EASL2015:ANTI衣壳药物HAP12和AT130 TARGET HBV核心蛋白核功能

StephenW

RS-3403
Viral hepatitis
Hepatitis B & D - Experimental

ANTI CAPSID DRUGS HAP12 AND AT130 TARGET HBV CORE PROTEIN NUCLEAR FUNCTIONS
Laura Belloni1, 2, Gianna Aurora Palumbo2, Leonardo Lupacchini2, Lichun Li3, Srinivas Reddy Chirapu4, Ludovica Calvo2, M G Finn4, Uri Lopatin5, Adam Zlotnick3, 5, Massimo Levrero* 1
1Center for Life Nano Science (CLNS), IIT-Sapienza University, 2Dept of Internal Medicine - DMISM, Sapienza University, Rome, Italy, 3Dept of Molecular & Cellular Biochemistry, Indiana University, Bloomington, 4Dept Chemistry, Georgia Institute of Technology, Atlanta, 5Assembly Bioscience, Bloomington, United States


Corresponding author’s email: [email protected]


Background and Aims:
HBV Core protein (Cp) represents an attractive new therapeutic target for HBV chronic infection. In addition to their role in capsid assembly, pgRNA packaging and reverse transcription, Cp has been shown to bind the nuclear cccDNA mini-chromosome as well as a number of cellular genes promoters. Several compounds that target Cp and HBV capsids assembly, including the Hetero-aryl-dihydropyrimidines (HAPs) and the phenyl-propenamide derivatives AT61 and AT130, have been shown to inhibit HBV replication in vitro and in vivo. HAPs and AT130 enhance the rate and the extent of Cp assembly leading to non-functional capsids and, at high concentration, stabilize preferentially non-capsid polymers of Cp. Here we investigated the ability of the Core protein Assembly Modulators (CaMPs) HAP12 and AT130 to affect both nuclear (cccDNA structure and transcription) and cytoplasmic (capsid maturation and replication) Cp functions as part of their antiviral activity against HBV.
Methods:
HAP12 and AT130 effects on 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 in: a) HBV-infected NTCP-HepG2 cells; b) AD38 inducible HBV stable cell line. Recruitment of HBc and histone modifications on the viral minichromosome were assessed using the cccDNA ChIP assay in AD38 cells.
Results:
CaMPs treatments started at day 6 post-infection (NTCP-HepG2) or day 6 post-induction (AD38 tet-off cells) resulted in a very strong inhibition of HBV replication (>95%) and a significant but incomplete reduction of the stable cccDNA pool. A strong effect on cccDNA-dependent HBeAg production (AD38 tet-off) and pgRNA transcription (AD38 tet-off / tet-on and NTCP-HepG2 infected cells) was also demonstrated. The ability HAP12 to target cccDNA transcription was confirmed by the reduced cccDNA-bound H3 histone acetylation and the descreased HBc occupancy on the cccDNA in induced AD38 cells. Importantly, when CaMPs treatment was started during infection, cccDNA formation/accumulation was completely inhibited (>95%) and viral replication was blunted.
Conclusions:
Anti-capsid compounds (CpAMs) have an impact on Cp nuclear functions at multiple levels: block of new cccDNA formation / accumulation, reduction of an established cccDNA pool and inhibition of HBc occupancy and histone acetylation on the cccDNA that translate into a reduced pgRNA transcription.


Disclosure of Interest: L. Belloni:  Grant: Gilead, G. A. Palumbo: : None Declared, L. Lupacchini: : None Declared, L. Li: : None Declared, S. R. Chirapu: : None Declared, L. Calvo: : None Declared, M. G. Finn: : None Declared, U. Lopatin:  Stockholder: Assembly Bioscience,  Employee: Assembly Bioscience, A. Zlotnick:  Stockholder: Assembly Bioscience, M. Levrero:  Consultant: Gilead, BMS, Janssen, Medimmune, Tekmira, Galapagos,  Sponsored Lectures (National or International): Gilead, BMS, Janssen, Roche, MSD

StephenW

RS-3403

病毒性肝炎

乙肝＆D - 实验





ANTI衣壳药物HAP12和AT130 TARGET HBV核心蛋白核功能

劳拉Belloni1，2，吉安娜极光Palumbo2，莱昂纳多Lupacchini2，丽春钒锂，SRINIVAS雷迪Chirapu4，卢多维卡Calvo2，MG Finn4，乌里Lopatin5，亚当Zlotnick3，5，马西莫Levrero * 1

