| [ISVHLD2012]当前肝炎的挑战——Prof.Charles M. Rice专访 -----
来源： 作者：国际肝病 发布时间：2012-7-17 13:55:59 阅读：26
过去二十多年来，我们欣喜地看到丙型肝炎的细胞培养系统已经取得巨大进步。对于丙型肝炎而言，在非A和非B的时代，人们试图对这种病毒进行细胞内培养，但没有取得成功。在丙肝病毒鉴定后，即使我们能够开发出敏感的检测复制水平的试剂，但是仍不能马上做到这一点。我认为我们确实为之奋斗了许多年，直到病毒发现十年之后第一个复制子才被鉴定出来。
Hepatology Digest: Your presentation on the first day of the conference was entitled ‘The Challenges of Hepatitis Today’. One of the challenges you discussed was the issue of getting virus replication in the lab both in in vitro models and small animal models and the problems that have been faced. Can you explain some of the challenges that are being encountered in in vitro models and the ways that researchers are now making hepatocyte models in the lab?
《国际肝病》：您在大会首日进行了“肝炎的挑战”的主旨演讲。在演讲中，您提到其中的一个挑战就是关于病毒复制方面的，您认为在实验条件下中要让病毒复制非常困难，无论是在体外模型以及小动物模型中均是如此，并且提及目前面临的困难。您能否解释一下在体外模型中，目前正面临的一些挑战以及现在研究者在实验室中如何制备肝细胞模型。
Dr Rice: We have seen an evolution in the cell culture systems that have been developed for hepatitis C over the last twenty-plus years. In the non-A/non-B era of hepatitis C， many people tried to show that they could grow this virus in cell culture but really without success. Even after the identification of the virus， it wasn’t immediately possible to do that even though we could develop fairly sensitive methods for following replication. I think we really struggled for many years and wasn’t until ten years after the discovery of the virus that the first replicons were identified and those were done using a drug selection where we could engineer a viral genome to express what is basically an antibiotic resistance marker that allowed us to identify cells that would harbor replicating RNAs. That really began with the hepatoma cells being permissive for some of these replicons and the identification of adaptive mutations and I think that was a great boom for the field because at least for the first time， we had a cell-based replication system where we began to study basic viral replication processes and also look for antiviral drugs. But even that system didn’t allow us to study replication in the real hepatocyte environment; this was a hepatoma cell which as a cancer cell is not the normal cell in which the virus replicates. In the last several years we have seen a number of systems that have been developed. One of them is using primary adult hepatocytes that are plated in micropatterned co-cultures under conditions where these hepatocytes maintain their differentiated function. Another one is taking fetal liver cells and plating those in defined media and those are also maintaining many hepatocyte functions and can be infected by the virus. The third area which I think is the most exciting， is the ability to take induced pluripotent human stem cells and differentiate these to cells that mimic hepatocytes and we and others have shown (Hengli Tang will be presenting at the meeting) that those cells are permissive for HCV replication. So now finally after almost twenty years， we actually have some systems where we can maintain primary hepatocyte function and infect them with the virus. That’s great news for those studying virus host cell biology.
Rice博士：过去二十多年来，我们欣喜地看到丙型肝炎的细胞培养系统已经取得巨大进步。对于丙型肝炎而言，在非A和非B的时代，人们试图对这种病毒进行细胞内培养，但没有取得成功。在丙肝病毒鉴定后，即使我们能够开发出敏感的检测复制水平的试剂，但是仍不能马上做到这一点。我认为我们确实为之奋斗了许多年，直到病毒发现十年之后第一个复制子才被鉴定出来。这些还是使用一种药物选择的方法鉴定出来的，在这里我们可以设计一个病毒基因组来表达一种标记物以确定哪些细胞能够复制RNAs，这种方法也是以抗菌素抗药性标记物为基础的。实际上，我们的研究是从对于这些复制子耐受的肝癌细胞开始的，并且逐步鉴定出一些适应性突变的细胞，我认为这是该领域的一个重大进步，因为至少我们首次建立了一个以细胞为基础的复制体系，同时我们可以以此为基础开展关于病毒复制进程的基础研究，并且开展抗病毒药物方面的研究。但是，即使该细胞模型并不能让我们研究真正肝细胞环境中的病毒复制进程，但是它是一个肝癌细胞系，而不是正常的细胞。在过去的几年里，我们欣喜地看到已经建立了一些模型体系。其中一个就是使用原代成人肝细胞种植在联合培养的微板上，在这种环境中，肝细胞能保持分化功能。另外一个就是使用胎儿肝细胞并且使用已知成分培养液进行接种，它们也能保持一些肝细胞功能，并且可被病毒感染。第三个领域，我认为这是最令人兴奋的，就是目前有能力使用多能干人类干细胞进行诱导分化为具有一些肝细胞功能的细胞，我们以及其他一些研究者证实在这些细胞中，HCV病毒可以进行复制（Hengli Tang将在会议上进行演讲）。所以，在经过20年的努力后，目前我们已经取得进步，建立了一些既具有基本肝细胞功能，同时又能被病毒感染的细胞体系。这对于那些研究病毒宿主细胞生物学的研究者来说确实是一大喜讯。
Hepatology Digest: In recent years we have seen more and more studies on IL28B. Some researchers though have remarked that with the new DAAs， IL28B polymorphism testing is still ten years away which be too late. What are your thoughts?
