Pharmaceutical Nanotechnology
Formulation and cytotoxicity of doxorubicin nanoparticles carried by dry powder aerosol particles

https://doi.org/10.1016/j.ijpharm.2006.03.052Get rights and content

Abstract

Regional drug delivery via dry powder inhalers offers many advantages in the management of pharmaceutical compounds for the prevention and treatment of respiratory diseases. In the present study, doxorubicin (DOX)-loaded nanoparticles were incorporated as colloidal drug delivery system into inhalable carrier particles using a spray-freeze-drying technique. The cytotoxic effects of free DOX, carrier particles containing blank nanoparticles or DOX-loaded nanoparticles on H460 and A549 lung cancer cells were assessed using a colorimetric XTT cell viability assay. The mean geometric carrier particle size of 10 ± 4 μm was determined using confocal laser scanning microscopy. DOX-loaded nanoparticles had a particle size of 173 ± 43 nm after re-dissolving of the carrier particles. Compared to H460 cells, A549 cells showed less sensitivity to the treatment with free DOX. The DOX-nanoparticles showed in both cell lines a higher cytotoxicity at the highest tested concentration compared to the blank nanoparticles and the free DOX. The cell uptake of free DOX and DOX delivered by nanoparticles was confirmed using confocal laser scanning microscopy. This study supports the approach of lung cancer treatment using nanoparticles in dry powder aerosol form.

Introduction

Pulmonary inhalation systems have been used to deliver pharmaceutical compounds for the prevention and treatment of respiratory diseases (Sharma et al., 2001, Wang et al., 2003). For sufficient drug deposition the particle size of the aerosol has to be between 1 and 5 μm (Finlay, 2001). Nanoparticles are generally too small for pulmonary delivery due to their small mass median aerodynamic diameter (MMAD). The MMAD determines the deposition location of particles within the lungs. We recently developed inhalable carrier particles, which were loaded with empty nanoparticles (Sham et al., 2004). In this work it was demonstrated that the carrier particles dissolve after coming in contact with the aqueous medium and release the nanoparticles. Carrier particles can be made with an appropriate MMAD to optimize alveolar deposition. Such an approach can be used for the local delivery of drug-loaded nanoparticles to the lungs.

Lung cancer is the second most common cancer and the most common cause of cancer related death in both men and women (Jemal et al., 2004). In the present study the anticancer agent doxorubicin (DOX) was used as a model drug. Its incorporation into poly(butylcyanoacrylate) nanoparticles by an emulsion polymerization process is well established (Gulyaev et al., 1999). Recent investigations of the translocation of nanoparticles in the alveolar region indicate that surfactants coated polystyrene particles translocated across the alveolar capillary barrier while uncoated particles did not (Kato et al., 2003). Therefore, in the present study we used polysorbate 80 coated nanoparticles which were loaded with DOX.

The cytotoxic effects of free DOX, carrier particles containing blank nanoparticles or DOX-loaded nanoparticles on H460 and A549 lung cancer cell lines were assessed using a colorimetric XTT cell viability assay. The cell uptake of free DOX and DOX delivered by nanoparticles was confirmed using confocal laser scanning microscopy.

Section snippets

Materials

Dextran 70,000 and polysorbate 80 were supplied by Sigma (Ontario, Canada). N-Butylcyanoacrylate monomer (Lot. 02GD9236) was a gift from Loctite Ltd. (Dublin, Ireland). DOX was purchased from Pfizer Canada Inc. (Ontario, Canada). Lactose monohydrate was obtained from FMC (Philadelphia, USA). Human non-small cell lung cancer H460 and A549 cell lines were obtained from American Type Culture Collection (ATCC, Rockville, MD, USA). Cytotoxicity was analyzed using a cell proliferating XTT kit (Roche

Results

The drug loading study showed that approximately 85% of DOX was adsorbed on the poly(butylcyanoacrylate) nanoparticles which was in accordance with the other published data (Gulyaev et al., 1999, Gelperina et al., 2002). The loading amount of DOX in the carrier particles after spray-freeze drying was calculated as 1.39 μg DOX/mg powder. The mean particle sizes of DOX-loaded nanoparticles after re-dissolving of the spray-freeze-dried powders were 173 ± 43 nm. The spray-freeze dried powder particles

Discussion

In the present study DOX-loaded nanoparticles were incorporated as model drug delivery system into inhalable carrier particles. In order to avoid chemical decomposition and a loss of drug activity, spray freeze-drying technology (Maa et al., 1999) rather than a conventional heat spray-drying technology was used to obtain the carrier microparticles. The spray freeze-drying into liquid nitrogen and storage of the particles at 4 °C can prevent a possible Millard reaction or decrease the reaction

Acknowledgements

This study was supported by a NSERC Strategic grant, Alberta Cancer Board and the University of Alberta. The authors thank Helena Orszanska for her help in spray-freeze drying.

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