DEVELOPMENT OF MULTIVALENT ANTHRAX TOXIN INHIBITORS

描述(申请人提供)：正如最近发生的事件所表明的那样，炭疽病作为恐怖主义和战争的生物武器构成了致命的威胁。炭疽毒素全身中毒几乎总是致命的，但目前还没有有效的治疗方法。这项应用的目标是开发有效的抑制剂，防止炭疽毒素复合体的组装。炭疽毒素是这种疾病的主要症状。这些毒素由一种名为保护性抗原(PA)的宿主受体结合蛋白和两种名为致死因子(LF)和浮肿因子(EF)的酶组成。这些蛋白质是由炭疽芽孢杆菌以无毒单体的形式释放出来的。它们扩散到宿主细胞表面，形成致死毒素(LF+PA)和水肿性毒素(EF+PA)两种有毒蛋白复合体。PA是将LF和EF从细胞表面输送到胞浆的必要载体，在胞浆中LF和EF发挥细胞毒作用。因此，阻止LF或EF与PA结合的抑制剂应该是一种有效的抗炭疽病药物。 在此之前，我们已经确定了几个与PA弱结合并抑制LF或EF与PA相互作用的多肽。我们假设，通过协同作用，多价抑制剂(MVI)，其中一个肽的多个副本偶联到一个载体分子，将显示出显著增强的抑制作用。在我们的初步研究中，我们合成了几种基于葡聚糖的MVI，它们显示出比单独多肽更高的抑制活性。我们建议以生物相容、无毒、线性聚合物和环状低聚物为载体，开发优化的微球载体。 目的1.优化以葡聚糖和果胶为载体的微球载体。 目的2.探索以多聚谷氨酸骨架为基础的微血管病变。 目的3.设计和开发基于(-环糊精)核的七价“冠状”MVI。 在目标1和目标2中，我们计划合成一系列多肽-聚合物偶联物，其中多肽与主链的比例和主链的分子尺寸是系统地变化的。在目标3中，我们将通过计算建模来辅助冠醚抑制剂的设计。我们将在我们建立的抑制试验中测试所有MVI，并在两个大鼠中毒模型中测试最活跃的MVI。在这项研究中开发的有效抑制剂可以帮助保护我们免受致命的炭疽病的伤害，并对抗炭疽生物恐怖主义的威胁。

DESCRIPTION (provided by applicant): As demonstrated so terribly by recent events, anthrax poses a deadly threat as a biological weapon of terrorism and warfare. Systemic intoxication by anthrax toxins is virtually always fatal but effective treatment is not available. The goal of this application is the development of potent inhibitors that prevent the assembly of anthrax toxin complexes. Anthrax toxins are responsible for the major symptoms of the disease. The toxins consist of a host receptor-binding protein termed protective antigen (PA) and two enzymes termed lethal factor (LF) and edema factor (EF). These proteins are released from Bacillus anthracis as nontoxic monomers. They diffuse to the surface of host cells and assemble into two types of toxic protein complexes, lethal toxin (LF+PA) and edema toxin (EF+PA). PA is the necessary vehicle that transports LF and EF from the cell surface to the cytosol where LF and EF exert their cytotoxic effects. Hence, inhibitors that prevent the binding of LF or EF to PA should provide an effective antitoxic therapy against anthrax. Previously, we have identified several peptides that bind PA weakly and inhibit the interactions of LF or EF with PA. We hypothesize that, through cooperative interactions, multivalent inhibitors (MVIs), in which multiple copies of a peptide are coupled to a carrier molecule, will display significantly enhanced inhibitory effects. In our preliminary study, we have synthesized several dextran-based MVIs that show higher inhibitory activities than peptides alone. We propose to develop optimized MVIs based on biocompatible, nontoxic, linear polymers and cyclic oligomers as carriers. Aim 1. To optimize MVIs based on dextran and pectin carriers. Aim 2. To explore MVIs based on polyglutamate backbones. Aim 3. To design and develop heptavalent "crown" MVIs based on (-cyclodextrin cores. In Aims 1 and 2, we plan to synthesize a series of peptide-polymer conjugates in which the peptide-to-backbone ratio and the molecular size of the backbone are systematically varied. In Aim 3, we will assist the design of crown inhibitors by computational modeling. We will test all MVIs in our established inhibition assays and the most active MVIs in two rat intoxication models. Potent inhibitors developed in this study can help to protect us from deadly anthrax disease and fight the threat of anthrax bioterrorism.