Zanubrutinib

Human cancer cell lines (obtained from ATCC, USA, or DSMZ, Germany) were cultured according to the distributors’ instructions. In brief, MCF7, SKBR3, BT20, JIMT1 and BT474 cells were maintained in Dulbecco’s modified Eagle’s medium supplemented with 10–15% FBS, and T47D, HCC1806 and EFM192A cells were maintained in RPMI 1640 medium supplemented with 10–20% FBS. All media were supplemented with 100 U/mL penicillin, 100 µg/mL streptomycin, and 2 mM glutamine. For treatment, cells were seeded at densities of 1.5-2 million cells per dish in 60 mm dishes and allowed to adhere overnight. The BTK inhibitors ibrutinib, evobrutinib, tirabrutinib, acalabrutinib and zanubrutinib were purchased from MedChemExpress, and the EGFR/HER2 inhibitor lapatinib was purchased from LC Laboratories.

Tirabrutinib, ibrutinib, and acalabrutinib were obtained from the Department of Drug Discovery Chemistry, Ono Pharmaceutical Co., Ltd. (Osaka, Japan). Zanubrutinib and refametinib were purchased from MedChemExpress (Princeton, NJ, USA). BMS-345541 and MK-2206 were purchased from Selleck Chemicals (Houston, TX, USA) and Shanghai Haoyuan Chemexpress (Shanghai, China), respectively. Human peripheral blood mononuclear cells (PBMC) were obtained from VERITAS Corporation (Tokyo, Japan; Lot No. A1929). The activated B-cell-like (ABC)-DLBCL cell lines TMD8 and U-2932 have been described previously [24 (link),25 (link)]; TMD8 cells were obtained from the Tokyo Medical and Dental University (Tokyo, Japan), and U-2932 cells from Deutsche Sammlung von Mikroorganismen und Zellkulturen GmbH (DSMZ, Braunschweig, Germany). PBMC and these cell lines were maintained in a culture medium, namely RPMI medium 1640 (Invitrogen Corporation) containing 10% v/v inactivated fetal bovine serum (Invitrogen Corporation) and 1% v/v penicillin-streptomycin liquid (Invitrogen Corporation) at 37°C, 5% CO₂/95% air.