heg类似物中断：用于HBV自发复制控制的肝内设定点的替代方he

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Nucleos(t)ide analogue interruption: Alternative approach to intrahepatic set point for spontaneous control of HBV replication?

Yiqi Yu†, Jing Wang†, Guojun Li, Chuan Shen, Shaolong Chen, Jun Huang, Xinyu Wang, Caiyan Zhao, Yuxian Huang, Bin Wang, Xuanyi Wang, Jiming Zhang'Correspondence information about the author Jiming ZhangEmail the author Jiming Zhang, Chao Qiu'Correspondence information about the author Chao QiuEmail the author Chao Qiu, Wenhong Zhang'Correspondence information about the author Wenhong ZhangEmail the author Wenhong Zhang
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DOI: http://dx.doi.org/10.1016/j.jhep.2017.09.026
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With great interest, we read the manuscript by Berg et al. in the Journal of Hepatology.[url=]1[/url] Although prolonged antiviral treatment reduced the levels of intrahepatic viral replicative forms,[url=]2[/url] during the initial course of tenofovir disoproxil fumarate (TDF) off-therapy, viral rebound developed in all patients who had achieved suppression of serum HBV DNA to an undetectable level for at least 3.5 years. This finding raises an unresolved question regarding the optimal virological set point for the spontaneous control of HBV replication. Herein, we evaluated whether such a set point could be accomplished through long-term nucleos(t)ide analogue (NA) therapy.

Durable viral suppression was observed in hepatitis B e antigen (HBeAg)-negative patients with chronic hepatitis B (CHB) and low or undetectable serum HBV DNA and normal aminotransferase levels, which was previously referred to as the “inactive carrier” phase.[url=]3[/url] Recapitulation of the inactive carrier state is considered an acceptable endpoint for cessation of therapy. Therefore, we compared the levels of the viral replicative forms of HBV in serum and liver samples between HBeAg-negative patients who had been successfully treated with entecavir (ETV) for >1 year and inactive carriers. Treatment-naive HBeAg-negative chronic patients experiencing active hepatitis were also included for comparison.

As shown in Fig. 1A, significantly lower levels of serum HBV RNA were observed in ETV-treated HBeAg-negative patients (2.57 log10 copies/ml, range = 2.00–3.73 log10 copies/ml) than in treatment-naive patients (4.74 log10 copies/ml, range = 2.76–5.75 log10 copies/ml; p <0.0001). In line with recent studies,[url=]4[/url] serum HBV RNA (<200 copies/ml) was undetectable in 6/22 (27.3%) ETV-treated patients. However, cases with undetectable serum HBV RNA were more frequent (12/16, 75%) among inactive carriers (Fig. 1A, p = 0.0076).

Fig. 1

Levels of viral replicative forms and spontaneous control of HBV replication. (A) Levels of serum HBV-RNA, (B) intrahepatic HBV-RNA, (C) intrahepatic total HBV DNA, (D) and cccDNA under ETV treatment in comparison to those of IA and IC patients. Level of cccDNA in one ETV-treated case, one IC case and one IA case were below the lower detectable limit. ETV, HBeAg-negative patients receiving ETV treatment; IA, HBeAg-negative patients in the immune-active phase who were naive to treatment, also known as HBeAg-negative chronic hepatitis B; IC, inactive carrier, also known as HBeAg-negative HBV infection (Mann–Whitney test, *p <0.05; **p <0.01; ***p <0.001). cccDNA, covalently closed circular DNA; ETV, entecavir; HBeAg, hepatits B e-antigen; HBV, hepatitis B virus.

