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标题: 美国基因和细胞治疗年会 AAV-TALEN直接清除染色体整合的HBV [打印本页]

作者: MP4 时间: 2015-6-21 17:20 标题: 美国基因和细胞治疗年会 AAV-TALEN直接清除染色体整合的HBV

美国基因和细胞治疗年会ASGCT的主角：AAV-TALEN直接清除染色体整合的HBV
http://m.weibo.cn/2209901433/384 ... 10&wm=2468_1001

作者: MP4 时间: 2015-6-21 17:21

本帖最后由 MP4 于 2015-6-21 20:17 编辑

南非，德国，日本……

作者: 相信会幸福 时间: 2015-6-21 17:27

哪位大神解释下这句话的意思

作者: 42年才知道 时间: 2015-6-21 18:34

就是用基因编辑技术把整合到肝细胞染色体上的HBVDNA敲除掉。

作者: 相信会幸福 时间: 2015-6-21 18:56

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那乙肝能根治了？

作者: 相信会幸福 时间: 2015-6-21 18:57

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那乙肝能根治了？

作者: MP4 时间: 2015-6-21 20:03

本帖最后由 MP4 于 2015-6-21 21:16 编辑

Inhibition of Hepatitis B Virus Replication In Vivo Following Delivery of Antiviral TALENs With Recombinant Adeno-Associated Viral Vectors

Samantha Nicholson, Buhle Moyo, Kristie Bloom, Claudio Mussolino, Toni Cathomen, Koichi Watashi, Abdullah Ely, Patrick Arbuthnot. Antiviral Gene Therapy Research Unit, Faculty of Health Sciences, University of the Witwatersrand, Johannesburg, South Africa; Institute for Cell and Gene Therapy, Center for Chronic Immunodeficiency, University Medical Center Freiburg, Freiburg, Germany; Department of Virology II, National Institute of Infectious Diseases, Tokyo, Japan

Hepatitis B virus (HBV) is hyperendemic to sub-Saharan Africa where there are approximately 600 000 deaths per year associated with chronic HBV infection. HBV may have long periods of dormancy in carriers of the virus, which is dependent on persistence of the nuclear replication intermediate covalently closed DNA (cccDNA). Current HBV therapies, including nucleot(s)ide analogs, target the actively transcribed virus. The cccDNA is not cleared and may act as a reservoir for re-initiation of viral replication following treatment withdrawal. This poses a significant challenge for the successful treatment of chronic HBV sufferers, and makes the cccDNA intermediate an important target for novel HBV treatments. Employing gene editing technology to disable cccDNA has emerged as a promising novel therapeutic strategy. TALENs targeting the surface and the core viral open reading frames, previously described by our group, demonstrated efficacy against HBV targets. To advance this technology to therapeutic application, efficient delivery and evaluation of the TALENs' safety and target specificity in vivo and in vitro need to be established. This study aimed to evaluate the safety and efficacy of TALENs delivered by adeno-associated viral (AAV) vectors in vivo and in vitro. The HBV-infectable HepG2-hNTCP-C4 cell line was used to establish that the TALENs could disable cccDNA. Capacity for capsid modification, low immunogenicity and good hepatotropic transduction efficiency are favorable features that were considered in selecting these vectors. However, the delivery of TALENs by AAVs is technically complicated, as sequences encoding the two subunits of the complete TALENs exceed the capacity of single stranded AAVs. Consequently, we generated AAVs that encode each of the TALEN subunits. To constitute the complete TALENs, pairs of vectors were thus administered to mice or cultured cells. In vitro testing of AAV-TALENs was undertaken using the HepG2-hNTCP-C4 cell line. These cells present a novel tool for testing anti-HBV therapeutics as they overexpress the human sodium taurocholate co-transporting polypeptide (hNTCP) gene allowing them to be infected by HBV. The viral life-cycle is accurately recapitulated and enables assessment of effects of therapeutic agents on cccDNA. Since viral gene expression is dependent on transcription from the cccDNA intermediate in these cells, measurement of markers of HBV replication may be used as an indicator of cccDNA function. This study evaluated the potential of AAV delivered TALENs in vivo and in vitro by testing their activity and evaluating off-target activity, immunogenicity and liver toxicity. The results provide evidence for the utility of applying AAVs to the delivery of anti-HBV TALENs and offers further support for the feasibility of employing TALENs to treat chronic HBV infection.
Keywords: TALENs; Gene Correction/Modification/Targeting; AAV Vectors

Session: Poster Session: Gene Targeting and Gene Correction I (5:15 PM-6:45 PM)
Date/Time: Wednesday, May 13, 2015 - 5:15 pm
Room: Elite Hall A
Epigenetic Silencing of Hepatitis B cccDNA In Vitro and In Vivo Using AAV-Delivered Engineered Repressor Transcription Activator-Like Effector

Buhle Moyo, Samantha Nicholson, Ilke Roelofse, Carol Crowther, Kristie Bloom, Claudio Mussolino, Toni Cathomen, Koichi Watashi, Abdullah Ely, Patrick Arbuthnot. Antiviral Gene Therapy Research Unit, School of Pathology, University of the Witwatersrand, Johannesburg, Gauteng, South Africa; Institute for Cell and Gene Therapy, Center for Chronic Immunodeficiency, University Medical Center Freiburg, Freiburg, Germany; Department of Virology II, National Institute of Infectious Diseases, Tokyo, Japan

