rePPI-i: A system for the rapid continuous evolution of protein-protein interaction inhibitors

项目摘要 该项目的长期目标是开发一种快速、经济、有效的进化 用于创建靶蛋白-蛋白相互作用（PPI）的选择性抑制剂的平台。的 癌症系统生物学方法的出现揭示了过多的PPI中心， 对于癌症的获得、维持和免疫逃避至关重要。然而，破坏这些 具有药效团的PPI，无论是用于治疗还是用于基础科学发现， 具有挑战性，在很大程度上是由于缺乏选择功能活动的方法， 分子文库我们新提出的系统，“PPI抑制剂的快速进化”（rePPI-i），将 协同两种最近开发技术：1）噬菌体辅助连续进化（PACE）， 和2）活性响应RNA聚合酶（AR），以解决这一长期存在的问题。在 在rePPI-i系统中，噬菌体将携带一个进化的遗传编码的 噬菌体的生命周期将取决于那些破坏靶PPI的先导物 选择性地超过脱靶PPI。因此，在几天之内，数十亿个目标可以 经过数百轮的快速进化由于AR PPI的多功能性 检测系统，rePPI-i将能够从目标识别移动到PPI库 抑制剂在几周内，大大加快了药物发现过程。此外，委员会认为， PPI通常被认为是传统药理学方法的“不可用药”靶点， 困难在于：1）破坏通常广泛的大分子界面;和2）限制 分子库和筛选方法目前用于癌症靶向运动。 由于快速、持续进化的力量，这种范式将被rePPI-i打破， 从而产生高度优化的基于肽的抑制剂分子。制定和 验证这种新的方法，我们将发展线性和环状肽，以及小 结构蛋白，以破坏c-Myc/Max相互作用和BCL-2家族蛋白Mcl-1， 都是需要治疗的有效致癌靶点。我们的技术将带来一个新的 除了发现PPI抑制剂的药理学发展之外， 这两个重要的癌症靶点的治疗线索。rePPI-i不仅有潜力 迅速加速PPI抑制剂药物的发现，并开辟以前“无法治疗”的目标， 药理学干预，而且还降低了靶向相互作用的“活化屏障”， 使研究人员能够探索更多的干预措施。一旦优化，rePPI-i将是简单的， 并且用于感兴趣的新靶点的成本低廉，使药物发现过程民主化 并允许癌症研究人员更容易地开发新靶点的治疗线索。

PROJECT SUMMARY The long-term goal of this project is to develop a rapid, cost-effective, efficacious evolution platform for the creation of selective inhibitors of target protein-protein interactions (PPIs). The emergence of cancer systems biology approaches has revealed a plethora of PPI hubs that are critical for cancer acquisition, maintenance, and immune evasion. However, disrupting these PPIs with pharmacophores, either therapeutically or for basic science discovery, has been challenging, in large part due to a lack of methods to select for functional activity and inadequate molecular libraries. Our new proposed system, “rapid evolution of PPI inhibitors” (rePPI-i), will synergize two recently developed technologies: 1) phage-assisted continuous evolution (PACE), and 2) activity-responsive RNA polymerases (ARs), to solve this long-standing problem. In the rePPI-i system, bacteriophage will carry an evolving population of genetically-encoded therapeutic leads, and the phage life cycle will depend on those leads disrupting a target PPI selectively over an off-target PPI. Therefore, in a matter of days, billions of targets can be screened through hundreds of rounds of rapid evolution. Due to the versatility of the AR PPI detection system, rePPI-i will be capable of moving from target identification to a library of PPI inhibitors in a matter of weeks, dramatically accelerating the drug discovery process. Moreover, PPIs are often considered “undruggable” targets with traditional pharmacological approaches due to the difficulty in: 1) disrupting the often extensive macromolecular interfaces; and 2) the limits of molecular libraries and screening approaches currently utilized in cancer target campaigns. This paradigm will be broken by rePPI-i due to the power of rapid, continuous evolution and the resultant ability to generate highly optimized peptide-based inhibitor molecules. To develop and validate this novel approach, we will evolve both linear and cyclic peptides, as well as small structured proteins, to disrupt the c-Myc/Max interaction and the BCL-2 family protein Mcl-1, both validated oncogenic targets in need of therapeutics. Our technology will result in a new paradigm for pharmacological development of PPI inhibitors in addition to discovering therapeutic leads for these two important cancer targets. rePPI-i has the potential to not only rapidly accelerate PPI inhibitor drug discovery and open up previously “undruggable” targets to pharmacological intervention, but also lowers the “activation barrier” to targeting interactions, allowing researchers to explore more interventions. Once optimized, rePPI-i will be both simple and inexpensive to employ for new targets of interest, democratizing the drug discovery process and allowing cancer researchers to more readily develop therapeutic leads for novel targets.