GS9620:针对慢性乙肝的先天免疫联合治疗的发展

StephenW

Targeting Innate Immunity: A New Step in the Development of Combination Therapy for Chronic Hepatitis B

    Fabien Zoulimemail address
    ,
    Souphalone Luangsay
    ,
    David Durantel

INSERM U1052, Hospices Civils de Lyon and Lyon University, Lyon, France



See “GS-9620, an oral agonist of Toll-like receptor-7, induces prolonged suppression of hepatitis B virus in chronically infected chimpanzees,” by Lanford RE, Guerra B, Chavez D, et al, on page 1508.

Current treatments for chronic hepatitis B (HBV) based on nucleos(t)ide analogs (NUC) allow control of viral replication and liver disease in the majority of patients.1, 2 However, because NUCs are unable to clear viral covalently closed circular DNA (cccDNA), lifelong therapy is required to maintain the antiviral effect. To define new treatments with finite duration, it is therefore necessary to develop new molecules acting on novel targets to design true combination therapy complementing already existing NUC-based treatment.3 The persistence of HBV infection and the maintenance of the cccDNA mainly result from a weak HBV-specific immune response. In this respect, the stimulation of the immune response against HBV-infected cells might represent a relevant approach. An efficient control of viral infections requires a concerted action of both innate and adaptive immune responses4 (Figure 1). In the course of viral infections, the host surveillance system represents the first line of antiviral defense and rapidly recognizes viral nucleic acids and proteins. The activation of the innate immunity is essential for triggering an antiviral response of infected cells by producing type I and III interferons (IFNs), by inducing natural killer (NK) cell-mediated killing of viral infected cells, and by inducing the maturation and recruitment of adaptive immunity through production of pro-inflammatory cytokines and chemokines. In a second step, the adaptive immunity takes place after maturation and expansion of B- and T-cell clones that are able to specifically recognize the virus and/or infected cells. Altogether, the coordination of both branches of the immune response can lead to the control of viral infection.4 In this issue of Gastroenterology, Lanford et al5 have investigated the antiviral effect of the activation of the innate immunity via a Toll-Like receptor (TLR)-7 agonist, in HBV chronically infected chimpanzees.

   
    Figure 1.

    The adaptive immune response is shaped by the activation of various cellular innate immune sensors. During chronic hepatitis B (HBV) infection, there is a weak host immune response against HBV-infected hepatocytes. Toll-like receptor (TLR)-7 is mainly expressed in myeloid (mDCs) and plasmacytoid (pDCs) dendritic cells as well as B cells. Its stimulation by agonists may restore a more vigorous immunity against chronic HBV infection. Indeed, TLR-7 activation may lead to (1) the secretion of antiviral cytokines, such as type I IFNs and expression of ISGs, such as IP-10, OAS1, and ISG15, by dendritic cells and/or infected cells; (2) an intercellular cross-talk and cross-activation through the secretion of pro-inflammatory cytokines and chemokines to enhance natural killer (NK) and T-cell recruitment indispensable to shape a persistent adaptive immune response against HBV infection. KC, Kupffer cells; LSEC, liver sinusoidal endothelial cells; mDC, myeloid dendritic cells; pDC, plasmacytoid dendritic cells.

During viral replication, the production of many pathogen associated molecular patterns, such as intermediates of replication, that is, single- or double-strand RNA, viral DNA, core proteins or envelope glycoproteins, may be detected by innate immune sensors, namely, pathogen recognition receptors (PRRs), and trigger an antiviral response. In this respect, PRR activation serves as a link between innate and adaptive immunity. These receptors, mainly represented by the TLRs, the RIG-I like receptors, and the NOD-like receptors, may direct an effective antiviral response through the secretion of type I IFNs and pro-inflammatory cytokines.6 In the course of HBV infection, activation of innate responses may occur in the infected hepatocytes and in cells of the immune system, including circulating dendritic cells and NK cells, liver resident Kupffer cells, as well as other liver nonparenchymal cells.7, 8, 9 The current knowledge from both experimental and clinical studies suggest that HBV is a weak inducer of innate responses,10 but also exhibits a mechanism to suppress innate responses to establish a persistent infection of the liver.4 Indeed, it was shown that HBV may modulate innate responses by interfering either with TLR signaling pathways11 the type I IFN signaling pathway12, 13 or the cross-talk between plasmacytoid dendritic cells (pDC) and NK cells.14

