Tenofovir Alafenamide is Hydrolyzed by Cathepsin A and Carboxylesterase 1 in the Human Liver and the Hydrolysis is Affected by CES1 Genetic Variation

Tenofovir alafenamide (TAF), a novel prodrug of tenofovir, is increasingly used to treat patients with chronic hepatitis B. Despite its improved efficacy and safety over tenofovir disoproxil and tenofovir, TAF therapy is associated with significant interindividual variability in clinical response. As a prodrug, TAF must be metabolized to its active metabolite in the liver to exert its intended anti‐hepatitis B virus effect. Thus, understanding the enzymes responsible for TAF activation and their regulation will lead to the improvement of TAF pharmacotherapy. TAF activation involves multiple steps, and the first step is a hydrolysis reaction catalyzed by hydrolases. In the present study, we quantified various hydrolases in human liver S9 fractions (HLS9) and determined the contributions of the hydrolases to TAF hydrolysis using the previously reported TAF hydrolysis activity from individual recombinant hydrolases. Our results suggested that cathepsin A (CatA) and carboxylesterase 1 (CES1) contribute to 55.0% and 44.4%, respectively, of TAF hydrolysis in the human liver. The findings were further confirmed by a TAF incubation study with HLS9 and the CatA inhibitors bortezomib and boceprevir, and the CES1 inhibitor bis‐(p‐nitrophenyl) phosphate. Moreover, the in vitro study of cell lines stably transfected with wild‐type CES1 and the CES1 variant G143E (rs71647871) revealed that G143E is a loss‐of‐function variant for CES1‐mediated TAF hydrolysis. In summary, our results demonstrated that TAF is hydrolyzed by CatA and CES1 in the human liver. Additionally, TAF activation could be affected by CES1 genetic variation and CatA and CES1 inhibitors.