Predictors of hepatitis B cure using gene therapy to deliver DNA cleavage enzymes: a mathematical modeling approach.
Predictors of hepatitis B cure using gene therapy to deliver DNA cleavage enzymes: a mathematical modeling approach.
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DOI：
10.1371/journal.pcbi.1003131
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发表时间：
2013
影响因子：
4.3
通讯作者：
Jerome KR
中科院分区：
文献类型：
--
作者：
Schiffer JT;Swan DA;Stone D;Jerome KR
大多数慢性病毒感染是用抑制复制但不能治愈的小分子疗法来管理的，因为尽管进行了数十年的抑制性治疗，非复制型病毒形式仍会持续存在。因此，有许多战略在发展中，以消除所有非复制病毒从身体。我们目前正在设计特异性靶向B型肝炎病毒共价闭合环状DNA（HBV cccDNA）的DNA切割酶，HBV cccDNA是病毒的附加体形式，尽管有有效的抗病毒治疗，但仍然存在。DNA切割酶，包括归巢核酸内切酶或大范围核酸酶、锌指核酸酶（ZFN）、TAL效应物核酸酶（TALEN）和CRISPR相关系统9（Cas9）蛋白，可以破坏病毒DNA的特定区域。因为DNA修复是容易出错的，当靶序列突变时，病毒可以在重复的切割事件后被中和。DNA裂解酶将作为进入肝细胞的病毒载体内的基因递送。在这里，我们开发的数学模型，描述了DNA切割酶的交付和细胞内活动。模型模拟预测，高载体与靶细胞比率、通过体液免疫对递送载体的有限去除以及酶与其DNA靶标之间的亲和结合将促进最高水平的cccDNA破坏。如果DNA切割和易错修复不会使病毒附加体复制失效，则可能发生对切割酶的从头耐药性的发展：我们的模型预测，靶向不同重要cccDNA区域的多种酶的同时递送或不同酶的顺序递送都是避免多酶耐药性的潜在有用策略。如果在根除试验期间同时给予抗病毒治疗，cccDNA持久性的潜在动力学不太可能影响治愈概率。最后，我们描述了可用于验证模型的实验，这反过来又为动物和最终人类潜在治疗试验的剂量选择提供了重要信息。正在开发创新的新方法来根除直到最近还被认为是不可治愈的病毒感染。我们感兴趣的是工程DNA切割酶，可以切割和失能持久的病毒。其中一个障碍是，这些酶必须作为病毒载体中对人类无害的基因传递到受感染的细胞。在本文中，我们开发了一系列方程，描述了这些酶向其预期目标的传递，以及细胞内DNA切割的活性。虽然我们的数学模型是迎合对肝炎B病毒感染，它是广泛适用于其他感染，如艾滋病毒，以及肿瘤和代谢疾病的异常基因表达的特点。某些酶可能比其他酶更强烈地结合DNA，而如果靶向病毒DNA的不同区域，不同的酶也可能协同结合。我们预测，如果将此类酶有效递送到高比例的受感染细胞，将对增加治愈可能性至关重要。我们还表明，我们的方程将作为一个有用的工具，用于确定最重要的功能，治愈方案，并最终指导临床试验的剂量计划，以确保根除B型肝炎的最小数量的可能的剂量。
Most chronic viral infections are managed with small molecule therapies that inhibit replication but are not curative because non-replicating viral forms can persist despite decades of suppressive treatment. There are therefore numerous strategies in development to eradicate all non-replicating viruses from the body. We are currently engineering DNA cleavage enzymes that specifically target hepatitis B virus covalently closed circular DNA (HBV cccDNA), the episomal form of the virus that persists despite potent antiviral therapies. DNA cleavage enzymes, including homing endonucleases or meganucleases, zinc-finger nucleases (ZFNs), TAL effector nucleases (TALENs), and CRISPR-associated system 9 (Cas9) proteins, can disrupt specific regions of viral DNA. Because DNA repair is error prone, the virus can be neutralized after repeated cleavage events when a target sequence becomes mutated. DNA cleavage enzymes will be delivered as genes within viral vectors that enter hepatocytes. Here we develop mathematical models that describe the delivery and intracellular activity of DNA cleavage enzymes. Model simulations predict that high vector to target cell ratio, limited removal of delivery vectors by humoral immunity, and avid binding between enzyme and its DNA target will promote the highest level of cccDNA disruption. Development of de novo resistance to cleavage enzymes may occur if DNA cleavage and error prone repair does not render the viral episome replication incompetent: our model predicts that concurrent delivery of multiple enzymes which target different vital cccDNA regions, or sequential delivery of different enzymes, are both potentially useful strategies for avoiding multi-enzyme resistance. The underlying dynamics of cccDNA persistence are unlikely to impact the probability of cure provided that antiviral therapy is given concurrently during eradication trials. We conclude by describing experiments that can be used to validate the model, which will in turn provide vital information for dose selection for potential curative trials in animals and ultimately humans. Innovative new approaches are being developed to eradicate viral infections that until recently were considered incurable. We are interested in engineering DNA cleavage enzymes that can cut and incapacitate persistent viruses. One hurdle is that these enzymes must be delivered to infected cells as genes within viral vectors that are not harmful to humans. In this paper, we developed a series of equations that describe the delivery of these enzymes to their intended targets, as well the activity of DNA cutting within the cell. While our mathematical model is catered towards hepatitis B virus infection, it is widely applicable to other infections such as HIV, as well as oncologic and metabolic diseases characterized by aberrant gene expression. Certain enzymes may bind DNA more avidly than others, while different enzymes may also bind cooperatively if targeted to different regions of viral DNA. We predict that such enzymes, if delivered efficiently to a high proportion of infected cells, will be critical to increase the probability of cure. We also demonstrate that our equations will serve as a useful tool for identifying the most important features of a curative regimen, and ultimately for guiding clinical trial dosing schedules to ensure hepatitis B eradication with the smallest number of possible doses.
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