TLR-7 is a PRR mainly expressed in endosomal compartments of pDCs and B lymphocytes that recognizes a pathogen-associated molecular pattern in viral single-stranded RNA. Upon stimulation of TLR-7, pDCs produce IFN-α and other cytokines/chemokines and induce the activation of NK cells and activation of cytotoxic lymphocytes, thereby orchestrating both innate and adaptive immune responses6 (Figure 1). The altered responsiveness of pDCs might contribute to the reduced innate immune responses during chronic viral infections. Agonist-induced activation of TLR-7 might therefore represent a novel approach for the treatment of chronic viral infections.15

Lanford et al5 have investigated the effects of immune activation with GS-9620, an orally administered agonist of TLR-7, in chimpanzees chronically infected with HBV. GS-9620 was administered to chimpanzees every other day for 4 weeks at 1 mg/kg and, after a 1-week rest, for a second cycle of 4 weeks at 2 mg/kg. A detailed evaluation was performed, including pharmacokinetics of GS-9620, viral load, IFN-stimulated gene (ISG) expression, cytokine and chemokine levels, and lymphocyte and NK cell activation, as well as safety and tolerability parameters. Short-term administration of the TLR-7 agonist provided long-term suppression of serum and liver HBV DNA. The mean maximum reduction of viral DNA was 2.2 logs and occurred within 1 week of the end of therapy; reductions of >1 log persisted for several months. Serum levels of hepatitis B surface antigen and hepatitis B e antigen, and numbers of HBV antigen-positive hepatocytes, were reduced as hepatocyte apoptosis increased. In parallel, GS-9620 administration induced the production of IFN-α and other cytokines and chemokines, up-regulated ISGs expression, and activated NK cells and lymphocyte subsets, confirming the activation of TLR-7 signaling.

This is a timely study; new antiviral targets are required to improve the rate of hepatitis B surface antigen loss in patients with chronic hepatitis B. Theoretically, the TLR-7 agonist-induced antiviral activity may complement the inhibition of viral DNA synthesis induced by NUCs.3 From the results presented in this work, one might envision several mechanisms by which the TLR-7 agonist administration might have led to the observed antiviral effect5 (Figure 1). The decrease of both intrahepatic and serum viral load as well as the number of hepatitis B core antigen-positive hepatocytes was concomitant with an increase in liver enzymes and immune cell infiltration, suggesting HBV-infected cell killing by specific CD8+ T-cell responses and NK cell responses. This seems to be consistent with the role of infected cell killing and hepatocyte turnover in the subsequent clearance of infection.16 The determination of the respective role of hepatocyte apoptosis and/or lysis will also be important to fully understand the mechanism involved in the viral clearance process induced by this TLR-7 agonist. The induction of type-I IFN and ISGs expression may also have contributed to enhance the initial antiviral effect, similarly to the T helper 1 cytokine expression observed previously in experiments performed with HBV transgenic mice.17 However, it is unlikely involved in the prolonged effect observed in these chimpanzees, because the induction of gene expression was only transient. It would also have been interesting to investigate the effect of this treatment regimen on the evolution of cccDNA amounts and its epigenetic control; it was previously shown in liver-humanized mice infected with HBV that IFN-α treatment induced the silencing of cccDNA by epigenetic reprogrammation18 and that immune-mediated killing of infected hepatocytes would lead to cccDNA decay.16 The determination of both integrated viral DNA and cccDNA would also have been extremely useful to ascribe the eventual cccDNA decay to the loss of infected cells or to infected cell curing.16 It will be also important to determine whether the positive effects observed in these infected chimpanzees receiving TLR-7 agonist monotherapy is confirmed, or even enhanced, in the context of combination therapy in chronically infected animals/patients who achieve viral DNA suppression in serum during NUC-based therapy.

One concern with TLR-7 agonist administration is the potential induction of adverse events, as previously observed with several other agonists evaluated in clinical trials for the treatment of chronic hepatitis C.19, 20 Flu-like syndromes and lymphopenia were observed in these clinical trials for hepatitis C.19, 20 The use of GS-9620 in HBV-infected chimpanzees was associated with lymphopenia as well as liver enzyme elevations; it is, however, interesting that there was no systemic production of IFN-α.5 Other potential adverse events resulting from an unregulated activation of TLR may include chronic inflammatory diseases and/or auto-immunity.6 Ongoing clinical trials will determine whether the administration of this specific TLR-7 agonist will provide the expected antiviral effect without the systemic effect associated with cytokine induction. A better knowledge of the precise mechanism involved in HBV-induced repression of the infected host innate immunity might also help to design novel approaches to specifically trigger an antiviral innate response restricted to infected cells, with the caveat that the adaptive immune responses may not be involved in such a strategies.