《国际肝病》：近年来，我们已经看到对于IL28B的研究越来越多。一些研究人员认为对于不断涌现的直接抗病毒药物而言，IL28B多态性检测用于临床仍需10年的时间，可能为时已晚。对此，您有何看法？
Dr Rice: We are already seeing in many clinical settings that at least in an interferon-based regimen that the IL28B genotyping is potentially useful for making treatment decisions. There is still nothing better than seeing how a patient responds in the first several weeks of interferon therapy and whether they or not they show a rapid decrease in viremia. The feeling is that the predictive value of the good CC genotype versus the TT and the CT genotypes is likely to become less important as we move towards interferon-free direct-acting antiviral oral regimens， but I think we are also seeing that the ability for patients to have a favorable response to interferon is some measure of their innate immune responses and that can influence what you see even in the context of oral direct-acting antiviral combinations. So there is still some， presumably immune component， and perhaps an innate immune component， that is operating for clearance even in the absence of exogenous interferon. I know in the US and Europe， many physicians will go ahead and determine the IL28B genotype of their patients just to add that to the discussion as to whether or not they should consider getting treatment or not.
Rice博士：我们已经注意到在许多临床试验中，至少在以干扰素为基础的药物治疗中，IL28B的基因型检测对于治疗策略的选择有潜在的价值。但是到目前为止，还没有比患者对于干扰素治疗前几周的应答水平以及病毒血症是否快速降低等表现更好的指标。当我们的治疗模式转变为无干扰素的口服直接抗病毒药物方案时，IL28B基因型的检测对于治疗效果的预测价值优势将不复存在，我们将无法观察到基因CC型比TT型和CT型的优势，但是我认为我们同时也观察到患者对干扰素的良好应答也是其先天免疫应答的一种度量方法，这一点即使在直接抗病毒药物组合治疗时也有影响。因此，仍然存在一些因素，可能是免疫组分，也许是一种先天免疫成分，即使在无外源性干扰素的情况下仍能够清除病毒。据我所知，在美国和欧洲，许多医生继续坚持并决定对患者进行IL28B基因型检测，以帮助决定是否对其进行治疗。
Hepatology Digest: In the United States， HCV is the leading cause of chronic hepatitis. At the same time， China has nearly ten million infected patients so the problem went long unrecognized due to the prevalence of HBV. Both countries are facing challenges. How has the US government confronted this problem and what are your suggestions for China?
