Development of a mouse model of tuberculosis that generates human-like pathology

摘要在过去的四十年中，虽然很少有新的结核病药物被引入 多年来，与结核病治疗发展相关的关键问题在整个领域得到了很好的阐述， 在生物医学界正以极大的活力进行研究。我们能找到一种方法来缩短治疗时间吗 从目前的六个月？哪些新的作用机制将支持完全绝育， 与现有的标准治疗相结合？持续存在的种群是否通过 具体治疗方案？ 为了在临床上取得成功，首先我们必须在临床前通过使用 预测动物模型。实验性结核病治疗的疗效评价通常是 使用小鼠模型进行，因为小鼠的尺寸相对较小且成本较低。然而，许多老鼠 模型不能再现地发展出类似人类的病理学，特别是缺氧性坏死性肉芽肿， 因此对于某些化合物可能具有有限的预测值。小鼠模型， 病理学可能遭受限制显著发病率所需的实质异质性或复杂方案。 我们最近的工作已经克服了这些重大的局限性，在这里，我们建议完善，验证和 描述了一种创新的结核病小鼠模型，该模型更能预测患者的结局。 我们已经完成了一系列在小鼠身上的研究，这些研究加强了我们推进更具预测性的模型的能力 结核病，一个始终产生人类一样的病理形式的缺氧坏死性肉芽肿。这 一个新的模型建立在Nos 2-/-小鼠缺氧性肉芽肿的研究基础上，但补充说， 通过简化方案并利用天然气溶胶途径感染小鼠， 部分减毒结核分枝杆菌（Mtb）菌株R1 Rv。我们将这一新颖的模型与 特异性标记药物和免疫耐受性Mtb细胞的荧光报告菌株。当一起部署时 为了支持对潜在结核病药物治疗方案的药理学评价，这两种工具可以提供独特的 深入了解为什么一些治疗方法可以很好地缩短患者的化疗时间， 其他国家则不会这样做。在这个建议中，我们寻求建立模型的预测值，以确定药物 疗效反应，并揭示耐药结核分枝杆菌生物学的新见解。 在前两个特定目标中，我们试图优化新的小鼠模型，并验证它是一个更严格的模型。 在体内抗结核药物疗效的措施，并在第三个目标，我们采用该模型来确定生态位 被耐药细胞占据，以更好地了解它们潜在的脆弱性。完成三个 该提案的目的是提供更具预测性和良好表征的Mtb小鼠模型 这种新的感染可能成为结核病药物小动物试验的新标准。

ABSTRACT Though very few new tuberculosis (TB) drugs have been introduced over the past forty years, the pivotal questions relevant to TB therapeutic development are well articulated across the field and are being pursued with great vigor in the biomedical community. Can we find a way to shorten treatment for from the current six months? Which novel mechanisms of action will support complete sterilization when combined with existing standard of care treatment? Are persister populations completely eliminated through a specific treatment regimen? In order to succeed in the clinic, first we must address these questions preclinically through the use of predictive animal models. Evaluation of the efficacy of experimental tuberculosis therapeutics is commonly performed using mouse models, due to the relatively small size and low cost of mice. However, many mouse models fail to reproducibly develop human-like pathology, specifically hypoxic necrotic granulomas, and thereby may have limited predictive value for certain compounds. Mouse models which do develop human-like pathology may suffer from substantial heterogeneity or complex protocols required to limit significant morbidity. Our recent work has overcome these significant limitations, and here we propose to refine, validate and characterize an innovative mouse model of TB that is more predictive of outcomes in patients. We have completed a series of studies in mice that strengthen our ability to advance a more predictive model of TB, one that consistently produces human-like pathology in the form of hypoxic necrotic granulomas. This new model builds on work that demonstrated the development of hypoxic granulomas in Nos2-/-mice, but adds significant value by simplifying the protocol and utilizing the natural aerosol route to infect mice with the partially attenuated Mycobacterium tuberculosis (Mtb) strain R1Rv. We have paired this novel model with fluorescent reporter strains that specifically mark drug and immune tolerant Mtb cells. When deployed together to support pharmacological evaluation of potential TB drug regimens, these two tools can provide unique insights into why some therapeutic approaches may work well to shorten chemotherapy in patients, while others will not do so. In this proposal, we seek to establish the predictive value of the model to determine drug efficacy responses, and to reveal new insights into the biology of drug-tolerant Mtb. In the first two Specific Aims we seek to optimize the novel mouse model and validate it as a more rigorous measure of in vivo anti-tubercular drug efficacy, and in the third Aim we employ the model to identify the niche occupied by drug tolerant cells, to better understand their potential vulnerabilities. Completion of the three Aims of this proposal will deliver a more predictive and well-characterized mouse model of Mtb infection that will have the potential to become a new standard for small animal testing of TB drugs.