Estimation of bioavailable concentration of endogenously formed N-nitrosamines by physiologically based kinetic modelling.

N-nitrosamines (NAs) are potentially carcinogenic organic compounds, and nitrosamine drug sub­stance-related impurities (NDSRIs) are currently regulated with class-specific thresholds in the low nanogram range according to the carcinogenic potency categorization approach (CPCA) classi­fication schema. Beyond direct exposure, NDSRIs can form endogenously in the human organism after ingestion of secondary amines. As recently shown, enalapril, propranolol, and fluoxetine form NDSRIs under conditions mimicking the acidic environment in the stomach. The MUTAMIND project investigated whether such endogenously formed NA levels lead to plasma or liver con­centrations which align with or exceed the acceptable intake limits based on the current CPCA. A generic physiologically based kinetic (PBK) model was built using compound-specific in vitro ADME parameters such as intestinal permeability and hepatic clearance. The predictions correlated well with measured in vivo ADME data for the data-rich APIs, so the same PBK approach was applied to the corresponding NDSRIs. While the modelling of propranolol was unremarkable, the highest NA conversion rate observed for N-nitrosoenalapril under gastric conditions resulted in plasma and liver levels exceeding those derived from the CPCA threshold by a factor of about 800 and 400, respectively. The long half-life of fluoxetine suggests a risk of bioaccumulation of its nitrosamine with chronic exposure. These findings indicate that PBK modelling could be a valuable tool as part of a weight of evidence approach in contributing to the risk assessment of nitrosamine impurities in pharmaceuticals.