In conclusion, the results presented by Lanford et al5 describing a novel approach to inhibit HBV replication and induce infected cell clearance provide refreshing perspectives on how to tackle chronic HBV infection (Figure 1). It represents a step toward the development of true combination therapy in chronic hepatitis B; with the aim of increasing the rate of serum HBsAg clearance.

StephenW

StephenW

目前的治疗慢性乙肝病毒（HBV）的基础上核苷（酸）类似物（NUC）允许控制病毒复制和肝脏疾病的广大患者，[2]然而，因为NUCs无法清除病毒共价闭合环状DNA （cccDNA的），需要终身治疗，以维持抗病毒作用。要定义新的治疗持续时间有限，因此，有必要制定新的目标，新的分子作用于设计真正的联合治疗，补充已有的基于NUC治疗的持久性HBV感染和维护的cccDNA主要由弱HBV特异性免疫反应。在这方面，对HBV感染细胞的刺激的免疫反应可能代表一个相关的方法。一个有效的控制病毒感染，先天免疫和适应性免疫responses4需要采取协调一致的行动，（图1）。在病毒感染的过程中，监视主机的系统代表抗病毒防御的第一行，并迅速识别病毒的核酸和蛋白质。生产Ⅰ，Ⅲ型干扰素（干扰素）感染的细胞的抗病毒反应的触发，通过诱导自然杀伤（NK）细胞介导的​​杀死病毒感染的细胞，并通过诱导成熟和招募的先天免疫系统的活化是必要的通过促炎性细胞因子和趋化因子的生产适应性免疫。在第二步骤中，发生适应性免疫后的B-和T-细胞克隆能够特异性识别的病毒和/或感染的细胞的成熟和扩展。总而言之，协调两个分支的免疫反应可导致病毒感染的控制，在这个问题上消化，兰福德等[5]研究了抗病毒效果的先天免疫激活Toll样受体（TLR通过）-7受体激动剂，在HBV慢性感染的黑猩猩。

  
图1。

    适应性免疫应答的形状由各种细胞的先天免疫传感器的激活。在慢性乙型肝炎（HBV）感染，是一种弱的对HBV感染的肝细胞的宿主免疫反应。 Toll样受体（TLR）-7主要表达在骨髓（MDC的）和浆细胞（pDCs细胞）的树突状细胞和B细胞。其激动剂刺激可以恢复更有力的抗慢性乙肝病毒感染。事实上，TLR-7的激活可能导致（1）的抗病毒细胞因子的分泌，如I型干扰素和表达的ISGs，如IP-10，OAS1，ISG15，树突状细胞和/或感染的细胞，（2 ）通过分泌促炎症细胞因子和趋化因子，以提高自然杀伤细胞（NK）和T-细胞不可或缺的招聘塑造一个持久的适应性免疫应答对乙肝病毒感染的细胞间的串扰和交叉激活。 KC，枯否细胞，肝窦内皮细胞，肝窦内皮细胞; MDC，髓样树突状细胞; PDC，浆细胞样树突状细胞。

病毒复制过程中，生产的许多病原体相关的分子模式，如复制的中间体，即单或双链RNA，病毒DNA，核心蛋白，包膜糖蛋白，可能会被检测到由先天免疫传感器，即，病原体模式识别受体（PRRS），并触发抗病毒反应。在这方面，PRR激活作为先天和适应性免疫之间的链接。这些受体，主要表示的Toll样受体，RIG-I相同的受体，NOD样受体，可以直接通过在HBV感染的过程中分泌的I型干扰素和亲炎症cytokines.6的有效的抗病毒反应，先天反应的激活可能发生在感染的肝细胞和免疫系统的细胞，包括从两个试验循环树突状细胞和NK细胞，Kupffer细胞肝驻地，以及其他肝脏非实质细胞的，8，9由当前的知识和临床研究表明，乙肝病毒是一种本能反应，10的微弱诱导剂，但也表现出一种机制来抑制本能反应，建立持续性感染的肝脏的确，它表明，乙肝病毒可调节先天反应干扰与TLR信号pathways11的I型IFN信号pathway12，13或浆细胞样树突状细胞（PDC）的之间的串扰和NK细胞。

TLR-7是的PRR主要表达的核内体的隔室的pDCs和B淋巴细胞识别病毒单链RNA的病原体相关的分子模式。 TLR-7刺激后，pDCs细胞产生的IFN-α和其他细胞因子/趋化因子诱导活化的NK细胞和细胞毒性T细胞的激活，从而编排先天和适应性：免疫responses6（图1）。 pDCs细胞的反应性改变，可能有助于减少先天免疫反应在慢性病毒感染。激动剂诱导的激活TLR-7，因此可能代表一种新的方法，用于治疗慢性病毒infections.15

