(3s) Characterization of Novel Inhalable Dry Powder Particles Containing Paclitaxel and the Development of Three-Dimensional Multicellular Spheroids for the Treatment and Study of Lung Cancer

Despite the significant advances in the treatment of lung cancer, it is a disease which still signifies poor prognosis and challenges in implementation of treatment. Targeted pulmonary inhalation drug delivery offers many advantages for lung cancer patients in comparison to conventional systemic chemotherapy. These include: the potential to deliver local therapeutically effective concentrations of drug directly to the lung, minimized side effects due to limited systemic delivery, and ease of use for the patient. Inhalable dry powder formulations of nanoparticles and microparticles containing a chemotherapeutic are advantageous in their ability to deliver drug deep in the lung via optimally sized particles, higher local drug dose delivery, and long-term storage capability. In this work, novel advanced spray-dried inhalable PEGylated phospholipid microparticle/nanoparticle powders containing the chemotherapeutic paclitaxel were successfully designed and produced via dilute organic solution spray drying under various conditions. Fixed ratios of dipalmatoylphosphatidylcholine (DPPC) and dipalmatoylphosphatidylethanolamine poly(ethylene glycol) (DPPE-PEG) were mixed with various ratios of paclitaxel in a dilute methanol solution. Upon optimization of the spray drying conditions (e.g. pump rate), the physicochemical characterization of the particles was completed. Scanning electron microscopy (SEM) images showed the spherical particle morphology of the inhalable particles. The size of the particles was statistically analyzed using these images SigmaScan software and determined to be 600 nm – 1.2 μm in diameter, which is optimal for deep lung alveolar penetration. Differential scanning calorimetry (DSC) and x-ray powder diffraction (XRPD) were performed to analyze solid-state transitions and long-range molecular order, respectively, and allowed for the confirmation of the presence of phospholipid bilayers and/or paclitaxel and their physical states. The amount of paclitaxel loaded into the particles was analyzed via high performance liquid chromatography (HPLC) and their aerosol performance was evaluated using a Next Generation Impactor (NGI) to determine the fine particle fraction and aerodynamic diameter. These results demonstrate this novel therapeutic platform as one capable of effectively delivering paclitaxel directly to the lung for the treatment of lung cancer.

As for all cancer but particularly for lung cancer, applying in vitro outcomes to in vivo applications has limitations because conventional two-dimensional (2D) cell culture does not recreate a physiologically representative model for cells. We investigated a three-dimensional (3D) cell culture technique to model lung tumors in vitro.  A 3D lung cancer model was created by applying collagen (a semi-non-adhesive material) to a cell culture Transwell, which allows for nutrient transfer through the collagen. Two lung cancer cells lines (H358, a bronchioalveolar carcinoma and A549, a lung adenocarcinoma) were seeded on the collagen. The non-adhesive collagen caused the cells to be more inclined to attach to one another rather than its surface allowing for the formation of multicellular spheroids (MCS). To better mimic the environment for lung cancer specifically, an air-interface culture (AIC) condition was created. For AIC conditions, the cell media on the apical side of the Transwell was removed and the basolateral side with media provided effective nutrient transport to the MCS while still exposing the cells to air. The AIC model yielded viable MCS at sizes similar to MCS formed in LCC conditions (100 to 150 μm in diameter). With the optimized 3D model, LCC cells were exposed to paclitaxel in media, and paclitaxel-loaded dry powder aerosol particles described above were delivered to AIC cells through direct application with an insufflator. Using viability analysis, it was shown that both applications of paclitaxel showed variance in efficacy when comparing 2D and 3D culture conditions (where the IC₅₀ values for paclitaxel were higher for 3D compared to 2D).  Transepithelial electrical resistance of A549 cell monolayers was evaluated before and after particle delivery to illustrate that the particle application does not affect the permeability of the cells, which infers that this form of drug therapy will not affect the permeability of lung tissue. Overall, a much more representative in vitro model has been developed that is expected to be an improved predictor of efficacy of alternative drug delivery methods such as direct pulmonary delivery for lung cancer, which could lead to more efficient drug therapies for lung cancer patients.