[AASLD 2012]HAPS抑制HBV核衣壳的成熟与cccDNA转录

肝胆速递

Heteroaryldihydropyrimidines（HAPS）抑制HBV核衣壳的成熟与cccDNA转录
CONTROL ID: 1426205
PRESENTATION TYPE: Oral or Poster
CURRENT CATEGORY: Hepatitis B
CURRENT DESCRIPTORS: I02. Treatment and Clinical Trials
TITLE: Heteroaryldihydropyrimidines (HAPs) inhibit HBV replication in tissue culture by targeting both nucleocapsid maturation and cccDNA transcription
AUTHORS (FIRST NAME, LAST NAME): Laura Belloni1, 3, Lichun Li4, Gianna Aurora Palumbo1, 2, Srinivas Reddy Chirapu 5, Ludovica Calvo1, 3, M. G. Finn5, Adam Zlotnick4, Massimo Levrero1, 2
Institutional Author(s):
INSTITUTIONS (ALL): 1. Dept of Internal Medicine, Sapienza University, Rome, Italy.
2. Life Nanosciences Laboratory, Sapienza University, Rome, Italy.
3. EAL Inserm U785, Sapienza University, Rome, Italy.
4. Department of Molecular & Cellular Biochemistry , Indiana University, Bloomington, IN, United States.
5. Dept Chemistry, Scripps Research Inst, La Jolla, CA, United States.
ABSTRACT BODY: Background: The development of novel combination based therapies for HBV infection requires new antivirals that block viral life cycle functions other than those associated with the viral polymerase. The HBV Core, that comprises the viral capsid, nucleic acid, and host and viral ancillary proteins, represents an attractive target. Proper assembly of the capsid is critical for RNA packaging, reverse transcription, and intracellular trafficking. Moreover, core proteins (Cp) have been shown to interact with histones and to bind the nuclear cccDNA, possibly contributing to the regulation of cccDNA function and the maintainance of the cccDNA stability. Hetero-aryl-dihydropyrimidines (HAPs) are a new class of antivirals inhibiting HBV replication in vitro and in vivo. HAPs enhance the rate and the extent of core protein (Cp) assembly over a broad concentration range and act as allosteric effectors to induce an assembly-active state or, at high concentration, stabilize preferentially non-capsid polymers of Cp. Here we investigated the impact of HAP12 on cccDNA formation, levels and transcription as part of its antiviral activity against HBV.
Methods: Capsid-associated HBV-DNA (TaqMan real-time PCR), cccDNA (TaqMan real-time PCR) and pgRNA levels (quantitative real-time PCR with specific primers), were assessed both in HepG2 cells transfected with full length HBV genomes and in a HepG2 stable clone that accumulates cccDNA, left untreated or treated with the hetero-aryl-dihydropyrimidine HAP12 at 1-5 microM.
Results: We confirmed, using the cccDNA ChIP assay, that Cp is recruited onto the cccDNA in HBV replicating cells. HAP12 treatment of cells transfected with wild type linear HBV genomes showed a complete suppression of HBV replication at 72 and 96 hrs with a peak >50% reduction of pgRNA transcription at 96 hours, without significant changes in cccDNA levels. In the HepG2 stable cell line we confirmed the strong HAP12 inhibitory effect on pgRNA transcription and HBV replication as well as the lack of significant effect on steady state cccDNA levels. By treating the cells with HAP12 already before the establishment of the cccDNA pool we could also show that cccDNA formation and accumulation are not targeted by HAP12.
Conclusions: HAPs binding to HBV capsid, in addition to conformational change resulting in a core protein that does not support HBV replication, also target the nuclear cccDNA. We show that inappropriate assembly and effective cytoplasmic trapping of Cp induced by HAPs affects, through mechanisms that have not been yet clarified, cccDNA function rather than the formation of cccDNA or the establishment/maintenance of the cccDNA nuclear pool.

