Spray-dried nanosuspensions for pulmonary drug targeting and in vitro testing thereof

Simkova, Katerina. Spray-dried nanosuspensions for pulmonary drug targeting and in vitro testing thereof. 2021, Doctoral Thesis, University of Basel, Faculty of Science.


Official URL: https://edoc.unibas.ch/83988/

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Pharmaceutical formulation development can be a challenging task during which many, sometimes contradicting factors need to be considered such as the formulation composition, suitability of excipients for each manufacturing step, or scalability of the process(es) applied. Development of dry powder formulations for inhalation can be especially challenging as only particles of very narrow aerodynamic particle size can enter the lungs while they are in fact designed by nature to prevent entry of foreign material. Additionally, there are mechanisms in place that quickly remove any particles that do enter them. Formulations with micronised drug substance particles are mostly being used to bypass the anatomical obstacles but the drug’s deposition efficiency is commonly rather low. Utilisation of particle engineering, which leverages on favourable particle properties being tailored through a manufacturing process, offers a possibility to address these anatomical and performance challenges; however, their manufacturing is often linked to usage of organic solvents. A formulation scientist also needs to keep in mind that for any kind of formulation, drug dissolution is the key prerequisite for bioavailability and thus also for the drug’s action. This can be a challenging step especially for a poorly water–soluble drug depending on its solubilities in the respective biological fluids as these are made up of water most of the time. Due to the obstacles inhaled drug particles encounter on their way to dissolved state, the drug needs to dissolve before it is cleared by the body’s defence mechanisms to effectively work. Thus, study of the dissolution kinetics of solid formulations is of considerable interest. Unlike for orally delivered drug substance, no fully established pharmacopoeial dissolution method for inhaled medicines exists, likely due to the complexity to mimic reasonably well the in vivo situation.
The objective of this work was to combine industry-viable manufacturing processes to engineer inhalation powders of superior pharmaceutical performance and prolonged lung residence time thanks to favourable physical-chemical properties, and to study dissolution behaviour and biological fate of these powders using a dissolution vessel and in vitro cell culture system, respectively. The work intentionally used industrially established processes and only water as a solvent to offer the possibility to ultimately introduce this as platform technology for pulmonary formulation development allowing optimised inhalation drug product performance.
High-energy wet media milling was used for production of nanoparticles of a poorly-water soluble model drug (budesonide). The nanosuspension was stabilised using D-α-tocopherol polyethylene glycol 1000 succinate, which allowed to create nanosuspension of ≈ 260 nm median particle size at specific energy input of ≈ 137 MJ/kg. Subsequently used particle engineering via spray drying aimed to create composite particles of maximal fine particle fraction for deep lung deposition, maximal geometric size, minimal density, and fast dissolution. Spray drying at high Peclet numbers was crucial to successfully achieve this goal and it guided the choice of formulation composition and process parameters. Among the tested additives were leucine, trileucine, mannitol, albumin, glycine, and a pore former ammonium carbonate. Different spray drying temperatures and atomising pressures, as well as feedstock concentration, were screened to obtain formulation of the above-mentioned desirable properties. The most favourable formulation, prepared with the nanoparticulate drug together with leucine and ammonium carbonate, exhibited a high fine particle fraction of 61% as assessed by the next generation impactor deposition and had a median particle size of ≈ 4.4 µm. The deposition efficiency correlated well also with the bulk and effective density measurements for which this formulation had lowest and second lowest values, respectively. The aerodynamic performance was well above the commercial, carrier-based product, which reached only 21% of fine particle fraction. Interestingly, when the micronised drug was not processed by wet media milling, even with the use of same additives and process conditions, only 22% of fine particle fraction could be reached. This suggested that the wet media milling step was indispensable for improved aerodynamic performance and that only the combination of the processes allowed to harness the advantages of both. Moreover, the geometric size of the composite spray-dried particles was larger than that of the micronised drug, offering additionally the potential to evade phagocytosis for a longer time period compared to the micronized drug as this is a size driven process.
