Real-Time Measurement of Cell Mechanics as a Clinically Relevant Readout of an In Vitro Lung Fibrosis Model Established on a Bioinspired Basement Membrane.

Lung fibrosis, one of the major post-COVID complications, is a progressive and ultimately fatal disease without a cure. Here, an organ- and disease-specific in vitro mini-lung fibrosis model equipped with noninvasive real-time monitoring of cell mechanics is introduced as a functional readout. To establish an intricate multiculture model under physiologic conditions, a biomimetic ultrathin basement (biphasic elastic thin for air-liquid culture conditions, BETA) membrane (<1 µm) is developed with unique properties, including biocompatibility, permeability, and high elasticity (<10 kPa) for cell culturing under air-liquid interface and cyclic mechanical stretch conditions. The human-based triple coculture fibrosis model, which includes epithelial and endothelial cell lines combined with primary fibroblasts from idiopathic pulmonary fibrosis patients established on the BETA membrane, is integrated into a millifluidic bioreactor system (cyclic in vitro cell-stretch, CIVIC) with dose-controlled aerosolized drug delivery, mimicking inhalation therapy. The real-time measurement of cell/tissue stiffness (and compliance) is shown as a clinical biomarker of the progression/attenuation of fibrosis upon drug treatment, which is confirmed for inhaled Nintedanib-an antifibrosis drug. The mini-lung fibrosis model allows the combined longitudinal testing of pharmacodynamics and pharmacokinetics of drugs, which is expected to enhance the predictive capacity of preclinical models and hence facilitate the development of approved therapies for lung fibrosis.

Aggregates Associated with Instability of Antibodies during Aerosolization Induce Adverse Immunological Effects.

BACKGROUND: Immunogenicity refers to the inherent ability of a molecule to stimulate an immune response. Aggregates are one of the major risk factors for the undesired immunogenicity of therapeutic antibodies (Ab) and may ultimately result in immune-mediated adverse effects. For Ab delivered by inhalation, it is necessary to consider the interaction between aggregates resulting from the instability of the Ab during aerosolization and the lung mucosa. The aim of this study was to determine the impact of aggregates produced during aerosolization of therapeutic Ab on the immune system. METHODS: Human and murine immunoglobulin G (IgG) were aerosolized using a clinically-relevant nebulizer and their immunogenic potency was assessed, both in vitro using a standard human monocyte-derived dendritic cell (MoDC) reporter assay and in vivo in immune cells in the airway compartment, lung parenchyma and spleen of healthy C57BL/6 mice after pulmonary administration. RESULTS: IgG aggregates, produced during nebulization, induced a dose-dependent activation of MoDC characterized by the enhanced production of cytokines and expression of co-stimulatory markers. Interestingly, in vivo administration of high amounts of nebulization-mediated IgG aggregates resulted in a profound and sustained local and systemic depletion of immune cells, which was attributable to cell death. This cytotoxic effect was observed when nebulized IgG was administered locally in the airways as compared to a systemic administration but was mitigated by improving IgG stability during nebulization, through the addition of polysorbates to the formulation. CONCLUSION: Although inhalation delivery represents an attractive alternative route for delivering Ab to treat respiratory infections, our findings indicate that it is critical to prevent IgG aggregation during the nebulization process to avoid pro-inflammatory and cytotoxic effects. The optimization of Ab formulation can mitigate adverse effects induced by nebulization.