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In liver biopsies, intrahepatic HBV RNA was detectable for all patients. The intrahepatic HBV RNA levels of patients treated with ETV (2.76 log10 copies/ml, range = 0.30–4.40 log10 copies/ml) were not significantly lower (p = 0.2786) than those of treatment-naive patients (2.06 log10 copies/ml, range = 1.09–3.10 log10 copies/ml), and remained higher than those of inactive carriers (1.21 log10 copies/ml, range = 0–1.87 log10 copies/ml; p = 0.0019) (Fig. 1B). In comparison to treatment-naive patients, levels of intrahepatic total HBV DNA (Fig. 1C) and covalently closed circular DNA (cccDNA) (Fig. 1D) were significantly decreased in ETV-treated patients and inactive carriers, but no differences were observed between the latter two groups. Therefore, our data suggested that although NA therapy induced the remission of liver disease and reduced the size of the cccDNA pool, the levels of intrahepatic HBV RNA could not be reduced to the comparably low levels of inactive carriers. This finding explains why patients under prolonged treatment remained at higher risk of HBV reactivation than patients with spontaneous HBeAg seroconversion.[url=]5[/url] Lower levels of intrahepatic HBV RNA approaching those of inactive carriers, might also be considered a useful virological endpoint for cessation of NA therapy.

Berg et al.’s study also disclosed some encouraging discoveries, inspiring further investigations. During the 144-week follow-up period, four patients who discontinued NA therapy experienced hepatitis B surface antigen (HBsAg) loss and three of them achieved HBsAg seroconversion, whereas no such change in HBsAg was detected in the patients that continued therapy. In the context of CHB, the T cell response to HBV is generally weak and dysfunctional, and could be partially restored by long-term NA therapy.[url=]6[/url] Moreover, most HBV-specific T cells are sequestered in the liver and do not circulate through the peripheral tissues.[url=]7[/url] Thus, HBV rebound after withdrawal of long-term NA therapy might directly activate liver-resident HBV-specific T cells,[url=]8[/url] which would more effectively restrict viral replication than the immune responses elicited by traditional HBV therapeutic vaccines inoculated peripherally.[url=]7[/url] We propose that “NA interruption” might serve as a novel therapeutic approach for directly boosting liver-resident immune responses against HBV and further suppressing viral activity toward an intrahepatic set point for achieving spontaneous control. However, further studies are needed to define the mechanisms of NA interruption required for realizing spontaneous control of HBV replication and to identify patients prone to clinical relapse after NA interruption using HBsAg quantification or other predicators.[url=][9][/url], [url=][10][/url]

Lastly, NA interruption as a therapeutic approach is associated with several limitations. Firstly, the durability of viral control that might be achieved by this approach is currently uncertain. Secondly, viral replication in the presence of a low drug concentration could potentially lead to the development of viral variants with drug resistance. Therefore, NA agents with a high genetic barrier are highly recommended for this approach. Thirdly, NA interruption as a therapeutic approach should not be recommended for patients with cirrhosis, because of the risk of liver failure due to viral reactivation. Close follow-up would be required to prevent uncontrolled conditions such as recurrent viremia, alanine transaminase flares, and clinical decompensation.

In conclusion, levels of intrahepatic HBV RNA might be a potential indicator and virological set point for withdrawal of NA therapy.

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Nucleos（t）ide类似物中断：用于HBV自发复制控制的肝内设定点的替代方法？


致编辑：
感兴趣的是，我们阅读了Berg等人的手稿。尽管延长的抗病毒治疗降低了肝炎病毒复制形式的水平[2]，但是在替诺福韦二吡呋酯富马酸盐（TDF）治疗的初始过程中，所有已经达到血清HBV DNA抑制的患者都出现了病毒反弹至少要3.5年才能检测出来。这一发现提出了关于HBV复制的自发控制的最佳病毒学设定点的尚未解决的问题。在此，我们评估了这种设定点是否可以通过长期的核苷（酸）类似物（NA）疗法来完成。

在乙型肝炎e抗原（HBeAg）阴性的慢性乙型肝炎患者（CHB）和低或不可检测的血清HBV DNA和正常转氨酶水平，以前被称为“非活动载体”阶段，观察到持久的病毒抑制作用.3重新评估的无活性载体状态被认为是用于停止治疗的可接受终点。因此，我们比较了恩替卡韦（ETV）治疗1年以上的HBeAg阴性患者和非活动性携带者血清和肝脏中HBV的病毒复制形式水平。未接受治疗的HBeAg阴性慢性活动性肝炎患者也被纳入比较。

