Tuning aminoglycosides for treatment of genetic diseases

描述（由申请人提供）：大量的人类遗传疾病是由于突变导致突变基因编码的蛋白质合成过早终止而导致的。目前，已知有数百种这样的无义突变，其中一些被证明可以解释某些致命疾病，包括囊性纤维化（CF）、杜氏肌营养不良症（DMD）、泰-萨克斯病等。对于其中的许多疾病，目前还没有有效的治疗方法。在过去的几年里，研究表明，一些氨基糖苷类抗生素（包括庆大霉素）有能力让哺乳动物核糖体选择性地读取假停止信号，而不是正常的终止信号，并产生全长功能蛋白。然而，这些药物对人体的高毒性限制了它们的治疗用途。本研究的主要目的是开发具有有效终止抑制活性的新型氨基糖苷，同时降低对哺乳动物细胞的毒性。迄今为止，氨基糖苷类药物的临床应用仅限于抗菌药物，尚未对其作为终止密码子穿透诱导剂的活性进行优化。为了达到这些目的，几个不同的实验室正在进行合作，并结合许多互补学科的专业知识来合成和阐明所设计药物的结构-活性-毒性关系。该提议背后的假设是将导致毒性的氨基糖苷结构元素与诱导无义抑制所需的元素分开：设计的结构对细胞质rRNA A位点具有广泛的特异性和选择性，可以减小功能剂量范围，从而降低预期的毒性，包括对线粒体蛋白质合成机制的有害影响，使其成为治疗人类遗传疾病的潜在药物。我们的假设是基于最近的一系列观察，我们已经表明，通过降低对原核核糖体的特异性，从而消除氨基糖苷的“天然”抗菌活性，我们减少了它们对真核线粒体蛋白质合成机制的作用，从而显著降低了它们对人类的毒性作用。预计该研究将带来实质性的治疗和经济效益。本文设计的结构制备简单，初步试验已经发现了一些具有较低毒性和更强的通读效力的变体，可以在体外和体内从突变基因中恢复CFTR蛋白的功能，包括与庆大霉素密切预测人类CF受试者结果的模型。实现这个项目的目标将为发现可能作为新药的新结构提供必要的知识和工具。

DESCRIPTION (provided by applicant): A large number of human genetic diseases result from mutations that cause premature termination of the synthesis of proteins encoded by mutant genes. Currently, hundreds of such nonsense mutations are known, and several where shown to account for certain cases of fatal diseases, including cystic fibrosis (CF), Duchenne muscular dystrophy (DMD), Tay-Sachs, and more. For many of those diseases there is presently no effective treatment. In the last several years, it was shown that some aminoglycoside antibiotics (including gentamicin) have the ability to allow the mammalian ribosome to selectively read past a false-stop signal, but not a normal termination signal, and generate full-length functional proteins. However, high toxicity of these drugs in humans limits their therapeutic use. The main objective of this research is to develop novel aminoglycosides that will have efficient termination suppression activity, and at the same time will have reduced toxicity against mammalian cells. To date, the clinical application of aminoglycosides is limited to their use as antibacterial drugs, and no efforts have been made to optimize their activity as stop codon read-through inducers. Toward these ends, a collaborative effort is under way between several different laboratories and combines expertise of number of complementary disciplines to synthesize and elucidate the structure-activity- toxicity relationships of the designed drugs. The hypothesis behind the proposal is to separate the elements of the aminoglycosides structures that cause toxicity from those that are required for inducing nonsense suppression: designed structures exhibiting extensive specificity and selectivity for the cytoplasmic rRNA A site can decrease the functional dosing ranges and subsequently decrease the anticipated toxicity, including deleterious effects on mitochondrial protein synthesis machinery, making them potential drugs for the treatments of human genetic disorders. Our hypothesis is based on a series of recent observations in which we have shown that by reducing the specificity to prokaryotic ribosome and as such wiping away from aminoglycosides their "natural" antibacterial activity we reduce their action on eukaryotic mitochondrial protein synthesis machinery and as such significantly reduce their toxic effects on humans. Substantial therapeutic and economic benefits are anticipated from this study. The designed structures presented here are simple for preparation and the preliminary tests already discovered some variants with lower toxicity and greater read-through efficacy to restore functional CFTR protein from the mutant gene both in vitro and in vivo, including models closely predictive of results with human CF subjects, than those of gentamicin. Fulfillment of the goals of this project will provide the necessary knowledge and tools for uncovering new structures, which may act as novel drugs. PUBLIC HEALTH RELEVANCE: Many human genetic diseases including cystic fibrosis, Duchenne muscular dystrophy, Usher syndrome, Hurler syndrome and numerous types of cancer are caused by nonsense mutations. According to the Human Gene Mutation Database, nonsense mutations represent about 12% of all mutations reported, and for many of those diseases there is presently no effective treatment. The proposed research aims to develop potent aminoglycosides derivatives with high potential for immediate therapeutic application for treatment of genetic diseases caused by nonsense mutations.