Hepatitis B virus (HBV) infection is hyper-endemic (>8 % chronic carriers) to parts of Asia and sub-Saharan Africa. Persistent HBV infection predisposes to liver diseases such as cirrhosis and hepatocellular carcinoma. Current anti-HBV therapies face several limitations. RNA interference-based therapies lack the robustness and specificity required for therapeutic effect while interferon-α and nucleotide/side analogs function post-transcriptionally and thus allow for the persistence of the covalently closed circular (cccDNA). cccDNA may persist indefinitely and enables re-initiation of HBV replication after withdrawal of treatment. Disabling the cccDNA is essential for the successful treatment of chronic HBV. Our group previously described effective anti-HBV transcription activator-like effector (TALE) nucleases (TALENs). Although potentially useful to counter HBV replication, one drawback is that viral sequences integrated into the host genome may be susceptible to digestion by the TALENs. Transcriptional silencing, rather than cleavage, of cccDNA may therefore be preferable to avoid causing undesirable mutations in the host. To this end, TALE binding domains designed to target the viral preS2 promoter and the basic core promoter/enhancer II regions were fused to a Krüppel-associated box repressor domain to generate repressor-TALEs (rTALEs). These rTALEs were shown to inhibit viral replication in vitro and in vivo without inducing significant toxicity. In an in vivo murine model using hydrodynamic transfection with an HBV replication-competent plasmid, a reduction in secreted HBV surface antigen (HBsAg) of 97% and 98% was seen at day 3 and 93% and 96% at day 5 for P1L and P1R respectively. To develop this approach as a feasible therapy, rTALE-encoding sequences were incorporated into recombinant adeno-associated viruses (rAAVs) and assessed in the HepG2.hNTCP-C4 cell line. These cells overexpress the HBV receptor, human sodium taurocholate co-transporting peptide (hNTCP). The HepG2.NTCP-C4 cell line is infectable with HBV and viral gene expression is dependent on formation of cccDNA. Measurement of viral markers of replication may thus be used as an indicator of inhibitory effects on cccDNA. The anti-HBV efficacy of the rTALEs and epigenetic modification of the targeted HBV DNA was characterized. Chromatin immunoprecipitation assays determined the binding of the rTALEs to the cccDNA and induction of epigenetic markers of viral gene suppression. Mobility shift assays confirmed specificity of rTALE binding. In vivo efficacy of rAAV-delivered rTALEs was evaluated in transgenic HBV mice. Our study provides valuable information on the potential therapeutic utility of rTALEs and demonstrates the feasibility of the approach for treatment of HBV.

Keywords: AAV Vectors; Gene Correction/Modification/Targeting; Infectious Diseases

Session: Simultaneous Oral Abstract Sessions: Gene Editing and Gene Regulation II (3:30 PM-5:30 PM)

Date/Time: Friday, May 15, 2015 - 4:30 pm

Room: Empire C

作者: MP4 时间: 2015-6-21 20:27

本帖最后由 MP4 于 2015-6-21 20:33 编辑

http://www.cansa.org.za/files/20 ... -Prof-Arbuthnot.pdf用药5天老鼠表面抗原消失

作者: 齐欢畅2 时间: 2015-6-21 20:28

The HBV-infectable HepG2-hNTCP-C4 cell line was used to establish that the TALENs could disable cccDNA.

作者: 齐欢畅2 时间: 2015-6-21 20:28

重大利好。

作者: 相信会幸福 时间: 2015-6-21 20:45

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老鼠那强大的免疫力，不能比啊

作者: MP4 时间: 2015-6-21 20:53

相信会幸福发表于 2015-6-21 20:45
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老鼠那强大的免疫力，不能比啊

这药完全不依赖免疫系统

作者: 战天斗hbv 时间: 2015-6-21 20:55

猛！！！

作者: 相信会幸福 时间: 2015-6-21 21:10

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这意思是HBV可以根除了？

作者: MP4 时间: 2015-6-21 21:17

相信会幸福发表于 2015-6-21 21:10
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这意思是HBV可以根除了？

先要动物身上做长期安全试验的，这是基因治疗不容易批准的

作者: 南湖淋雨 时间: 2015-6-21 21:24

比较靠谱，对天朝的专家我实在不能说什么。。。。

作者: MP4 时间: 2015-6-21 21:34

南湖淋雨发表于 2015-6-21 21:24
比较靠谱，对天朝的专家我实在不能说什么。。。。

http://www.ebiotrade.com/newsf/2013-11/201311194704262.htm

作者: 9病成医 时间: 2015-6-22 08:48

盼望加快研究进度，尽快用于临床。

作者: styxl 时间: 2015-6-22 15:31

哈,很神奇的样子

作者: 相信会幸福 时间: 2015-6-22 18:24

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您觉得这个有希望吗

作者: MP4 时间: 2015-6-22 19:49

相信会幸福发表于 2015-6-22 18:24
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您觉得这个有希望吗

有……

作者: 咬牙硬挺 时间: 2015-6-23 13:47

不知道副作用怎样

作者: 沁海 时间: 2015-6-27 12:54

人体实验有需要支我一声

作者: tonychant 时间: 2015-6-27 14:50

望梅止渴

作者: 相信会幸福 时间: 2015-6-27 15:25

希望成功

作者: newchinabok 时间: 2015-10-26 09:02

这个技术如何？

作者: hfdr 时间: 2016-8-22 15:35

这个成功，才是釜底抽薪！

作者: 山野菜面 时间: 2016-8-22 16:43

不知道得等到猴年马月

作者: yelin1208 时间: 2016-8-22 22:41

看到希望了

作者: 东霸天 时间: 2016-8-24 06:13

什么意思啊？楼主帮解释一下呗？

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