Sorafenib

Osimertinib and AZ5104 were purchased from Beijing Sunflower and Technology Development Co., Ltd. (Beijing, China). Omeprazole, lansoprazole, and rabeprazole were purchased from Shanghai Canspec Scientific Instruments Co., Ltd. (Shanghai, China). Sorafenib was purchased from Shanghai Macklin Biochemical Technology Co., Ltd. (Shanghai, China). Pooled RLMs and HLMs were from Corning Life Sciences Co., Ltd. Recombinant human CYP3A4 and cytochrome b5 were prepared by our group as indicated previously (Zhou et al., 2019 (link)). Reduced nicotinamide adenine dinucleotide phosphate (NADPH) was purchased from Roche Pharmaceutical Ltd. (Basel, Switzerland). All other chemicals and solvents not mentioned were of analytical grade. The information on 114 drugs is presented in Supplementary Table S1.

6‐ to 8‐week‐old male C57BL/6 mice were provided by Anhui University of Traditional Chinese Medicine. The animal experimental protocol was approved by the University Animal Care and Use Committee. Mice were randomly divided into five groups (n=6 per group) including the vehicle group, CCl₄‐treated group, CCl₄ + sorafenib (2.5, 5, 10 mg/kg)‐treated groups. The groups treated with CCl₄, CCl₄ + sorafenib were subjected intraperitoneal injections of olive oil with 10% CCl₄ (#56‐23–5, MACKLIN, China; 10 ml oil/kg; twice a week) for 8 weeks to induce liver fibrosis. The vehicle group was subjected intraperitoneal injections of the same dose of olive oil. After 4 weeks, each mouse in the groups treated with CCl₄ + sorafenib was orally administered sorafenib (#sc‐357801A, Santa Cruz, USA) daily. The vehicle group and the CCl₄‐treated group were orally administered the same dose of saline daily. 8 weeks later, all mice were euthanized; serum and livers were obtained. The serum was used to detect liver function indicators. Livers were paraffin‐embedded or stored at −80°C for further analysis.

Transfected cells were distributed into 96-well plates at a density of 4000 cells per well. Following overnight adherence, the culture medium was substituted with 100 μl of fresh medium containing varying concentrations of sorafenib (Macklin, Shanghai, China). The cells were then cultured for either 24 or 48 h. Subsequently, 5 mg/ml of MTT (Beyotime, China) was introduced to each well and incubated for 4 h. The supernatant was discarded, 150 μL DMSO (Beyotime, China) was added, and a microplate reader (ALLSHENG, Hangzhou, China) measured the OD value at 490 nm.