Multilayered Cultures of NSCLC cells grown at the Air-Liquid Interface allow the efficacy testing of inhaled anti-cancer drugs

Abstract

Evidence supports the advantages of inhalation over other drug-administration routes in the treatment of lung diseases, including cancer. Although data obtained from animal models and conventional in vitro cultures are informative, testing the efficacy of inhaled chemotherapeutic agents requires human-relevant preclinical tools. Such tools are currently unavailable. Here, we developed and characterized in vitro models for the efficacy testing of inhaled chemotherapeutic agents against non-small-cell lung cancer (NSCLC). These models recapitulated key elements of both the lung epithelium and the tumour tissue, namely the direct contact with the gas phase and the three-dimensional (3D) architecture. Our in vitro models were formed by growing, for the first time, human adenocarcinoma (A549) cells as multilayered mono-cultures at the Air-Liquid Interface (ALI). The in vitro models were tested for their response to four benchmarking chemotherapeutics, currently in use in clinics, demonstrating an increased resistance to these drugs as compared to sub-confluent monolayered 2D cell cultures. Chemoresistance was comparable to that detected in 3D hypoxic tumour spheroids. Being cultured in ALI conditions, the multilayered monocultures demonstrated to be compatible with testing drugs administered as a liquid aerosol by a clinical nebulizer, offering an advantage over 3D tumour spheroids. In conclusion, we demonstrated that our in vitro models provide new human-relevant tools allowing for the efficacy screening of inhaled anti-cancer drugs.

Non-Coding RNAs in Lung Tumor Initiation and Progression

“…Due to limited current data, future study of targeting common cancer initiating genes with non-coding RNAs using organoid models could serve as promising indicators for clinical therapeutic response and tumorigenesis. A form of in vitro modeling includes air-liquid interface models, which attempt to mimic the microenvironment of the lungs [301]. Movia et al utilized these multilayered cultures to test response chemotherapy administered through inhalation and they found that these models demonstrated high chemoresistance [301].…”

“…A form of in vitro modeling includes air-liquid interface models, which attempt to mimic the microenvironment of the lungs [301]. Movia et al utilized these multilayered cultures to test response chemotherapy administered through inhalation and they found that these models demonstrated high chemoresistance [301]. Further study of therapeutic efficacy using miRNAs by mimicking the lung microenvironment with air-liquid interface models could serve as promising future study.…”

Preclinical Development of Orally Inhaled Drugs (OIDs)—Are Animal Models Predictive or Shall We Move Towards In Vitro Non-Animal Models?

“…Also, they can replicate some of the key features that need to be kept into account when developing an inhalation therapy, namely (i) the constitution and thickness of the pulmonary lining fluid [67] and (ii) mucociliary clearance [60][61][62]. For example, ALI cultures have been used to model the effects of smoke exposure on epithelial cells [104] and the authors have created a complex, diseased ALI culture model capable of reproducing the chemoresistance mechanisms observed in patients affected by non-small-cell lung cancer [105,106]. Also, culturing human airway epithelial cells isolated from patients, makes it possible to conduct patient-specific research and drug-screening, for example in cystic fibrosis, asthma and COPD [107][108][109][110].…”

Modifying and Integrating in vitro and ex vivo Respiratory Models for Inhalation Drug Screening

“…Despite these limitations, primary respiratory epithelial ALI culture models have been used to recapitulate in vivo airway epithelia as a biologically relevant in vitro drug screening platform (Ong et al, 2016). ALI culture systems have been heavily reviewed in the literature and have delivered highly impactful research outcomes (Movia et al, 2018;Upadhyay and Palmberg, 2018) however, the use of in vitro ALI models to determine toxicity, delivery and efficacy of inhaled therapeutics requires greater physiological and anatomical relevance. Specifically, in vitro respiratory models that implement breathing mechanics, aerosol deposition, and the co-culture of different cell types allow greater mechanistic and efficacious insights of novel inhalable therapeutic compounds.…”