肝胆速递

TITLE：Heteroaryldihydropyrimidines（HAPS）抑制HBV核衣壳的成熟与cccDNA转录在组织培养



姓氏，名字）：劳拉Belloni1，3，丽春LI4，吉安娜极光Palumbo1，2，5 SRINIVAS雷迪Chirapu，Ludovica Calvo1 3，MG Finn5，亚当Zlotnick4，马西莫Levrero1，2
机构（S）：
机构（ALL）：1。内科部，Sapienza大学，罗马，意大利。
2。生命纳米科学实验室，Sapienza大学，罗马，意大利。
3。 EAL INSERM的U785，Sapienza大学，罗马，意大利。
4。 ，IN，美国，印地安那大学伯明顿分校分子和细胞生物化学系。
5。 ，斯克里普斯研究部化学研究所，拉霍亚，加利福​​尼亚州，美国。
抽象的身体：背景：新的组合为基础的治疗HBV感染的发展需要新的抗病毒药物，阻止病毒的生命周期与病毒聚合酶以外的功能。 HBV核心，包括病毒衣壳，核酸，以及主机和病毒辅助蛋白，是一个有吸引力的目标。合适的组装衣壳的是，反转录，RNA包装和胞内运输的关键。此外，已经示出核心蛋白（Cp）的交互与组蛋白和核的cccDNA绑定，可能作出贡献到cccDNA的功能和修的cccDNA的稳定性的调节。杂芳基 - 二氢嘧啶（HAPs）等在体外和体内的是一类新的抗病毒药物抑制乙肝病毒复制。有害空气污​​染物提升率和核心蛋白​​（CP）组装的程度，在很宽的浓度范围内，并作为变构效应诱导的组件激活状态，或者，在高浓度下，稳定的CP优先非衣壳聚合物的的。在这里，我们调查的影响，HAP12 cccDNA的形成，其对乙肝病毒的抗病毒活性的水平和转录。
评估方法：衣壳相关HBV-DNA（荧光定量实时PCR），，cccDNA的（TaqMan实时PCR）和pgRNA水平（实时定量PCR特异性引物），在转染的HepG2细胞全长HBV基因组和的HepG2的稳定克隆累积cccDNA的，留下未处理的或与杂芳基 - 二氢嘧啶HAP12处理在1-5 MICROM。
结果：我们证实，cccDNA的ChIP实验，cp是招募到cccDNA的乙肝病毒复制的细胞。 HAP12治疗与野生型线性HBV基因组转染的细胞在72和96小时时表现出一个完整的抑制HBV复制具有峰值pgRNA转录减少> 50％在96小时，没有显着的变化cccDNA水平。在肝癌稳定的细胞系中，我们证实了抑制作用强HAP12，对的pgRNA转录和HBV复制，以及缺乏稳定状态cccDNA水平的显着影响。已经成立前的cccDNA的池处理的细胞HAP12，我们也可能会出cccDNA的形成和积累的目标HAP12。
结论：结合到HBV衣壳，在除了不支持HBV复制的核心蛋白，也针对核的共价闭合环状DNA的构象变化导致的有害气体污染物。我们表明，不适当的装配和有效的细胞质捕获CP引起的有害空气污​​染物的影响，通过机制还没有得到澄清，的cccDNA的功能，而不是形成cccDNA的建立/维护的cccDNA的核池。

lin12345

谢谢分享 最好翻译过来再稍微修改下  

咬牙硬挺

似乎是针对ccCDNA的新开发的药物啊？自动翻译确实不好

StephenW

HAPs在2003年首次被发现. 上述的研究
是最新了解HAPs具体机制. BAY 41-4109仍是

I期临床试验

HAP Compound                       Bay 41-4109

Inhibits viral nucleocapsid

AiCuris,                       Germany

Phase I

http://www.hbvhbv.com/forum/viewthread.php?action=printable&tid=304009
作者: 清照    时间: 2003-12-17 01:16

异芳基二氧嘧啶 (heteroaryldihydropyrimidines HAPs)是一类新的化合物，具有抑制HBV病毒复制的效果，而与其他几类抗病毒药物的作用机理完全不同。

与核苷类药物不同，HAPs没有抑制HBV的DNA聚合酶，目前研究人员还没有查明HAPs抑制病毒复制的具体机制。由于这类药物对病毒聚合酶没有作用，因此推测它可能不会出现耐药的病毒变异。

2003年2 月发表在Science 上的一项研究表明，它是通过抑制病毒核蛋白外壳的形成，来抑制病毒复制的，与HBV核心蛋白水平减少相关。而核蛋白壳是病毒复制所需装配的病毒颗粒。

在所有测试过的HAPs化合物中， Bay 41 4109 是最有潜力的，已经证明它在HBV转基因小鼠的治疗中有效，并且潜在的药物不良反应和毒性也较为合适（可以耐受）。目前尚未在人体中测试，还需要进一步的临床试验来证明它的效果。