267894 Development of Three-Dimensional Lung Multicellular Spheroids in Air and Liquid Interface Culture for the Evaluation of Anti-Cancer Therapeutics
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. This work investigated a three-dimensional (3D) cell culture technique to model lung tumors in vitro. A 3D lung cancer model was created by applying collagen 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. 2D versus 3D cell behavior in response to paclitaxel and aerosol particles containing paclitaxel was evaluated using the viability of the cells.
Through the use of brightfield and fluorescence microscopy imaging, 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 were delivered to AIC cells through direct application with an insufflator. This alternative delivery method using dry powder particles was used since paclitaxel could not be delivered through media as in LCC conditions. The particles containing paclitaxel were comprised of a PEGylated phospholipid excipient mixture which encapsulated the drug. 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.
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