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本帖最后由 StephenW 于 2015-3-7 18:56 编辑
回复 战天斗hbv 的帖子
当然，结果令人失望，相比黑猩猩的结果. 但是，我们不是专家, 不能说GS9620是失败的.
http://www.natap.org/2014/AASLD/AASLD_139.htm
Cytokines Induced by a Toll-Like Receptor 7 Agonist Potently Inhibit HBV RNA, DNA, and Antigen Levels in Primary Human Hepatocytes
Reported by Jules Levin
AASLD 2014 Nov 7-11 Boston
Congrong Niu, Stephane Daffis, Mei Yu, Guofeng Cheng, William E. Delaney IV, Simon P. Fletcher
Gilead Sciences, Inc., Foster City, CA
Background & Aims: GS-9620, an oral agonist of toll-like receptor 7 (TLR7), is in phase 2 trials for the treatment of chronic hepatitis B (CHB). Infrequent dosing of GS-9620 (e.g. once a week) induced prolonged suppression of serum viral DNA and antigens in animal models of CHB. Here we investigated the molecular mechanisms that contribute to the antiviral response to GS-9620 by evaluating the antiviral activity of TLR7 agonists in vitro.
Methods: Primary human hepatocytes (PHH) were infected with HBV and ≥3 days later were treated with a TLR7 agonist (compound A), with media from human PBMCs treated with the TLR7 agonist (TLR7 conditioned media; TLR7-CM) or with recombinant cytokines. Antiviral activity was evaluated by quantifying extracellular HBV DNA (qPCR), HBeAg and HBsAg (ELISA), as well as intracellular HBV RNA (qRT-PCR).
Results: The TLR7 agonist compound A (a close analog of GS-9620), had no direct antiviral activity in HBV-infected PHH, consistent with the lack of functional TLR7 in hepatocytes. In contrast, sustained exposure of HBV-infected PHH to TLR7-CM strongly reduced the levels of HBV DNA and HBeAg (>90%), as well as HBsAg and HBV RNA (>75%), without detectable toxicity. Reductions in viral parameters were observed when PHH were treated with TLR7-CM early (3 days) or late (13 days) post-infection, but were not induced by media from control DMSO-treated PBMCs. We next compared the durability of short (3 day) and long (10 day) treatment of HBV-infected PHH with TLR7-CM. Short duration treatment (days 3-6 post-infection) only transiently reduced HBeAg. In contrast, prolonged treatment (days 3-13 post-infection) induced more sustained suppression of HBeAg, with rebound still incomplete 7 days after removal of TLR7-CM (day 20). A survey of cytokines in TLR7-CM identified recombinant IFN-α2a as a potent antiviral cytokine (EC50 ≤ 10 IU/mL for HBV DNA and HBeAg) in HBV-infected PHH. Recombinant TNF-α, IFN-γ and IFN-λ1 also strongly reduced HBV DNA, RNA and antigen levels, whereas IL-6 had only weak antiviral activity. Since TLR7 agonists induced substantially more IFN-α and IL-6 in human PBMCs than other cytokines, these data indicated that IFN-α was likely the principal mediator of TLR7-CM antiviral activity in HBV-infected PHH.
Conclusion: Sustained exposure to antiviral cytokines directly induced by TLR7 activation, such as IFN-α, potently inhibited HBV in PHH in vitro. However, since short duration exposure had only a transient antiviral effect, additional components of the TLR7-induced immune response may also play an important role in the antiviral response to GS-9620 in vivo.
细胞因子Toll样受体激动剂7强效抑制HBV RNA，DNA诱导和抗原水平的人原代肝
儒勒·莱文报道
AASLD 2014年11月7日至11日波士顿
从戎牛，斯特凡Daffis，梅宇，郑华国锋，威廉E.德莱尼IV，西蒙·P·弗莱彻
吉利德科学公司，福斯特市，CA
背景与目的：GS-9620，toll样受体7（TLR7）的口服激动剂，是在阶段2试验为慢性乙型肝炎（CHB）的治疗。 GS-9620的罕见剂量（如每周一次）诱导血清病毒DNA和慢性乙型肝炎的动物模型抗原长期抑制。在这里，我们调查了有助于通过评估TLR7激动剂的抗病毒活性在体外对GS-9620的抗病毒反应的分子机制。
方法：原代人肝细胞（PHH）感染了乙肝病毒和≥3天之后进行与TLR7激动剂（化合物A）的处理，用从与TLR7激动剂（TLR7条件培养基; TLR7-CM）的处理的人PBMC中的介质或用重组细胞因子。抗病毒活性通过定量细胞外HBV DNA（定量PCR），HBeAg和HBsAg的（ELISA），以及细胞内的HBV的RNA（QRT-PCR）来评价。
结果：TLR7激动剂化合物A（GS-9620的亲密模拟），并没有直接的抗病毒活性的HBV感染PHH，在肝细胞缺乏TLR7功能一致。与此相反，持续HBV感染PHH暴露于TLR7-CM大大减少HBV DNA和HBeAg的（> 90％）的水平，以及HBsAg和乙肝病毒的RNA（> 75％），没有可检测的毒性。在病毒参数的减少观察时PHH分别与TLR7-CM早期（3天）或晚期（13天）在感染后治疗，但没有引起从控制DMSO处理的PBMC介质。我们接下来比较短（第3天）和长（10天）治疗HBV感染PHH与TLR7-CM的耐久性。短时间的治疗（3-6天感染后）只有短暂减少大三阳。相比之下，引起的HBeAg更持续抑制，随着反弹的长期治疗（3-13天感染后）仍然是不完整切除TLR7-CM后7天（20天）。细胞因子TLR7-CM的调查鉴定重组IFN-α2A作为一种有效的抗病毒细胞因子（EC50≤10 IU /毫升HBV DNA和HBeAg），乙肝病毒感染PHH。重组的TNF-α，IFN-γ和IFN-λ1还强烈降低的HBV DNA，RNA和抗原水平，而IL-6只有弱的抗病毒活性。自的TLR7激动剂诱导的基本上更多的IFN-α和IL-6在人PBMC比其它细胞因子，这些数据表明，IFN-α是可能的TLR7-CM的抗病毒活性在HBV感染PHH的主要介体。
结论：持续暴露于直接诱导TLR7活化的抗病毒细胞因子如IFN-α，有效抑制乙肝病毒在PHH体外。然而，由于时间短曝光只有一个短暂的抗病毒效果，TLR7诱导的免疫应答的附加组分也起到体内GS-9620的抗病毒反应中起重要作用。 |
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楼主: newchinabok
发表于 2015-3-7 17:16
