Directed evolution of inhibitors of anthrax toxin

描述（由申请人提供）：对新的生物恐怖威胁和新出现的传染病的快速反应的需求因药物发现的缓慢步伐而受挫。我们假设蛋白质合成机制可以被设计用于任何靶分子的高亲和肽和肽类似物抑制剂的体外快速进化。这里的目标是开发针对播散性炭疽的候选药物，这种疾病用抗生素无法治愈。以炭疽毒素为靶点，通过我们的“纯翻译展示”技术，将生产出抗肽和蛋白酶、膜渗透的n -甲基肽类似配体。这项技术允许合成天然和非天然氨基酸的聚合物，并且凭借遗传编码，筛选比化学合成文库多数百万倍的聚合物变体。因此，我们的库将在结构上更加多样化，并将包含更强大的绑定器。

DESCRIPTION (provided by applicant): The need for a rapid response to new bioterror threats and emerging infectious diseases is frustrated by the slow pace of drug discovery. We hypothesize that the protein synthetic machinery can be engineered for the rapid evolution in vitro of high-affinity peptide and peptide analog inhibitors of any target molecule. The goal here is to develop drug candidates for disseminated anthrax, a disease incurable with antibiotics. With anthrax toxin as the target, both peptide and protease-resistant, membrane-permeable, N-methyl-peptide analog ligands will be produced by our "pure translation display" technology. This technology allows synthesis of polymers of both natural and unnatural amino acids and, by virtue of genetic encoding, the screening of millions of times more polymer variants than chemically-synthesized libraries. Thus, our libraries will be much more diverse in structure and will contain binders that are much more potent. Specific Aim 1. To test the hypothesis that pure translation display with natural aminoacyl-tRNA substrates ill yield higher-affinity peptide binders for anthrax toxin's heptameric protective antigen fragment than prior methods. Specific Aim 2. To test the hypothesis that pure translation display with N-methylated arhinoacyl-tRNA substrates will yield binders for anthrax toxin that are resistant to proteases. Specific Aim 3. To test the hypothesis that the binders inhibit anthrax toxin function and have pharmacologically-desirable properties. Peptide and N-methyl-peptide analog binders will be tested in cellular assays of toxin function and for transport across Caco cells. Relevance of this research to public health. Anthrax is one of the biological weapons most likely to be used with devastating effect on a large population, and improved prophylactic and treatment options are urgently needed. Work in this proposal should yield several compounds that will have come a long way towards candidates suitable for testing for clinical use, either as injectables or pills. Furthermore, the technology developed will have potential utility in diagnosis, target validation and treatment of any disease.