Fully Synthetic Prodiginines as Probes of Protein-Protein Interactions Regulating

描述(由申请人提供)：在癌细胞中重建细胞凋亡为靶向、无毒治疗提供了相当大的希望。我们之前通过开发“Smac模拟物”证明了这一点，Smac模拟物是一种小分子，它可以破坏蛋白质之间的相互作用，从而抑制caspase的正常激活。自那以后，来自几个组织的Smac模拟药物已进入人体临床试验。这些实验仍在进行中。在这个提案中，我们概述了针对第二组蛋白质-蛋白质相互作用的新化学，这些蛋白质-蛋白质相互作用可以阻止癌症中的细胞凋亡并导致耐药性。BCL-2家族蛋白门控线粒体外膜通透性，调节正常细胞死亡所必需的细胞因子的释放。抗凋亡蛋白Bcl2的过度表达在化疗和放射耐药癌症中很常见。因此，作为药物开发的靶点，Bcl2系统受到了广泛的研究。这些努力中最先进的已经进入临床阶段。然而，尽管这一机制有巨大的希望，但意想不到的靶外效应，特别是基于Mcl-1的耐药性正在限制进展。需要具有更好轮廓的新化合物。ANSA桥联的前列腺素是由细菌产生的细胞毒性色素。其中一种天然产物，链霉菌红素B，被Shore鉴定为一种在细胞培养中促进细胞凋亡的分子--据报道是通过与Bcl-2蛋白相互作用来实现的。这项观察为奥布克拉克斯的开发奠定了基础，奥布克拉克斯是一种合成的前列腺素，目前正在进行临床试验，用于治疗淋巴细胞性白血病。Obatoclax被描述为一种针对BH3螺旋结合表面的PAN Bcl-2拮抗剂，包括Mcl-1上的Noxa结合位点。数据是有希望的，但受到选择性的影响。斜纹夜蛾中的三吡咯类发色团在金属离子存在的情况下产生ROS，并能通过跨膜离子转运中和pH梯度。这两种作用都会损害正常细胞。我们建议研究链霉菌红素B的新的吡咯罗芬变种，这些变种缺乏Obtoclax的结构敏感性。在我们最近完成的亲红素全合成的基础上，我们概述了由海洋细菌产生的新分离的抗白血病前列腺素Marineosin A和Marineosin B的合成方法。在与麦吉尔大学的戈登·肖尔的合作中，我们发现了在我们的玫瑰亲油素合成中的中间体，它在体外阻止了t-Bid依赖的Mcl-1功能。我们建议在这些结果的基础上，沿着几条路线推进我们的合成化学。基于计算模型，我们相信所讨论的天然产物的ANSA桥联杂环核心可以支撑一类新的螺旋多肽。描述了检验这一假说的实验。其中包括选择性地针对Mcl-1/Noxa、Bcl-xl/Bim和p53/MDM2相互作用的定制合成。最后，作为在细胞培养和体内鉴定非基于Bcl2的效应的手段，我们建议使用基于亲和力的生化方法和最新的功能基因组筛选来寻找这一结构类别的额外靶点。

DESCRIPTION (provided by applicant): Reestablishing apoptosis in cancer cells holds considerable promise for targeted, non-toxic therapy. We demonstrated this previously with the development of 'smac mimetics', small molecules that disrupt protein- protein interactions that suppress proper caspase activation. Smac mimetics from several organizations have since advanced to human clinical trials. These experiments remain ongoing. In this proposal we outline new chemistry aimed at a second set of protein-protein interactions that block apoptosis in cancer and contribute to drug resistance. Bcl-2 family proteins gate mitochondrial outer membrane permeability and regulate cytoplasmic release of factors essential for normal cell death. Over-expression of anti-apoptotic Bcl-2 proteins is common in chemotherapy and radiation resistant cancers. As a result, the Bcl-2 system has been intensely studied as a target for drug development. The most advanced of these efforts have reached clinic stages. However, despite tremendous promise for the mechanism, unanticipated off-target effects and, notably, Mcl-1 based resistance to agents targeting Bcl-XL are limiting progress. New compounds with superior profiles are needed. Ansa-bridged prodiginines are cytotoxic pigments produced by bacteria. One of these natural products, streptorubin B, was identified by Shore as a molecule that potentiates apoptosis in cell culture - reportedly through interactions with Bcl-2 proteins. This observation seeded development of obatoclax, a synthetic prodiginine now in clinical trials as therapy for lymphocytic leukemia. Obatoclax is described as a pan Bcl-2 antagonist that targets BH3 helix-binding surfaces, including the Noxa binding site on Mcl-1. Data is promising but tempered by selectivity. The tri-pyrrolic chromophore in obatoclax generates ROS in the presence of metal ions and can neutralize pH gradients via transmembrane ion transport. Both effects damage normal cells. We propose to study new pyrrolophane variants of streptorubin B that lack the structural liabilities of obatoclax. Based on our recently completed total synthesis f roseophilin, we outline new methods to synthesize marineosins A and B - newly isolated anti-leukemic prodiginines produced by marine bacteria. In collaboration with Gordon Shore at McGill University, we have discovered intermediates in our roseophilin synthesis that block t-Bid dependent Mcl-1 functions in vitro. We propose to build on these results by advancing our synthetic chemistry along several lines. Based on computational models, we believe the ansa- bridged heterocyclic cores of the natural products in question can scaffold a new class of ¿-helix peptidomimetics. Experiments to test this hypothesis are described. These include custom syntheses of ligands targeting the Mcl-1/Noxa, Bcl-XL /Bim and p53/mdm2 interactions selectively. Lastly, as means to identify non Bcl-2 based effects in cell culture and in vivo, we propose to search for additional targets of this structural class using affinity based biochemical methods and state-of-the-art functional genomic screens.