In Vivo Tumor Xenograft Model

A subcutaneous tumor model of breast cancer was established using both MCF-7 and MDA-MB-231 cell lines. Female 5-6-week-old athymic balb/c nude mice provided by Charles River Laboratories were housed in a specific pathogen-free environment and received food and water ad libitum. The SPF room is under a temperature of 22 ± 2°C with a 12 h light/12 h dark cycle and relative humidity of 40–60%. After the mice had been in quarantine for 1 week, 2*10⁶ MCF-7 cells suspended in a 100 μL mixture of PBS and matrigel (PBS: matrigel = 1:4) or 2*10⁶ MDA-MB-231 cells suspended in 100 μL PBS were injected subcutaneously into the right flank of each mouse. When the tumor volume reached 60 mm³, the mice were randomly allocated into two groups and treated intraperitoneally with vehicle (10% β-cyclodextrin) or JB 20 mg/kg every day. Mice weight and tumor volumes were recorded. At the end of the experiment, mice were euthanized and tumors were harvested, photographed, and weighed. All procedures of animal experiments were conducted in compliance with standard ethical guidelines and with the approval (202105007Z) of the Institutional Animal Care and Use Committee of Fudan University. According to its guidelines and suggestions, the experiment was finished before tumors grew to the permitted maximum length/width of no more than 20 mm. The MCF-7 tumors grew rapidly under the action of matrigel containing a number of growth factors and were harvested on D17. The MDA-MB-231 tumor model was established by 2*10⁶ cells suspended in PBS, and the tumors were harvested on D28. Tumor volume was calculated by vernier caliper using the following formula: tumor volume = length×width²/2.