of beclomethasone liposome aerosol in volunteers. Tolerance and safety. Chest 111:316–323 (1997).

    Article  PubMed  CAS  Google Scholar 

  60. Y. Morimoto and Y. Adachi. Pulmonary uptake of liposomal phosphatidylcholine upon intratracheal administration to rats. Chem. Pharm. Bull. (Tokyo) 30:2248–2251 (1982).

    Article  CAS  Google Scholar 

  61. A. Pettenazzo, A. Jobe, M. Ikegami, R. Abra, E. Hogue, and P. Mihalko. Clearance of phosphatidylcholine and cholesterol from liposomes, liposomes loaded with metaproterenol, and rabbit surfactant from adult rabbit lungs. Am. Rev. Respir. Dis. 139:752–758 (1989).

    PubMed  CAS  Google Scholar 

  62. R. Dhand. Nebulizers that use a vibrating mesh or plate with multiple apertures to generate aerosol. Respir. Care 47: 1406–1416 (2002).

    PubMed  Google Scholar 

Download references

Acknowledgment

We wish to thank Karin Quanz, Eva Mohr, Nicole Bamberger and Klaus Keim for their excellent technical assistance.

Author information

Authors and Affiliations

  1. Department of Pharmaceutics and Biopharmacy, Philipps-University, D-35037, Marburg, Germany

    Elke Kleemann, Udo Bakowsky & Thomas Kissel

  2. University of Giessen Lung Center (UGLC), D-35392, Giessen, Germany

    Thomas Schmehl, Tobias Gessler & Werner Seeger

Authors
  1. Elke Kleemann
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Thomas Schmehl
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Tobias Gessler
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Udo Bakowsky
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Thomas Kissel
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Werner Seeger
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Elke Kleemann.

Rights and permissions

Reprints and permissions

About this article

Cite this article

Kleemann, E., Schmehl, T., Gessler, T. et al. Iloprost-Containing Liposomes for Aerosol Application in Pulmonary Arterial Hypertension: Formulation Aspects and Stability. Pharm Res 24, 277–287 (2007). https://doi.org/10.1007/PL00022055

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/PL00022055

Key words

Search

Navigation