Within the first part of the work, five formulations of comparable geometric particle size, but different densities and particle shapes were studied in depth to assess their dissolution behaviour. For this purpose, a USP2 paddle apparatus was modified with the aim to mimic closely the in vivo conditions in terms of liquid hydrodynamics and volumes. For the modification, an insert from impactor stage, on which aerodynamically classified particles were deposited, was placed into the dissolution vessel; the setup thus resembled a USP5 (paddle-over-disk) apparatus. Using such setup, the dissolution behaviour of powders from three different stages was studied as a function of particle properties such as aerodynamic particle size, shape, or specific surface area. A permeable polycarbonate membrane was fixed onto each insert, which enabled the creation of an inner and outer compartment of different volumes. In the inner compartment, between the membrane and the insert surface, a small liquid amount of 200 µL was in direct contact with the powders and allowed dissolution in small, unstirred liquid volume. In the outer compartment was 300 mL of the stirred dissolution media into which the drug permeated upon its dissolution in the inner compartment. Dissolution of all aerodynamically classified fractions showed a very fast onset and was largely completed within 30 minutes irrespective of the formulation and the impactor stage. To further analyse this observation, mathematical kinetic modelling was used to deduce the drug’s dissolution rate coefficients in each formulation in all three stages. From this it was found that the dissolution rate was determined by the properties of the drug nanoparticles, mainly particle size, rather than by the variable properties of the composite microparticles., This then explained why no differences among the formulations and stages were observed when same drug nanoparticles were used in the formulations.
Second part of the work aimed to investigate these aerodynamically classified composite powders even further using an in vitro cell culture system, which should have provided more representative in vivo conditions compared to the dissolution vessel. For this purpose, the next generation impactor was successfully modified for the first time to allow deposition on an A549 cell culture, cultivated on a low-profile, Matrigel®-coated cell culture insert. These alveolar type II cells were grown at an air liquid interface, which allowed formation of a surfactant layer similar to the one present in the alveolar lung region. It was again of interest to evaluate whether particle properties like shape, density, or size affect the dissolution behaviour in this miniaturised setup. In this setup, it was assumed that the drug dissolution starts immediately upon particle deposition and any dissolved solute is translocated into the intracellular compartment, where it may be metabolised, and is eventually transported into the basal compartment. The drug amount in the basal solution was determined for four to eight hours upon deposition, while its amount on the cell surface and in the interior of the cell monolayer was evaluated at the end of the experiments. Any induced cell damage was assessed also at the end of the experiment by measurement of the lactate dehydrogenase leakage from the cell membrane. Significant differences in the total deposited drug amount and the amount remaining on the cell surface at the end of the experiment were found between different formulations and impactor stages. The deposited amount negatively affected the dissolution of the drug as it took rather long (≥ 4 hours) for larger powder amounts to dissolve despite the drug’s nano-range size. In fact, the dissolution took considerably longer than in the dissolution vessel setup, implying potential negative impact on local bioavailability as alveolar phagocytic clearance has similar half-life. Prolonged time required for complete drug dissolution and cell uptake in case of the large deposited powder amounts also suggested initial drug saturation of the surfactant layer. Interestingly, irrespective of the stage or formulation, roughly half of the deposited drug amount was taken up by the cells and metabolised to a large extent to its metabolic conjugate with oleic acid. Additionally, kinetic modelling was performed to evaluate the kinetics of drug dissolution and its uptake into the cells, metabolism into the oleate metabolite, and release into the basal solution, and supported the conclusions made based on the experimental results. However, it is important to note that partial cell damage was observed, which was possibly caused by the impaction of particles on the cells. This clearly indicated the need to further improve the experimental setup to reduce the cells membrane damage.
This work provided many insights into dry powder for inhalation formulation development and in vitro testing of inhalation powders. It focused on formulation composition and process optimisation and use of industrially established, solely water-based processes. This potentially allows establishment of the presented approach as a platform technology. It clearly showed that when formulating inhalation powders using this platform, equal importance needs to be given to drug pre-processing by particle size reduction as to spray drying if advantageous aerodynamic performance over carrier-based formulations should be achieved. It also showed that large particles of low density and enhanced aerodynamic performance, which could be used for targeted drug delivery, can be engineered using only water-based processes. Thorough in vitro testing using two different drug dissolution configurations showed the clear need to consider the test’s purpose to select a relevant setup. For predicting local bioavailability, the newly developed cell culture in vitro system was able to provide useful insights into the process and kinetics of drug dissolution and cell uptake following powder deposition on an alveolar cell surface and it further highlighted the importance of fluid volume for formulation properties’ study. As stage-specific drug distribution in different cell compartments and drug’s amount in each compartment are relevant for local bioavailability and therapeutic effect, this setup offers more possibilities for biopharmaceutical performance prediction of dry powder for inhalation formulations while using state-of-the-art equipment.
Advisors:Imanidis, Georgios
Committee Members:Štěpánek, František
Faculties and Departments:05 Faculty of Science
UniBasel Contributors:Imanidis, Georgios
Item Type:Thesis
Thesis Subtype:Doctoral Thesis
Thesis no:14242
Thesis status:Complete
Number of Pages:113
Identification Number:
  • urn: urn:nbn:ch:bel-bau-diss142425
edoc DOI:
Last Modified:10 Sep 2021 04:30
Deposited On:09 Sep 2021 07:49

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