Split Proteins As Boolean Circuits and Drugs of a New Kind

描述（由申请人提供）：在这份题为“将蛋白质拆分为布尔电路和新型药物”的 EUREKA 申请中，我们建议验证、优化和实施一种治疗“复杂”疾病的新方法。在这种被称为删除特异性靶向（DST）的方法中，异常细胞以一种新颖的方式通过其 DNA 的突变变化，特别是纯合 DNA 删除（HD）来靶向。例如，许多癌症已被证明含有 HD。 （虽然癌症是一个相关的例子，但相同的“DST”逻辑适用于任何可以证明含有 HD 的不良细胞谱系。换句话说，DST 策略也与癌症以外的疾病相关。）纯合性缺失的一个显着特性是这种“阴性”目标无法恢复，从而充当治疗的不可变路标。为了针对HD，即不存在的“目标”，DST策略以一种新颖的方式将现有的和新的方法结合在一起，包括泛素融合技术、分裂泛素测定、锌指DNA识别蛋白和分裂限制性大范围核酸酶。 DST 策略还采用反馈机制，该机制接收来自作为布尔 OR 门运行的电路的输入，并涉及分裂大范围核酸酶的激活，从而破坏正常（非目标）细胞中的 DST 向量。 DST 的逻辑使得治疗选择性基本上无限增加成为可能。我们建议首先在酿酒酵母中测试和开发 DST，酿酒酵母是一种遗传上易于处理的生物体，复杂的操作速度更快 并且比哺乳动物细胞更容易。该项目的这一阶段之后将进行结果测试 （优化）培养的哺乳动物细胞中的电路，以及随后在患有特定疾病的完整小鼠中的电路。 项目叙述：本 EUREKA 申请中提出的研究旨在验证、开发和实施一种治疗“复杂”疾病的新方法。这种方法被称为删除特异性靶向（DST），使得在异常细胞（例如癌症（但不仅仅是癌症）细胞）中靶向纯合删除（HD）成为可能。最终在临床环境中实施 DST 策略可能会被证明可以治愈一系列 HD 相关疾病，而且基本上没有副作用。

DESCRIPTION (provided by applicant): IIn this EUREKA application, entitled "Split Proteins As Boolean Circuits and Drugs of a New Kind", we propose to verify, optimize, and implement a new approach to therapy of "complex" diseases. In this approach, termed deletion-specific targeting (DST), the abnormal cells are targeted, in a novel way, through mutational changes in their DNA, specifically homozygous DNA deletions (HDs). For example, many cancers have been shown to harbor HDs. (Although cancer is a relevant example, the same "DST" logic applies to any undesirable cell lineage that can be shown to contain HDs. In other words, the DST strategy is also relevant to diseases other than cancer.) A salient property of a homozygous deletion is that this "negative" target cannot revert, thereby serving as an immutable signpost for therapy. To target an HD, i.e., a "target" that is absent, the DST strategy brings together, in a novel way, both existing and new methodologies, including the ubiquitin fusion technique, split-ubiquitin assay, zinc-finger DNA-recognizing proteins and split restriction meganucleases. The DST strategy also employs a feedback mechanism that receives input from a circuit operating as a Boolean OR gate and involves the activation of split meganucleases, which destroy DST vector in normal (non-target) cells. The logic of DST makes possible an essentially unlimited increase in the selectivity of therapy. We propose to test and develop DST initially in the yeast S. cerevisiae, a genetically tractable organism in which sophisticated manipulations are faster and easier than in mammalian cells. This stage of the project will be followed by tests of resulting (optimized) circuits in mammalian cells in culture, and thereafter in intact mice with specific diseases. Project Narrative: The studies proposed in this EUREKA application are designed to verify, develop and implement a new approach to therapy of "complex" diseases. This approach, termed deletion-specific targeting (DST), makes possible the targeting of homozygous deletions (HDs) in abnormal cells, e.g., in cancer (but not only cancer) cells. The eventual implementation of DST strategy in a clinical setting may prove to be curative with a range of HD-relevant diseases, and in addition substantially free of side effects.