Tuning aminoglycosides for treatment of genetic diseases

描述（由申请人提供）：大量人类遗传性疾病是由突变引起的，这些突变导致突变基因编码的蛋白质合成提前终止。目前，已知有数百种这样的无义突变，其中几种被证明是某些致命疾病的原因，包括囊性纤维化（CF），杜氏肌营养不良症（DMD），泰-萨克斯等。对于其中许多疾病，目前没有有效的治疗方法。在过去的几年中，研究表明，一些氨基糖苷类抗生素（包括庆大霉素）具有允许哺乳动物核糖体选择性读取假终止信号而不是正常终止信号的能力，并产生全长功能蛋白。然而，这些药物在人体中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新的氨基糖苷类，将具有有效的终止抑制活性，并在同一时间将有减少对哺乳动物细胞的毒性。迄今为止，氨基糖苷类的临床应用仅限于其作为抗菌药物的用途，并且没有努力优化其作为终止密码子通读诱导剂的活性。为了实现这些目标，几个不同的实验室之间正在进行合作，并结合了一些互补学科的专业知识，以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引起毒性的氨基糖苷类结构的元素与诱导无义抑制所需的元素分开：对细胞质rRNA A位点表现出广泛特异性和选择性的设计结构可以降低功能剂量范围并随后降低预期的毒性，包括对线粒体蛋白质合成机制的有害作用，使它们成为治疗人类遗传疾病的潜在药物。我们的假设是基于一系列最近的观察，其中我们已经表明，通过降低对原核核糖体的特异性，从而消除氨基糖苷类的“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。预计本研究将带来实质性的治疗和经济效益。这里提出的设计结构易于制备，并且初步测试已经发现了一些具有较低毒性和更大通读效力的变体，以在体外和体内从突变基因恢复功能性CFTR蛋白，包括与庆大霉素相比，与人类CF受试者密切预测结果的模型。该项目目标的实现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.