EASL2015:核酸聚合物对乙肝病毒感染者的抗病毒效果

StephenW

RS-3064
Viral hepatitis
Hepatitis B & D - Experimental

ANTIVIRAL EFFECTS OF NUCLEIC ACID POLYMERS ON HEPATITIS B VIRUS INFECTION
Clément Guillot* 1, Olivier Hantz1, Andrew Vaillant2, Isabelle Chemin1
1Cancer Research Center of Lyon UMR Inserm 1052 CNRS 5286, Lyon, France, 2REPLICor Inc., Montreal, Canada


Corresponding author’s email: [email protected]


Background and Aims:
Hepatitis B virus (HBV) infection remains a major public health problem worldwide and current therapies are rarely able to cure HBV infection. Nucleic acid polymers (NAPs) have been shown to inhibit duck HBV infection in vitro and in vivo (Noordeen et al., 2013). NAPs have been shown to eliminate the hepatitis B surface antigen (HBsAg) from the blood (in ducks infected with the duck hepatitis B virus) but the mechanism whereby NAPs achieve the removal of surface antigen from the blood has yet to be clearly elucidated.  Due to their phosphorothioate oligonucleotide structure, NAPs are chemically analogous to sulfated polyglycans such as heparin which are known to block entry of hepatitis B virus. In this study we investigated the in vitro antiviral activity of NAPs in HBV infected HepaRG cells and primary human hepatocytes.
Methods:
NAP uptake into cells was assessed using Cy3 labeled NAPs. In order to evaluate potent effects of NAPs on HBV entry as well as post-entry, HBV infected differentiated HepaRG cells and primary human hepatocytes were treated with NAPs every two days starting at the time of infection or starting two days post-infection. The Elecsys HBsAg II quant automated system was used to quantitatively measure the secreted HBsAg. PreS1 containing particles were also assessed by ELISA and extracellular HBV DNA was measured by real-time PCR.
Results:
Fluorescent NAPs were observed to enter into differentiated HepaRG cells and in primary cultures more particularly in hepatocytes rather than biliary cells. 5µM and 1µM NAPs significantly reduced extracellular HBsAg by 80% and 40% respectively as well as PreS1 containing particles when added at the time of infection. Interestingly, 5µM NAPs seems also to decrease extracellular HBsAg by 25% as well as PreS1 containing particles when added two days post-infection, suggesting NAPs exert a post-entry antiviral effect.
Conclusions:
In this study, we showed a strong antiviral activity of nucleic acid polymers against HBV infection in HepaRG cells and primary human hepatocytes. Our results suggest that NAPs are able to block entry of HBV but also to have an effect on the replication cycle following entry of the virus which results in a reduction of HBsAg released into the supernatant. These antiviral activities both on virus entry and within the cells promise a strong potential of NAPs alone or in combination with already existing antiviral treatments.


Disclosure of Interest: C. Guillot: : None Declared, O. Hantz: : None Declared, A. Vaillant:  Stockholder: REPLICor,  Employee: REPLICor (VP, CSO), I. Chemin: : None Declared

StephenW

RS-3064

病毒性肝炎

乙肝＆D - 实验





核酸聚合物对乙肝病毒感染者的抗病毒效果

克莱门特GUILLOT * 1，奥利维尔Hantz1，安德鲁Vaillant2，伊莎贝尔Chemin1

里昂INSERM UMR CNRS 1052的1Cancer研究中心5286，法国里昂，2REPLICor公司，加拿大蒙特利尔



通讯作者的邮箱：[email protected]



背景和目的：乙型肝炎病毒（HBV）感染仍然是全球主要的公共健康问题，目前的治疗方法很少能够治愈乙肝病毒感染。核酸聚合物（行动方案）已经显示出抑制鸭HBV感染在体外和体内（Noordeen等人，2013年）。行动方案已经示出，以消除从血液中的B型肝炎表面抗原（HBsAg）（以感染鸭乙型肝炎病毒鸭），但该机制，使行动方案达到去除从血液表面抗原尚未清楚地阐明。由于它们的硫代磷酸酯寡核苷酸的结构，行动方案在化学上类似于硫酸化多聚糖如肝素已知阻断乙肝病毒的条目。在这项研究中，我们调查的行动方案的体外抗病毒活性于HBV感染HepaRG细胞和原代人肝细胞。

方法：NAP摄取到细胞内，用Cy3标记的国家行动方案进行评估。为了评估对HBV条目行动方案有力效果以及入境后，HBV感染HepaRG分化的细胞，并用行动方案处理每两天原代人肝细胞中起始于感染时或开始后两天感染。在Elecsys测定的HBsAg II定量自动化系统用于定量测量分泌的HBsAg。前S1含颗粒也进行了评估，通过ELISA和细胞外HBV DNA通过实时PCR测定。

结果：荧光行动方案，观察进入分化HepaRG细胞和原代培养物更特别地在肝细胞中，而不是胆细胞。 5μM和1μM行动方案由80％和40％分别以及当在感染时加入前S1含颗粒显著降低细胞外的HBsAg。有趣的是，5μM行动方案似乎也减少细胞外的HBsAg 25％，以及当加入2天感染后，提示行动方案施加一个后条目抗病毒作用前S1含有的颗粒。

结论：在本研究中，我们发现在HepaRG细胞和原代人肝细胞对HBV感染核酸聚合物的一个强有力的抗病毒活性。我们的研究结果表明，行动方案是能够阻止HBV的条目，但也有以下这导致减少的HBsAg释放到上清液中的病毒的条目上的复制循环的效果。无论在病毒进入和细胞内的这些抗病毒活性承诺的单独或与现有的抗病毒治疗的组合行动方案一个强大的潜力。



股权变动：C. GUILLOT：无申报，O. Hantz：无申报，A.威能：股东：REPLICor，员工：REPLICor（VP，CSO），一舍曼：无申报