Epirubicin

Cell lines

human glioma cell line U-87

Preparation Method

The human glioma cell line U-87 cells were cultured in a standard tissue culture incubator (37˚C; 5% CO₂; 95% air; 100% humidity). Cells were regularly passaged by treatment with trypsin (0.05%) and were grown in Eagle's minimum essential medium (EMEM) supplemented with 10% fetal bovine serum (FBS), penicillin and streptomycin, non-essential amino acids, 1mg/ml glucose and 1mM pyruvate. The U-87 and neuronal cells were plated on 96well flatbottomed microplates at a density of 1×10⁴cells/well in 100µl complete growth medium. Prior to drug treatment, the growth medium was substituted with fresh medium containing 2% FBS. The U-87 and neuronal cells were exposed to Epirubicin at concentrations of 0.5-100µM. After 48h of incubation at 37˚C in a humidified atmosphere of 5% CO₂, the cytotoxic effect of Epirubicin was estimated using an MTT assay. DNA synthesis in proliferating cells was evaluated by measuring Bromodeoxyuridine (BRDU) incorporation using a commercial Cell Proliferation ELISA System. MMP-9 levels in the conditioned media of the U-87 cells were determined using a commercial human MMP-9 immunoassay kit. Tumor cell migration was assessed using the wound healing assay.

Reaction Conditions

0.5-100µM; 48h

Applications

Epirubicin dose-dependently suppressed U-87 glioma cell proliferation, reduced MMP-9 secretion, and inhibited cell migration.

Animal models

Female BALB/c mice

Preparation Method

Female BALB/c mice (8 weeks, weighting 18 to 22g) were housed in a specified chamber with controlled air conditions (temperature 20-25°C, humidity 50–65%) and free access to sterile food and water. A 4T1 cell suspension (100μL, 5×10⁶cells/mL) was injected subcutaneously into the right side of the fourth mammary gland of each mouse. The tumor length (L) and width (W) were measured every other day, and the tumor volume was calculated by using the following formula: V (mm³)=(L×W²)/2. On day 8 of 4T1 cells injection, the size of tumors was approximately 100 mm³. Mice were randomly divided into two groups and accepted treatments every other day for eight times: Control group (Cont, n=8): mice received intragastrically administered (i.g.) and a tail intravenous injection of saline. Epirubicin group (n=8): mice received i.g. saline and a tail intravenous injection of Epirubicin (2.5mg/kg). The mice weights were recorded every other day. Blood was collected and the mice were sacrificed after 16 days. The tumors, hearts, and other tissues were removed rapidly and weighed. Several tissues were fixed in formalin for HE staining, TdT-mediated dUTP nick end labeling, and immunohistochemistry, whereas others were rapidly frozen in liquid nitrogen and stored in −80°C for Western blotting analysis.

Dosage form

2.5mg/kg; every other day for 16 days; i.v.

Applications

Epirubicin reduced tumor volume and increased tumor cell apoptosis in 4T1 breast cancer-bearing BALB/c mice.

References:
[1] Wang XF, Zhao ZF, Chen MH, Yuan QH, Li YL, Jiang CL. Epirubicin inhibits growth and alters the malignant phenotype of the U87 glioma cell line. Mol Med Rep. 2015;12(4):5917-5923.
[2] Wu J, Xue X, Zhang B, et al. Enhanced antitumor activity and attenuated cardiotoxicity of Epirubicin combined with Paeonol against breast cancer. Tumour Biol. 2016;37(9):12301-12313.