如图1A所示，ETV治疗的HBeAg阴性患者（2.57log10copies / ml，范围= 2.00-3.73log10copies / ml）比未治疗的患者（4.74log10）显着降低了血清HBV RNA水平拷贝/ ml，范围= 2.76-5.75log10copies / ml; p <0.0001）。根据最近的研究，在6/22（27.3％）ETV治疗的患者中检测不到4种血清HBV RNA（<200copies / ml）。然而，血清HBV RNA检测不到的病例在非活动性携带者中更频繁（12/16，75％）（图1A，p = 0.0076）。

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图。1
病毒复制形式的水平和HBV复制的自发控制。 （A）在ETV治疗下血清HBV-RNA，（B）肝内HBV-RNA，（C）肝内总HBV DNA，（D）和cccDNA的水平与IA和IC患者相比较。一个ETV治疗病例，一个IC病例和一个IA病例的cccDNA水平低于可检测下限。 ETV，接受ETV治疗的HBeAg阴性患者; IA，HBeAg阴性患者在免疫活跃期谁被天真去治疗，又称为HBeAg阴性慢性乙型肝炎; IC，无活性的载体，也称为HBeAg阴性HBV感染（Mann-Whitney检验，* p <0.05; ** p <0.01; *** p <0.001）。 cccDNA，共价闭合环状DNA; ETV，恩替卡韦; HBeAg，乙肝e抗原; HBV，乙型肝炎病毒。

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在肝脏活组织检查中，所有患者均可检测到肝内HBV RNA。 ETV治疗组患者肝内HBV RNA水平（2.76log10copies / ml，范围= 0.30-4.40log10copies / ml）与治疗前未治疗患者（2.06log10copies / ml）差异无显着性（p = 0.2786） （1.21log10copies / ml，范围= 0-1.87log10copies / ml; p = 0.0019）（图1B）。与未接受治疗的患者相比，接受ETV治疗的患者和非活动性携带者的肝内总HBV DNA水平（图1C）和共价闭合环状DNA（cccDNA）水平（图1D）均显着降低，但未观察到差异后两组。因此，我们的数据表明，虽然NA治疗诱导了肝病的缓解，并减小了cccDNA库的大小，但是肝内HBV RNA水平不能降低到相对较低水平的非活动性携带者。这一发现解释了为什么接受长时间治疗的患者与自发性HBeAg血清学转换的患者相比，HBV再激活的风险仍然较高[5]。接近那些无活性的携带者的较低水平的肝内HBV RNA也可能被认为是停止NA治疗的有用的病毒学终点。

伯格等人的研究也披露了一些令人鼓舞的发现，激发了进一步的调查。在144周的随访期间，4名停止NA治疗的患者经历了乙型肝炎表面抗原（HBsAg）丧失，其中3人获得了HBsAg血清学转换，而在继续治疗的患者中没有检测到HBsAg的这种变化。在慢性乙型肝炎背景下，T细胞对HBV的反应一般较弱且功能失调，并且可以通过长期的NA疗法部分恢复。而且，大多数HBV特异性T细胞被隔离在肝脏中，并且不通过因此，停止长期NA治疗后HBV的反弹可能直接激活肝脏HBV特异性T细胞[8]，比传统的HBV治疗疫苗外周接种引起的免疫反应更有效地限制病毒的复制[7]。我们提出“NA中断”可以作为一种新的治疗方法，用于直接提高肝脏对HBV的免疫应答，并进一步抑制病毒活性达到肝内设定点以实现自发控制。然而，还需要进一步的研究来确定实现HBV复制的自发控制所需的NA中断的机制，并使用HBsAg定量或其他预测因子来鉴定在NA中断后容易发生临床复发的患者[9,10]

最后，作为治疗方法的NA中断与几个限制相关联。首先，通过这种方法可能实现的病毒控制的持久性目前是不确定的。其次，在药物浓度低的情况下病毒复制可能导致具有耐药性的病毒变体的发展。因此，强烈推荐具有高耐药屏障的NA药物。第三，NA中断作为一种治疗方法不应该被推荐用于肝硬化患者，因为病毒再激活导致肝衰竭的风险。需要密切随访，以防止不受控制的情况，如复发性病毒血症，丙氨酸转氨酶闪光和临床代偿失调。

总之，肝内HBV RNA水平可能是NA治疗撤退的潜在指标和病毒学设定点。