(700g) Lipid-Based Particle Engineering Via Spray-Drying for Drug Targeting to the Lung

Rifampicin (RIF) is an antibiotic listed as an essential medicine by the World Health Organization. It is administered orally or intravenously in the therapy against Tuberculosis. However, RIF, like other antibiotics, can cause serious adverse effects, such as hepatotoxicity. Delivery of RIF directly to the site of action via inhalation can reduce the risk of such effects by diminishing systemic exposure. Furthermore, pulmonary delivery of RIF can lead to dose reduction and less frequent administration [1]. For that, RIF-loaded particles that are able to reach deep lung deposition and trigger phagocytosis of alveolar macrophages (AM) are required. These requirements can be provided by loading RIF into lipid-based excipients (LBE) followed by particle engineering via spray-drying. Targeted particle attributes are given by the interplay of lipid’s material attributes and process parameters. In this work, selected LBE were used to engineer particles for pulmonary delivery of RIF to be administered as dry powder for inhalation (DPI).

Materials

Diglycerol full ester of behenic acid (PG2-C22f) commercially available as Witepsol® PMF222 was selected as LBE due to its superior processability via spray-drying. As previously shown by our group in [2], the high melting point (Tm=72.5°C), monophasic crystallization and absence of polymorphism of PG2-C22f lead to high process yield. Glyceryl stearate citrate (Imwitor®372P) was used as lipid-based emulsifier, selected due to the similar solid state structure to PG2-C22f and high HLB (10-12).

Methods

Setting critical particle attributes: Inhalable particles with mass median aerodynamic diameter (MMAD) between 1-5µm were required for suitable lung deposition. After deposition, activation and uptake by AM were envisaged. Therefore, particles of volume mean diameter (VMD) below 4µm (optimal 2.1 µm) and negative surface charge, measured via Zeta Potential (ZP) [3] were designed.

Spray-drying: An aqueous-based suspension was prepared for spray-drying. PG2-C22f and Imwitor372P (1:1 mass ratio) were melted at 80°C. RIF was dissolved in the molten mixture (2.5% over lipid mass). Pre-heated water was added to the mixture to provide a solid content of 0.5% w/w. High-shear stirring was applied. The mixture was left to cool down at room temperature to allow the formation of solid lipid-particles loaded with RIF within a suspension. The suspension was atomized through a bi-fluid nozzle of 0.4mm (1.14 bar) at 2 g/min into the spray-drying chamber at inlet temperature (Tin) of 75°C. Particles were recovered via cyclone.

Particle characterization: The morphology of particles was observed via Scanning Electron Microscopy (SEM). Particle size distribution was analyzed via light diffraction. Bulk and tapped densities were used to assess Hausner ratio. Solid state and stability were evaluated via differential scanning calorimetry (DSC) and small and wide angle X-ray scattering (SWAXS) on freshly prepared samples and after storage for 3 months at 25°C/60%r.H (LTC) and 40°C/75%r.H (AC). The aerodynamic performance was assessed via the next generation impactor (NGI) coupled to Aerolizer dry powder inhaler. Six hypromellose capsules were loaded with 30mg of powder. An inhalation flow of 60 l/min for 4 seconds was used. MMAD and in vitro deposition were obtained. Particle uptake was assessed in THP-1 cells by exposing them to suspended particles for 24 hours, followed by drug extraction and quantification via HPLC.

Results

Physical characterization

Spray-drying of RIF-loaded lipid-suspension resulted in high yield of particles (83%). This is in agreement with our previous findings [2]. SEM images evidenced spherical particles of slightly rough surface. No internal voids were formed during spray-drying due to slow evaporative conditions (Tin<Tb,water). A suitable VMD of 2.41 ± 0.07µm was provided. The flowability of particles was characterized by a Hausner ratio of 1.90, associated to high powder cohesiveness. Negative surface charge (ZP = -36.25 ± 7.97 mV) was found in the particles, which is attributed to the carboxyl groups within the structure of PG2-C22f and glyceryl stearate citrate.

Solid state

The solid state of particles and stability thereof was confirmed via DSC and SWAXS. No alterations of endothermic events were observed. Endothermic transitions of RIF were not found, depicting the solubilization of the drug within the lipid matrix. SWAXS analysis confirmed the absence of crystalline RIF signals (Fig. 3). The crystalline structure of the lipid matrix corresponds to a stable α-form of both excipients, arranged in a mixed lamellar structure of 68A. After storage, however, slight inclination of lamellae due to potential appearance of the β-form of Glyceryl stearate citrate was observed.

Aerodynamic and cellular performance

The aerodynamic performance of the particles as lipid-based DPI showed that, in spite of the high cohesivity of the particles, they were successfully aerosolized. A high emitted fraction of 94.8 ± 0.2% was found and associated to the lipidic particle surface. Suitable MMAD of 2.44 ± 0.34µm depicted the readily inhalability of the particles with high FPF of 61.2 ± 0.1%. Exposure of THP-1, model cell line of AM, to the particles showed 88.8 ng of RIF/Mio cell taken up; whereas 50.5 ng /Mio cell were taken up from exposition to a RIF-solution.

Conclusion

Lipid-microparticles loaded with RIF were successfully prepared through an aqueous-based spray-drying process of high yield. The particles were produced through an exclusively lipid-based formulation using diglycerol full ester of behenic acid and glyceryl stearate citrate. Optimal particle attributes and outstanding aerodynamic performance as lipid-based DPI were achieved. Such attributes resulted in the targeted delivery of RIF to AM, which in turn offers a vast potential for dose reduction and reduced adverse side effects.

Acknowledgment: IOI Oleo GmbH, the Austrian Funding Agency (FFG) and the Doctoral Academy NanoGraz.