The Role Of ?-adrenergic Receptor Blockers And COX-2 Inhibitors In Liver Metastasis Of Colorectal Cancer

Background:Colorectal cancer(CRC)is one of the most common malignancies in the digestive tract.With the changes of living conditions and dietary habits in China,the incidence of CRC is rising.Till now,there are many new progresses in the treatment of colorectal cancer,but the metastasis rate and mortality rate are still at a high level.It has been reported that 20-40%of patients who diagnosed with colorectal cancer has the liver metastases simultaneously.And the rate of recurrent liver metastasis after radical surgery is about 50%.Furthermore,the mortality rate of in patients with liver metastases is as high as 45-71%.Although the comprehensive treatment of colorectal cancer has improved a lot,no big breakthrough has been made yet.Since of the recurrence and metastasis,the prognosis of CRC is poor.Liver is the most common metastatic site of colorectal cancer.Current treatments for liver metastases of colorectal cancer include surgery,chemotherapy,radiotherapy,radiofrequency ablation,targeted therapy and so on.Surgery is still the most important therapy.For patients who cannot get operated,chemotherapy and targeted therapy are very important.Parts of patients can have the chance of surgical resection through conversion therapy.However,most of the chemotherapeutic drugs and targeted drugs are expensive,and the side effects make it difficult for some patients to receive complete treatment.Therefore,searching for new anti-tumor drugs has always been the interest of researchers.?-adrenergic receptor is a G-protein coupled receptor,which,upon activation,regulates tumor cell growth and promotes tumor cell angiogenesis through a variety of signal transduction pathways.Cyclooxygenase-2(COX-2)has the dual function of cyclooxygenase and peroxide synthase.Studies have shown that COX-2 can inhibit apoptosis and induce epidermal growth factor receptor to promote tumor development.The role of ?-adrenergic receptor blockers and COX-2 inhibitors in liver metastasis of colorectal cancer has not been reported yet.Aims:To explore the role and possible mechanisms of ?-adrenergic receptor blockers and COX-2 inhibitors in liver metastasis of colorectal cancer.Methods:1.CCK-8 assay was used to detect the effects of different concentrations of?-adrenergic receptor blockers(propranolol)and COX-2 inhibitors(etodolac)on the proliferation of the mouse colon cancer cell CT-26.2.The effects of different concentrations of propranolol and etodolac on the secretion of vascular endothelial growth factor(VEGF)in colon cancer cells was studied by enzyme-linked immunosorbent assay(ELISA).3.Flow cytometry was used to detect the effects of different concentrations of propranolol and etodolac on the expression of reactive oxygen species(ROS)in colon cancer cells.4.Nude mice were pre-treated with propranolol and etodolac,followed by the construction of animal model of colorectal cancer liver metastasis,the number of metastasis tumor,maximum weight of tumor and VEGF mRNA expression level in mouse model were determined.Results:1.Propranolol and etodolac both had inhibiting effects on colon cancer cells in a dose-dependent manner.After 50,100 and 200 ?M propranolol stimulation of CT-26 cells for 48 h,the cell proliferation inhibition rates were(81.00±4.58)%,(70.33±2.08)%and(39.00±2.00)%,respectively.The difference was statistically significant(p<0.05).The cell proliferation inhibition rates were(75.33±3.06)%,(57.00±2.65)%and(33.67± 1.15)%,respectively after treated with 0.5,1,2 ?M etoposide for 48 h.2.Propranolol and etodolac inhibited the secretion of VEGF in a dose-dependent manner.ELISA was used to detect the concentration of VEGF in the supernatant of CT-26 cells treated with propranolol and etodolac.The VEGF concentration decreased from 95.94±7.94 pg/mL to 55.44±7.65 pg/mL with increasing concentration of propranolol treatment.The difference was statistically significant(p<0.05).Compared with the control group(127.97±7.72 pg/mL),the VEGF secretion in CT-26 cells treated with 0.5,1,2 ?M etodolac decreased to 81.97±2.39pg/mL,58.70±3.52 pg/mL and 37.75±4.08 pg/mL,respectively.The difference between each group was statistically significant(p<0.05).3.Propranolol and etodolac promoted the expression of ROS in CT-26 cells.Expression of ROS in CT-26 cells after propranolol and etodolac treatment was determined by flow cytometry.Results showed that the rates of ROS positive cell after propranolol stimulation at 50,100 and 200 ?M were 8.1%,18.0%and 33.2%,respectively.And the rates of ROS positive cell were 14.6%,29.6%and 56.4%,respectively after 0.5,1,2 ?M etodolac stimulation.4.Propranolol and etodolac inhibited the liver metastasis of colon cancer cells,tumor proliferation,and down-regulated the expression of VEGF.Furthermore,the combined effects of the propranolol and etodolac are better.We constructed the liver metastasis model of CT-26 cells in nude mice,all the mice in the control group developed liver metastases and the tumor metastasis rate was 100.0%.12 mice developed liver tumor after propranolol treatment,the metastasis rate was 80.0%.8 mice in the etodolac-treated group developed liver metastasis and tumor metastasis rate was 53.3%.5 mice were found liver metastases in in propranolol/etodolac group,the metastatic rate was 33.3%.PCR was used to detect the expression of VEGF mRNA in tumor tissues.Compared with the control group,the expression of VEGF in the propranolol-treated group was 0.75±0.08,0.50±0.08 in the etodolac-treated mice,and 0.22±0.06 in the propranolol/etodolac treated-group.All were statistically significant(p<0.05).Conclusions:1.?-adrenergic receptor blockers and COX-2 inhibitors can inhibit the proliferation of colon cancer cells,and the possible mechanism is to promote ROS expression and induce cells apoptosis.2.?-adrenergic receptor blockers and COX-2 inhibitors can inhibit the secretion of VEGF in colon cancer cells.Furthermore,the combined two have synergistic effects,thereby inhibiting neovascularization of tumor cells and inhibiting liver metastasis of colon cancer cells.3.?-adrenergic blockers and COX-2 inhibitors may be a potential drug for the treatment of colorectal cancer.