1Center生命纳米科学（CLNS），IIT-Sapienza大学，内科2Dept - DMISM，Sapienza大学，罗马，意大利，分子与细胞生物化学3Dept，印地安那大学伯明顿分校，4Dept化学，技术，亚特兰大佐治亚理工学院， 5Assembly生物科学，布卢明顿，美国



通讯作者的邮箱：[email protected]



背景和目的：乙肝病毒核心蛋白（CP）代表HBV慢性感染一个有吸引力的新的治疗靶点。除了其在衣壳包装，pgRNA包装和反转录的作用是：Cp已显示结合核cccDNA的微型染色体以及若干细胞基因的启动子。几个化合物组件靶向Cp和HBV衣壳，包括杂芳基dihydropyrimidines（HAPS）和苯基丙烯酰胺衍生物AT61和AT130，已被证明能抑制HBV复制的体外和体内。污染物和AT130提升幅度1和Cp组件导致非功能性衣壳的程度，并且在高浓度，稳定优先Cp的非衣壳聚合物。在这里，我们调查的核心蛋白大会调制器（营）HAP12和AT130的核常兼备（cccDNA的结构和转录）和细胞质（衣壳的成熟和复制）CP功能影响他们对乙肝抗病毒活性的一部分的能力。

方法：1）的HBV：HAP12和衣壳相关HBV-DNA AT130效果（TaqMan TM实时PCR）的cccDNA（TaqMan TM实时PCR）和pgRNA水平（定量实时PCR用特异性引物），在进行了评估 - 感染NTCP-HepG2细胞;二）AD38诱导乙型肝炎病毒的稳定细胞系。的HBc和病毒微染色体组蛋白修饰的招聘都用AD38细胞cccDNA的ChIP实验评估。

结果：CAMPS治疗开始在第6天在感染后（NTCP-HepG2细胞）或第6天诱导后（AD38 TET断细胞）产生了一个很强的HBV复制（> 95％）和一个显著但不完全降低的抑制稳定的cccDNA池。对cccDNA的依赖HBeAg的生产（AD38 TET-关闭）和pgRNA转录（AD38 TET-关闭/ TET-上NTCP-HepG2细胞感染的细胞）有很强的影响也被证实。能力HAP12靶向cccDNA的转录是由降低的cccDNA结合的H3组蛋白乙酰化和在cccDNA的所述descreased的HBc占用诱导AD38细胞证实。重要的是，在感染过程中，当开始治疗营地，cccDNA的形成/积累完全被抑制（> 95％）和病毒复制被减弱。

结论：抗衣壳化合物（CpAMs）具有多层次上的Cp核功能的影响：在转化为降低cccDNA的HBC的占用和组蛋白乙酰化建立cccDNA的游泳池和抑制新的cccDNA的形成/堆积块，减少pgRNA转录。



股权变动：L.贝罗尼：格兰特：Gilead公司，GA帕隆博：无申报，L. Lupacchini：无申报，L.李：无申报，SR Chirapu：无申报，L.卡尔沃：无申报， MG芬兰人：无申报，U.洛帕廷：股东：生物科学大会，职工：生物科学大会，A. Zlotnick：股东：生物科学大会，M. Levrero：顾问：Gilead公司，BMS，扬森，MedImmune公司，Tekmira，加拉帕戈斯群岛，赞助商讲座（国家或国际）：基列，BMS，扬森，罗氏，MSD

newchinabok

at61，at130不能被人体吸收，没有开发价值，hap12和莫非赛定一样，结果难以预料

StephenW

"at61，at130不能被人体吸收" - 我没有听说过, 你可以给一个参考?

newchinabok

我用手机上网，回去找，


莫非赛定试验停顿下来，sw您怎么看，是药物本身问题，还是其它原因，按理论上讲应该是有希望的，东阳光丙肝药也没看见临床，批文是批了，东阳光药物项目好像都停了

战天斗hbv

newchinabok 发表于 2015-4-12 10:22
我用手机上网，回去找，

理论上我认为莫非赛定上市简直就是小意思啊、真是奇了怪了

newchinabok

http://db.yaozh.com/zhuce/zhuce2.php

newchinabok

http://db.yaozh.com/zhuce/zhuce2.php

zgct

中国的不少科技及关键技术一到最后亮相时都好像被阉割了似的！牛b容易吹，30多年来都经常这样，。。。。。真心不如筹个亿元级别资金去，唯一求拜美国专家，奖励国外英才！

StephenW

莫非赛定试验停顿下来 ?
http://db.yaozh.com/zhuce/zhuce2.php? 哪个项目？