兰福德等[5]研究免疫激活激动剂口服TLR-7，GS-9620，在慢性HBV感染的黑猩猩的影响。 GS-9620管理黑猩猩隔日1毫克/公斤，连续4周，休息1周后，在2毫克/千克为第二个周期为4周。进行的详细评估，包括GS-9620的药代动力学，病毒载量，IFN-α刺激基因（ISG）的表达，细胞因子和趋化因子水平，淋巴细胞和NK细胞的激活，以及安全性和耐受性的参数。短期管理的TLR-7受体激动剂提供长期抑制血清和肝脏中的乙肝病毒DNA。平均最高减少病毒DNA为2.2日志和治疗结束后1星期内发生减少> 1日志坚持了好几个月。血清乙肝表面抗原和乙肝e抗原，HBV抗原阳性肝细胞的数目，减少肝细胞凋亡增加。与此同时，GS-9620给药诱导IFN-α和其他细胞因子和趋化因子，上调ISGs的表达，活化的NK细胞和淋巴细胞亚群的生产，确认激活TLR-7信令。

这是一个及时研究新的抗病毒的目标要求的提高从理论上讲慢性乙型肝炎患者的B型肝炎表面抗原的损失率，TLR-7激动剂诱导的抗病毒活性，让病毒DNA合成的抑制诱导NUCs 0.3从在这项工作中呈现的结果，人们可能会设想一些机制，TLR-7受体激动剂给药有可能导致所观察到的抗病毒effect5（图1）。的减少两个肝内和血清病毒负荷，以及数乙肝核心抗原阳性肝细胞是伴随肝酶和免疫细胞浸润增加，提示由特定的CD8 + T细胞的反应和NK细胞的HBV感染细胞杀害细胞反应。这似乎是符合感染细胞的杀伤作用和肝细胞成交金额在随后的间隙感染的肝细胞凋亡和/或溶解的确定各自的作用也将是重要的是要充分理解这一机制参与病毒清除处理这TLR-7受体激动剂诱发。提高初始抗病毒效果，类似于先前在与HBV转基因mice.17的实验中观察到的T辅助细胞1细胞因子的表达，但是，它是不太可能参与的长期效果，也可能导致I型IFN的诱导和ISGs表达在这些黑猩猩中观察到，，因为诱导的基因表达仅是短暂的。这也将是有趣的调查的演变的cccDNA的金额和其后生控制这种治疗的效果，它是先前的研究显示，感染HBV的肝人性化小鼠诱导IFN-α治疗的沉默cccDNA的后生reprogrammation18和感染的肝细胞，免疫介导的杀死会导致cccDNA的decay.16也将是极其有用​​的测定两个整合的病毒DNA与cccDNA归于最终的共价闭合环状DNA感染的细胞的损失或衰减到感染细胞curing.16这也将是重要的，以确定是否观察到的这些受感染的黑猩猩接收TLR-7受体激动剂单药治疗的积极效应被确认，或什至增强，在​​联合治疗的背景下，在慢性感染的动物/实现的患者血清中病毒DNA的抑制期间NUC为基础的治疗。

TLR-7受体激动剂给药的一个关注的是的潜在诱导的不良事件，和以前观察到与其他几个激动剂评估在临床试验中用于治疗慢性丙型肝炎C.19，20流感样综合征和淋巴细胞减少在这些临床试验中观察到的肝炎C.19，20在HBV感染的黑猩猩的使用GS-9620，淋巴细胞减少症，以及肝酶升高相关联，它是，但是，有趣的是，有没有全身性生产的IFN-α.5其他的潜在不利从不稳定的事件导致激活TLR可能包括慢性炎症性疾病和/或自动immunity.6正在进行的临床试验将决定是否将这个特定的TLR-7受体激动剂的管理提供预期的抗病毒效果，而无需与细胞因子诱导的全身效应。一个更好的了解HBV感染的宿主先天免疫诱导镇压的精确机制也可能有助于设计新的方法来触发仅限于受感染的细胞抗病毒先天反应，需要提醒的是适应性免疫反应可能不参与在这样的策略。

总之，结果兰福德等[5]描述了一种新的方法来抑制HBV的复制，并引起感​​染的细胞间隙提供清凉的观点，就如何解决慢性HBV感染者（图1）。它代表了一个步骤，向真正的联合治疗慢性乙型肝炎的发展，增加血清HBsAg清除率的目的。