The Anti-HBV Effec Of The Compound Of DC9

Backgrounds and ObjectiveAt present,Hepatitis B is a serious threat to human health worldwide especially in China.Statistically,there are more than 257 million people with hepatitis B virus S antigen positive in the world,which leads to about 887,000 deaths per year,following development of liver disease,such as cirrhosis,liver cancer or other complications from the WHO.Current options to treat chronic HBV infection are resitricted to the use of interferon,nucleoside,thymosin,but these treatments have some drawbacks,including serious side effects,drug resistance associated with HBV and expensive drug price.In addition,although the combination of the chemical drugs in the antiviral therapy more effectively reduce morbidity and mortality in HBV infected individuals than the use of sigle drug,emergence of multidrug resistance and side effects for long-term antiviral treatment limited the use of these drugs.Therefore,it is an urgent task for the medicine practitioners to look for new,safe and effective anti-HBV drugs.Natural products as the most consistently successful soure in drug discovery may offer opportunities for finding anti-HBV druga or lead compounds.Dibenzo cyclooctadiene compounds are extracted from the Schisandra plant,and most of them have biological activity,such as antitussive,anti-HIV,anti-HBV and other activities.Ten compounds of DC series is from the structural transformation of the non-nucleoside dibenzo cyclooctadiene,in order to play its anti-HBV effect for the clinical treatment of chronic hepatitis B.This thesis mainly aims to choose anti-HBV activity medication from DC series,then presents research on anti-HBV mechanism,for providing theory foundation for clinical therapy.Materials and MethodsThe human HBV-transfected cell line HepG2.2.15 which stably express HBV was emploied to evaluate anti-HBV effect of ten compounds of DC series in vitro.Selected the compounds which have anti-HBV activity for the further experiment.Detected the changes of cell supernatant and intracellular HBV DNA and studied on the mechanism.The contents were as follows:The 10 compounds were evaluated for cell toxicity by MTT method.The HBsAg and HBeAg level of cells supernatant collected at day 6 were detected by ELISA method.The Median Toxic Concentration(TC₅₀),50%Inhibiting Concentration(IC₅₀),and Therapeutic Index?TI?as indexes to evaluate the anti-HBV activity of the 10 compounds.The cell supernatant and intracellular HBV DNA at day 3,6,9 were detected by fluorescence quantitative polymerase chain reation?FQ-PCR?,the inhibition rate of cell supernatant and intracellular HBV DNA of DC9 were obtained.And expression of mRNA of p gene of HBV was analyzed by reverse thanscriptase PCR?RT-PCR?.Results1:10 compounds screening were completed.TI values of DC9 were greater than 1.2:DC9 revealed cytotoxity on HepG2.2.15 cell,and the TC₅₀ value was greater than 400?M;and the IC₅₀ values of the secretion of HBsAg and HBeAg were54.49 and 28.17?M,the TI values were greater than 7.34 and greater than 14.2,respectively.DC9 also reduced cell supernatant and intracellular HBV DNA copies in a dose-and time-dependent manner.The result of RT-PCR show that high concentration of DC9 can cause the inhibition of the expression of mRNA of p gene of HBV DNA at the transcription level,compared with the control group?P<0.01?,and the inhibitory effect was stronger than 3TC.Conclusions1:Among 10 compounds,the compound of DC9 have strong anti-HBV activity.2:DC 9 is effective in inhibiting HBV in vitro,and not only has better inhibition of secretion of HBsAg and HBeAg,but also could reduce HBV DNA level in cell supernatant and intracellular significantly.Furthermore,DC9 can inhibite the expression of mRNA of p gene of HBV.