《国际肝病》：在美国，丙型肝炎是慢性肝炎的主要病因。同时，中国HCV感染者接近1000万，因为乙肝的流行导致这个问题长期被忽视。两国都面临着严峻的挑战。请问，美国政府是如何面对这个问题的？您对中国有什么建议吗？
Dr Rice: I wouldn’t necessarily follow the example of the United States government. The funding for viral hepatitis research and HCV research in particular is really tiny compared to the disease problem if you compare it to HIV for instance which is also a huge problem but there are fewer HIV infections in the US. If I had to make some recommendations to China， there are various aspects that need to be taken care of. Some of these we have done well in the United States like blood screening. The chances of getting hepatitis C from a blood transfusion or a blood product are essentially zero. If that isn’t the case in China now， it certainly should be something that should be taken care of to prevent a lot of potential new infections. The other thing is unsafe medical practices such as safe injections in the medical community. At least in the US， I think many people take for granted that when you go to a physician to get an injection that it is a clean non-reusable syringe and there is not much chance that you would spread something like hepatitis C by injecting people with an unclean syringe or a multidose vial. Those are things that you can definitely implement and take care of. The other thing， and this is something we have been slow on in the United States， is to make decisions about finding out who is actually infected because it is a slow progressive disease and the majority of people that have this virus don’t know that they are infected. I would guess that is also probably true in China. Historically there maybe hasn’t been the pressure to do that from a public health standpoint because the treatment for HCV was really pretty difficult for the patients to tolerate i.e. six to twelve months of interferon and ribavirin is not an easy regimen to take. Now as treatments become more effective and easier to tolerate then I think that really should drive us in the direction of identifying people who are infected， treating them and then having the opportunity to eliminate these slow progressive disease processes. The sooner you do that， the less trouble you will have in terms of liver disease. Finally this year， the CDC has made a recommendation in the US that those in the highest risk age groups， who are those born in the 1945 to 1965 era when people were using drugs and there was a lot of transmission going on， should be tested to identify those that are HCV positive so that they might get in contact with a physician and decide whether or not they want to be treated. Another thing that needs to be assessed here in China is the relative importance of chemotherapy for treating hepatitis C versus vaccination. What is the new incidence of infection? Are there high risk groups where a vaccine would really help? If you think of the success of developing a licensed HBV vaccine here in China， should the vaccine developers here be taking on the challenge of developing an HCV vaccine? These topics I think should be discussed and kept in mind as China moves forward to deal with the problem. The other thing is to find out what the prevalence really is. Even that in the Unites States is somewhat controversial because it depends on how unbiased the sampling of the population has been to determine the frequency of positivity. In the United States， Brian Edlin wrote a piece on this and he feels that the sero-prevalence rate that is reported in the United States is actually a significant underestimate of the number of people that are actually infected just because some of the groups that have the highest prevalence are not represented in those sero-surveys. We need to know what the scope of the problem is; we need to identify the people that are infected; and then consider the various treatment options for them.
Rice博士：我可能不一定完全按照美国的模式进行说明。与另外一种严重疾病HIV感染相比较而言，对于病毒性肝炎和HCV的研究资金资助实在是非常少，但事实上美国感染HIV的人数相对较少。如果我必须对中国的问题提出推荐意见的话，我认为需要在以下方面需要引起注意。首先，有一些在美国已经做得非常好了，比如血液筛查，目前由输血或者血制品而感染HCV的几率已经降低到零。如果在中国还不能做到，毫无疑问，如果在这一点上下工夫的话，将可能避免非常多的新发感染者。其次就是杜绝不安全的医疗行为，例如在医疗界推广安全注射。至少在美国，我认为许多人会想当然地认为，当你去医院接受治疗的时候，一定会使用清洁的一次性输液器，因此通过不洁注射感染HCV的可能性较低，这一点非常容易就可以办到。再次，就是进行筛查发现哪些是感染者，这一点在美国也是进展非常缓慢，因为HCV感染是一个进展非常缓慢的疾病，大部分感染者并不知道自己已经感染HCV。我想，中国的情况可能也是如此。从历史上，从公共卫生角度上看并不可能有进行筛查的压力，因为患者能否耐受决定对于HCV的治疗事实上相当困难，例如使用6至12个月的干扰素和利巴韦林是不是一个简单的方案。现在，治疗方案变得越来越有效，同时耐受性大大提高，我认为应该推动在人群中进行筛查，以便发现患者并进行治疗，达到减缓这些疾病进程的目标。终于在今年，美国CDC建议在风险较高的年龄组，即在1945年至1965年出生的人群，当有吸毒史时进行HCV筛查，因为已经发现大量传播。这些人群应该进行检测，并决定是否需要开始进行治疗。在中国，另一点需要评价的就是丙型肝炎抗病毒治疗与疫苗的相对重要性的问题。新发感染率如何？对于高危人群疫苗是否能真正起到作用？如果您认为乙肝疫苗在中国已经取得成功，那么疫苗开发商是否为开发HCV疫苗做好准备？我认为这些问题需要展开讨论并引起足够的重视，以便更好地处理这些难题。另外还需要对真正的流行现状进行调查。即使在美国，这一点也存在争论，因为它决定于人口抽样的阳性率是否存在偏倚。在美国Brian Edlin曾经为此进行讨论，认为美国所报道的血清阳性率显著低估了实际情况，因为高流行率人群并未包括在内。我们需要知道问题的严重性；我们需要筛查出感染者；然后再考虑哪些治疗方法适合于他们。
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发表于 2012-7-18 11:41
