Use of the MucilAir Airway Assay, a New Approach Methodology, for Evaluating the Safety and Inhalation Risk of Agrochemicals

Introduction: The United States Environmental Protection Agency (US EPA) has issued a directive to reduce mammalian studies to 30% by 2025. The application of new approach methodologies is being implemented to provide information on chemical hazard and risk assessment. In vitro methods, such as the MucilAir™ assay, are used to detect airway damage and acute irritation potential in agrochemicals. Materials and Methods: Four agrochemicals, including Adepidyn™ and two developmental chemicals (DC 1 and 2), were evaluated using the MucilAir assay to determine the need for future in vivo inhalation studies. Chlorothalonil (CTN), which exhibits irritant properties in mucous membranes in vivo was assessed in the MucilAir assay to inform chronic inhalation risk. Evaluation of irritation potential and tissue damage were determined through transepithelial electrical resistance (TEER), lactate dehydrogenase (LDH) release, resazurin reduction, or histology. Each agrochemical was applied to the MucilAir for 24 hours at six concentrations (Adepidyn: 0.005–0.5 mg/L; chemicals no. 1–2: 31–317 mg/L) and 10 concentrations (CTN, 2–200 mg/L). As controls, vehicle, positive (sodium dodecyl sulfate, 4 mM) and untreated air–liquid interface groups were tested in parallel. Results: The control treatments confirmed correct assay functioning for each endpoint. Exposure to Adepidyn or DC 1 and 2 did not cause a change in TEER or LDH release. There was no change in resazurin reduction. Altogether, the irritation potential panel and histological evaluation indicated no evidence of toxicity. A benchmark dose (BMD) level of 0.00730 mg/cm2 was derived for CTN based on TEER, LDH release, and resazurin endpoints. Discussion: A good concordance of response across endpoints and human tissue donors was observed. A wide range of concentrations were tested to derive points of departure for risk assessment through BMD modeling. Conclusion: The MucilAir assay evaluating chemically induced airway cytotoxicity is fit for purpose for respiratory tract hazard screening and refining inhalation risk assessment of agrochemicals.