Small molecule inhibitors of ricin and shiga toxins

描述(申请人提供)：蓖麻毒素是一种在蓖麻籽中发现的有效的B类蛋白毒素。它易于准备，使其成为国内和国际恐怖分子最喜欢的毒剂。由于针对该毒素的疫苗接种并不是保护普通民众的合理策略，NIAID已将识别和生产蓖麻毒素小分子抑制剂作为其生物防御计划的首要任务。为了实现这一目标，我们的项目是三个实验室的合作努力，具有促进抑制剂设计、合成和测试的互补技能。该项目将通过筛选现有的商业化合物来识别蓖麻毒素抑制剂。由于蓖麻毒素的X射线结构已知，它可以作为大型商业图书馆基于现代计算机的虚拟屏幕的模板。最具吸引力的候选人将在体外测试系统中进行评估。对1000种化合物的初步筛选已经确定了三种抑制剂，其中包括一种名为Z3911的抑制剂，其IC50为1 DM。此外，使用新开发的微孔板分析的高通量筛查将在哈佛大学的NSRB设施中进行。然后，来自虚拟和物理筛选的抑制剂将在设计周期中使用基于结构的方法进行优化，并使用最先进的组合化学专门创建多样性文库。我们希望将IC50值提高至少10到100倍，进入纳摩尔范围。我们提高抑制剂强度的第一项努力是使Z3911平台在三个位置多样化，创建一个小型衍生物库。提出了一种详细的综合战略，并作为一个例子，说明了如何从尚不清楚的平台上开发更多的线索。在设计方案中，相当强的体外抑制剂，如Z3911，将在Vero细胞试验的早期进行测试。这将使我们能够评估生物利用度和细胞毒性，并限制对有用化合物的进一步设计努力。然后将在动物挑战模型中对有效的、无毒的蓖麻毒素抑制剂进行评估，以进行化合物的最终提纯。我们的目标是创造可以预防或减轻动物毒性的蓖麻毒素(和同源志贺毒素)抑制剂，并作为商业或政府机构开发真正解毒剂药物的基础。

DESCRIPTION (provided by applicant): Ricin is a potent class B protein toxin found in castor seeds. Its ease of preparation has made it a favorite agent for domestic and international terrorists. Since vaccination against the toxin is not a plausible strategy for protection of the general populace, NIAID has made identification and production of small molecule inhibitors of ricin a top priority in its program for Biodefense. Toward this goal, our project is a collaborative effort from three laboratories with complimentary skills that facilitate inhibitor design, synthesis and testing. The project will be initiated with screens of existing commercial compounds to identify ricin inhibitors. Since the x-ray structure of ricin is known, it can serve as a template for a modern computer based virtual screen of large commercial libraries. The most attractive candidates will be assessed in an in vitro assay system. An initial screen of 1000 compounds has already identified three inhibitors including one, called Z3911, with an IC50 of 1 DM. In addition, high throughput screens using a newly developed micro-plate assay will be conducted at the NSRB facilities at Harvard. Inhibitors from both virtual and physical screens will then be optimized in design cycles using both structure-base methods and specifically created diversity libraries using state of the art combinatorial chemistry. We expect to improve IC50 values at least 10 to 100 fold into the nanomolar range. Our first effort to improve inhibitor strength is to diversify the Z3911 platform at three positions, creating a small library of derivative compounds. A detailed synthetic strategy is presented, and serves as an example for the manner in which additional leads will be developed from as yet unknown platforms. In the design scheme, reasonably strong in vitro inhibitors, like Z3911, will be tested early on in a Vero cell assay. This will allow us to assess bioavailability, and cell toxicity, and to restrain further design efforts towards useful compounds. Potent, non-toxic, ricin inhibitors will then be assessed in an animal challenge model for the final refinement of compounds. Our goal is create ricin (and homologous shiga toxin) inhibitors that can prevent, or alleviate toxicity in animals and serve as the basis for development, by commercial or government agencies, of true antidote drugs.