| Background and Objective:The liver cancer is the major cancer in China,The annual incidence rate is 10-150/100000 people.It is also the common diseases in the world,there are about 260,000 new cases of primary liver cancer each year,about 110,000 people died of liver cancer in China each year,that is 45 per cent of liver cancer deaths in the the world.Most patients diagnosed as liver cancer are already in advanced,the operation rate only 5%-10%.With radiation therapy equipment and technology development,and the Three-dimensional Conformal Radiotherapy(3DCRT) technology applicated in clinic,the radiotherapy has gradually become one of the main methods of non-surgical treatment on liver cancer.However,due to liver cancer cells have the relative resistance of radiation,making the cure dose of liver cancer was higher than the tolerated dose of the liver,especially in the most advanced liver cancer are the basis of liver disease,also limit the radiation dose increased,leading to liver cancer radiotherapy ineffective.Therefore,if we can select a drug combined with radiation therapy to increase radiation sensitivity of liver cancer cells(Radiosensitivity) on radiotherapy,then to improve the liver relative tolerance dose and improve the effect of advanced liver cancer is one of the important means with important value in clinical application.The research of radiation sensitizer has been aconcerned with always,the early studying about the pro-electronics of nitro compounds was restricted in application in the clinic because of the toxicity.As the further development of the study on Radiotherapy mechanism,it found that the radiation sensitivity is related to internal molecular mechanism of cells in the tumor.Therefore,as a specific role to act on the tumor angiogenesis and signal transduction pathway of molecular in cells of tumor, it become a hotspot and was payed an close attention for the molecular targeted therapy drugs,Relevant researches' results showed it will be the trend of development on radiation sensitizer in the futureMolecular targeted drugs is one of the significant progress in medical treatment of tumor recently.In some tumors the treatment was make a great breakthrough. But the related research reports are little on using molecular targeted drugs combined with radiotherapy.It is reported that the Theory of molecular targeted drug on radiation sensitizing effect is based on the mechanism associated with the following:①To inhibit tumor cell proliferation and radiation damage repair.The damage repair mechanisms of tumor cells are activated by the injury,and the repair mechanisms is related to many key molecular target of signal transduction pathway, molecular targeted drugs can inhibit the activity of these molecular target to sensitize the effect of radiation.②the tumor cell cycle is redistribution.③effect of cytotoxicity and induced apoptosis.Many molecular targeted drugs have a direct cytotoxic effect or apoptosis effect to reduce the number of cloned cells,and will play a space synergies with radiation,then enhancing the sensitivity of radiation.④inhibit tumor angiogenesis.The target is to affect on endothelial growth factor (VEGF).It is reported that the PI3K/Akt signaling pathway in cells the play a very important role for radiosensitivity.A Nigerian-acid Sorafenib Chinese name called Duojimei,is a new multi-target anticancer drugs and a multi-kinase inhibitors(multi-kinase inhibitor),The relatively clear mechanism for the anti-tumor effect,on the one hand is by inhibiting the RAF / MEK/ERK signaling pathway directly to inhibit tumor growth and the other hand is by inhibiting VEGF and platelet-derived growth factor(PDGF) receptors to block the formation of tumor angiogenesis then indirectly inhibit tumor cell growth. Currently sorafenib for the treatment of liver cancer three clinical trials have been completed by the end of 2006.Foreign literature in the sorafenib combined with chemotherapy for kidney cancer,colon cancer treatment has been a small number of reports,but the study about sorafenib combined with radiotherapy for liver cancer at home and abroad are now also not been reported.Methods:1.By MTT experimental to observe the effect and toxicity of sorafenib to inhibit the growth of cells,to learn the value of IC50 and the degree of inhibition by different concentration of drug.And preliminary to observe the survival curves of cell under different concentrations of the drug.2,Cell cloning experiments:By choosing the appropriate concentration of drug and the different doses of radiotherapy combined acting on cells to observe the number of cell Cloning to learn cell survival curve.Through the comparison to the relevant parameters of curve and the sensitization enhancement ratio to observe the drugs on liver cancer cell line HepG2 sensitizing effect of radiation.3,By using PI staining method and FCM to detect distribution of the cell cycle; By using Annexin V tagging Phosphatidylserine valgus,PI staining method and FCM to detect apoptosis of cells.Preliminary to learn the mechanism of sensitizing effect for Sorafenib on liver cancer cell line HepG2.Results:1,MTT Experimental results showed that Sorafenib used alone has significantly inhibited effect on the HepG2 cells,the efficiency of inhibition is related the concentrations of the drug, but also with the role of time.From 3 days to 4days the value of IC50 is 11.5ug/ml and 8.0ug/ml.For the cell survival curves,its effect of inhibition,Likely linear relationship with its concentrations.From the role of time,The longer of action time,the greater of the inhibition effect.2,The cell cloning experiment results:9ug/ml Sorafenib act on the HepG2 cell with radiotherapy,the curve for survival fraction(SF) fit by the Linear quadratic equation.To compare the parameters of the two curve(α,α,α/β)(all P＜0.0001) showed a statistically significant. Compare the SF2,the irradiation group was 0.5125±0.0106,Sorafenib+irradiation group was 0.4607±0.0094,by comparison with two samples mean,P=0.03 show the significant difference;To compare the SER at the level of 2.0 Gy radiation dose,radiation +sorafenih group (SER 1.3478±0.0758) compared to the radiation group(SER 1.11753±0.0055),P=0.017, showed statistical significance(t=- 3.928,n=3).The experimental data show that sorafenib can obvious enhance sensitizing effect of radiation,on liver cancer cell line HepG2.3,Radiate 6.0Gy on HepG2 cells to combine with or without sorafenib,the results showed that sorafenib can lift of the cell cycle block,after using sorafenib the ratio of cells were arrested the decreased significantly in G2 / M phase.Apoptosis experiment showed that addition of sorafenib the radiation group apoptosis rate(14.45±2.72) compare to only drug(apoptosis rate:10.37±2.28) or radiation group(apoptosis rate of 10.06±1.30) the rate of apoptosis increased significantly, analysis of variance(P=0.008) show statistical significance.Conclusion:1,MTT Experimental results showed sorafenib alone act on liver cancer cell lines HepG2 with strong inhibition of cell growth,the inhibit extent and effect is related the concentrations of the drug and continuing role of time,with the increase in drug concentration,the effect enhanced.The effect and concentration of drug similar linear relationship.2,Cloning experiment results showed that Sorafenib can enhance the sensitizing effect of radiation on liver cancer cell line HepG2.Consider with the results of MTT experiment,the enhancement of their role model,is 1+1＞2 and stacking pattern and enhance the effect.3,Experiment of the cell cycle distribution and apoptosis analysis by the FCM found that Sorafenib combined with radiation on cells can lift of the cell cycle block and the rate of apoptosis Increased significantly.Taking into account the pharmacological mechanisms of Sorafenib,it make liver cancer cell line HepG2 radiation enhancement,the mechanism may be related to inhibit radiation-related injury repair mechanism after radiation.Its suppression mechanism of the molecular after radiation damage may be related to inhibit injury of repair and lift of the cell cycle block of G2/M phase.
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