Penicillin streptomycin

Two lines of human gastric cancer cells (AGS and HGC-27) were purchased from Procell Life Science & Technology Co., Ltd (Wuhan, China), and the other GC cell lines (GT38 and SNU-719) were purchased from Honsun Biological Technology Co., Ltd (Shanghai, China). AGS cells were cultured in Hams' F12 medium (Shanghai BasalMedia Technologies Co., Ltd.) that was supplemented with 10% fetal bovine serum (FBS; ExCell Bio. Jiangsu, China) and 1% penicillin-streptomycin (Shanghai BasalMedia Technologies Co., Ltd.). HGC-27, SNU719, and GT38 cells were cultured in RPMI-1640 medium (Shanghai BasalMedia Technologies Co., Ltd.) that was supplemented with 10% FBS and 1% penicillin-streptomycin. All cells were cultured in a humidified incubator with 5% carbon dioxide (Thermo Fisher Scientific, USA).

Human embryonic kidney 293 T (HEK293T), human mammary epithelial cell lines (HMEC and MCF10A), human breast cell lines (MDA‐MB‐231, MDA‐MB‐453, MDA‐MB‐468, MDA‐MB‐157, Hs578T, BT549, BT20, HCC1806, and HCC1937) cell lines were obtained from Shanghai Key Laboratory of Breast Cancer, Fudan University Shanghai Cancer Center. MDA‐MB‐231‐derived LM2‐4175 cells were obtained from Guohong Hu (University of Chinese Academy of Sciences), and SUM159PT cell line was kindly provided by Asterand. The above cell lines were authenticated by short tandem repeat profiling and monitoring cell vitality. MCF10A and HMEC cells were cultured in DMEM (BasalMedia, #L110) containing 5% donor horse serum (Gibco, #16050–114), 20 ng/ml epidermal growth factor (Sino Biological Inc, #10605 HNAE), 0.5 mg/ml hydrocortisone (Yeasen, #40109ES08), 10 mg/ml human recombinant insulin (Yeasen, #40107ES76), and 1% penicillin/streptomycin (BasalMedia, #S110B). The others were maintained in DMEM with 10% fetal bovine serum (Gibco, #10270–106), and 1% penicillin/streptomycin (BasalMedia, # S110B). Transwell chambers and Cell Counting Kit‐8 (CCK8) were purchased from Corning Falcon (#353097) and Yeasen (#40203ES92), respectively.

HEK293T cells were obtained from the American Type Culture Collection (Manassas, VA, USA) and maintained in suspension culture in Balance CD (Chuangling Cell-wise, Shanghai, China, CW01001) medium supplemented with 1% penicillin/streptomycin (BasalMedia, Shanghai, China, S110JV) at 37 °C and 5% CO₂. Sixteen hours before transfection, cells were transferred to adherent cultured in DMEM (BasalMedia, L110KJ) with 2% fetal bovine serum (Thermo Fisher Scientific GIBCO, Waltham, MA, USA, 10099141C) and 1% penicillin/streptomycin (BasalMedia, Shanghai, China, S110JV). Briefly, AAV vectors were produced by triple plasmid transient transfection with linear polyethylenimine (Polysciences, Warrington, PA, USA, 24765-1), viral particles were harvested at 72 h post-transfection and purified via iodixanol gradient ultracentrifugation [42 (link)], and buffer exchange was performed by using phosphate buffered saline (PBS) with 0.001% Pluronic F68 (Thermo Fisher Scientific, Waltham, MA, USA, 24040032) via Amicon filtration (Merck Millipore, Billerica, MA, USA, UFC910024). The purified rAAVs were titered via qPCR using the iQ SYBR Green Supermix kit (Bio-Rad, Hercules, CA, USA, 1708884). The obtained virus titers are shown in Supplementary Table S1. All viral vectors were aliquoted and stored at −80 °C until use.