EASL 2018 SAT-365 抗病毒特性和肝脏中特异性递送TLR1 / 2配体 HBV

StephenW

EASL 2018 SAT-365
Antiviral properties and liver specific delivery of a TLR1/2 ligand in
HBV-infected in vitro and in vivo models
J. Lucifora1, F. Fusil2, L. Myriam3, P. Capucine3, M. Maud1, D. Laura1,
A. Ludovic1, B. Maud1, F.-D. Suzanne1, R. Michel4, S. Daffis5,
S. Fletcher5, F. Zoulim1, A. Salvetti1, F.L. Cosset6, V. Bernard7,
D. Durantel1. 1INSERM U1052, Cancer Research Center of Lyon, Lyon,
France; 2Centre International de Recherche en Infectiologie, Lyon, France;
3Institut de Biologie et Chimie des Protéines (IBCP), Lyon, France; 4Centre
Léon Bérard, Lyon, France; 5Gilead Sciences, Foster City, United States;
6Centre International de Recherche en Infectiologie (CIRI), LYON, France;
7Institut de Biologie et Chimie des Protéines (IBCP), Laboratoire de
Biologie Tissulaire et d’Ingénierie Thérapeutique (LBTI), Lyon, France
Email: [email protected]
Background and Aims: Current therapies for hepatitis B virus (HBV)
chronic infections are effective at decreasing viremia in long-life
treatment regimen, but do not lead to viral eradication. Recent studies
highlighted the therapeutic potential of immune-stimulators to
restore immune responses to HBV in various animal models as well
as in early clinical trials. Our overall aim was to explore the anti-HBV
effect of free or particulated TLR1/2 agonists in vitro and in vivo in
monotherapy approaches.
Method: HBV-infected primary human hepatocytes (PHH), HBVinfected
differentiated HepaRG cells (dHepaRG), as well as HBVinfected
liver-humanized (HuHep) mice or AAV-HBV transduced
micewere treated with TLR1/2 agonist. HBV replication and immune
markers/correlateswere followed by ELISA, qPCR, qRT-PCR, Southern
Blot, and IHS.
Results: Pam3CSK4 (TLR1/2-ligand)was amongst the best TLR agonists
tested that reduced viremia and antigenemia in HBV-infected models.
Importantly, its antiviral effect was long-lasting in vitro, as a result of a
strong transcriptional/post-transcriptional inhibition of cccDNA and a
slight, but significant, decrease in its level. This effect was associated
with an activation of the NFkB pathways in hepatocytes and immune
cells, as demonstrated by analyses of mRNA and cytokines production
as well as knock down experiments. To prevent systemic immune
activation and improve its efficacy, Pam3CSK4 was encapsulated in
either “naked or functionalized” nanoparticles that mostly accumulate
in the liver of injected mice. Interestingly, nano-Pam3CSK4 led to a
stronger antiviral activity as compared to free Pam3CK4 in HBV  infected
models, without any toxicological issues.
Conclusion: Our data highlight the potential of innate immunity
stimulators, such as TLR1/2 agonists, as direct antiviral effectors in
hepatocytes and overall modulators of immune responses. This work
further supports the clinical evaluation of TLR agonists as immune
adjuvants in more complex immune-therapeutic strategies, based on
either prime-boost vaccination or T-cell adoption to cure HBV
infections.

StephenW

EASL 2018 SAT-365
抗病毒特性和肝脏中特异性递送TLR1 / 2配体
HBV感染的体外和体内模型
J.Lucifora1，F.Fusil2，L.Myriam3，P.Capucine3，M.Maud1，D.Laura1，
A. Ludovic1，B.Maud1，F.-D. Suzanne1，R.Michel4，S.Daffis5，
S.Fletcher5，F.Zoulim1，A.Salvetti1，F.L. Cosset6，V. Bernard7，
D.杜兰特尔1。里昂癌症研究中心1INSERM U1052，里昂，
法国;法国里昂市国际传染病研究中心;
3法国里昂生物和化学学会（IBCP）; 4Centre
法国里昂的LéonBérard; 5Gilead Sciences，福斯特城，美国;
法国里昂的6Centre International de Recherche en Infectiologie（CIRI）;
7生物和化学学会（IBCP），Laboratoire de
Biologie Tissulaire et d'IngénierieThérapeutique（LBTI），法国里昂
电子邮件：[email protected]
背景和目标：目前治疗乙型肝炎病毒（HBV）
慢性感染可有效减少长期存在的病毒血症
治疗方案，但不会导致病毒根除。最近的研究
强调了免疫刺激剂的治疗潜力
恢复在各种动物模型中对HBV的免疫应答
如在早期临床试验中那样。我们的总体目标是探索抗HBV
免疫或微粒TLR1 / 2激动剂在体外和体内的作用
单一疗法方法。
方法：HBV感染原代人肝细胞（PHH），HBV感染
分化的HepaRG细胞（dHepaRG）以及HBV感染
肝人源化（HuHep）小鼠或AAV-HBV转导
用TLR1 / 2激动剂处理小鼠。 HBV复制和免疫
标记物/相关物随后进行ELISA，qPCR，qRT-PCR，Southern
Blot和IHS。
结果：Pam3CSK4（TLR1 / 2-配体）是最好的TLR激动剂之一
在HBV感染模型中测试了减少的病毒血症和抗原血症。
重要的是，它的抗病毒作用在体外是持久的，因为a
强烈的转录/转录后抑制cccDNA和a
轻微但显着的下降。这种影响是相关的
激活肝细胞和免疫中的NFkB途径
如通过分析mRNA和细胞因子产生所证明的
以及击倒实验。为了防止系统性免疫
激活并提高其功效，将Pam3CSK4包裹在内
无论是“裸体还是功能化”纳米粒子，大部分都是积累的
在注射的小鼠的肝脏中。有趣的是，纳米Pam3CSK4导致了
与感染HBV的游离Pam3CK4相比，抗病毒活性更强
模型，没有任何毒理学问题。
结论：我们的数据突出了先天免疫的潜力
刺激物，如TLR1 / 2激动剂，作为直接抗病毒效应
肝细胞和免疫应答的整体调节剂。这项工作
进一步支持TLR激动剂作为免疫的临床评估
佐剂在更复杂的免疫治疗策略中，基于
不论是主动增强疫苗接种还是采用T细胞治疗HBV
感染。