Experimental Animal Models of TB: Chemotherapeutics and Imaging

结核病实验动物模型：化疗和影像学

• 批准号: 10692048
• 负责人: Clifton Barry
• 金额: $ 153.83万
• 依托单位: NATIONAL INSTITUTE OF ALLERGY AND INFECTIOUS DISEASES
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10692048
• 关键词: Advanced Development; Animal Model; Animals; Antibiotics; Benchmarking; Callithrix; Cell Wall; Clinical Trials; Computed Tomography Scanners; Data; Deoxyglucose; Development; Disease; Disease Progression; Dose; Drug Combinations; Drug Kinetics; Emission-Computed Tomography; Enzymes; Eukaryotic Cell; Experimental Animal Model; Extracellular Matrix; Extreme drug resistant tuberculosis; Failure; Foundations; Functional Imaging; Granuloma; Human; Image; Imaging Techniques; Individual; Infection; Inflammation; Kinetics; Knowledge; Label; Lesion; Linezolid; Lung; Measures; Metabolism; Methods; Microbiology; Modeling; Modernization; Monitor; Moxifloxacin; Mus; Mycobacterium tuberculosis; Mycobacterium tuberculosis antigens; Necrotic Lesion; Neutrophil Infiltration; Nitroimidazoles; Organ; Organ Culture Techniques; Oryctolagus cuniculus; Outcome; Oxazolidinones; Pathology; Patients; Penetration; Pharmaceutical Preparations; Positron-Emission Tomography; Pre-Clinical Model; Predisposition; Pyrazinamide; RNA analysis; Randomized; Regimen; Regulation; Residual state; Residual volume; Resolution; Rodent; Scientist; Signal Transduction; Site; South Africa; Spatial Distribution; Structure of parenchyma of lung; System; Techniques; Technology; Testing; Therapeutic; Therapeutic Effect; Time; Treatment Protocols; Trehalose; Tuberculosis; Work; X-Ray Computed Tomography; bacterial metabolism; bactericide; base; black hole; chemotherapy; cytokine; drug candidate; drug distribution; drug efficacy; drug testing; experience; experimental study; extensive drug resistance; fluorodeoxyglucose; human data; improved; in vivo; in vivo evaluation; industry partner; inflammatory marker; inhibitor; nonhuman primate; novel; novel therapeutics; phase II trial; predictive modeling; programs; quinoline; radiological imaging; radiotracer; rational design; response; small molecule; success; treatment effect; treatment group; treatment response; trend; tuberculosis chemotherapy; tuberculosis drugs; tuberculosis treatment

This project encompasses approaches to understand how current anti-tubercular chemotherapy works using the most modern technologies and to develop new and improved therapies and therapeutic approaches. Individual projects within this framework are (1) developing structural and functional imaging techniques using PET/CT for use in live, M. tuberculosis (Mtb) infected animals, (2) development of advanced animal models for predicting drug efficacy under conditions that exactly mimic those experienced by TB patients, (3) understanding the activity of various drugs in animal models of tuberculosis therapy, (4) correlating responses seen in animal models with the pathology and response to therapy observed in human TB, and (5) developing techniques for assessing drug distribution, penetration, and pharmacokinetics in vivo. In 2021 we developed moderate capacity to image mice in the Mediso PET/CT scanner using a chamber that holds 4 mice per imaging session. In 2022, we used this capacity to characterize murine lesions that were stimulated by cytokine treatment to recruit neutrophils and test their susceptibility to pyrazinamide. Most of our PET/CT studies have used 18F-2-fluoro-2-deoxyglucose (FDG) to image the metabolism of the eukaryotic cells in TB lesions in our animal models of tuberculosis. Yet, we would prefer a small molecule that could be used to label Mtb in vivo endogenously as a PET radiotracer. For several years we have focused employing Mtb antigen 85 enzymes that are expressed on the exterior of Mtb cell walls to incorporate exogenous-provided 18F 2-deoxy trehalose (FDT) as either the mono- or dimycolate in or near the cell wall. Use of FDT in the imaging of Mtb in non-human primates, successfully allows the specific imaging of TB-associated lesions and to monitor the effects of treatment in marmosets. This is a promising sign that the FDT will be able to give a more accurate indication of treatment success or failure compared to FDG. We are continuing to develop this probe in the marmoset. In the past we explored if the marmoset model accurately reflects the response to treatment by providing standard TB treatment (RIF, INH, PZA, and EMB) to infected symptomatic marmosets and demonstrated that marmosets show similar treatment results as humans. As a counterpart to an early bactericidal activity (EBA) and paired PET/CT clinical trial, NexGen EBA conducted in South Africa; NCT02371681, in 2018-2020 we have replicated the treatment groups and observations in randomized Mtb infected marmosets. In that study, the standard regimen was deconstructed, and each drug was administered by itself or in pair-wise combinations to measure the effect of the drugs on the microbiological and radiographic markers as well as two 4 drug regimens HRZE and MRZE where M is moxifloxacin. In 2022, we determined that there were significant differences in treatment response between the MRZE and HRZE groups based on the lesion volumetric disease in both the CT and FDG-PET data at 2 months, but not 2 weeks (the duration of the typical EBA study). When the PET/CT image scoring system was applied, we found that the pathology in HRZE-treated marmosets resolved significantly faster and the increased resolution rate led to a significantly different residual volume of disease. In 2021-2022 we have deconstructed the Nix-TB regimen of Bedaquiline (BDQ, B), Pretomenid (Pa) and LZD (L) with all the drug combinations in roughly 35 marmosets. Previously we, found that BDQ accumulated significantly in both marmoset (and rabbit) TB lesions at steady state, including in cavity caseum, possibly explaining its outstanding efficacy in hard-to-treat patients with XDR TB. The lower more human-like dose of BDQ modeled by collaborators gave a very good approximation of the Cavg seen in humans but was less effective in sterilizing lung tissue than our original dose indicating that a higher BDQ does in humans might be even more effective if it was tolerable. Each of the individual agents are active against MTB in the marmoset although BDQ is more active alone that LZD. We have found that the BPaL regimen is very active in the marmoset but no more so than BPa. In fact, adding LZD to Pa appears antagonistic by some measures and adding LZD to BDQ is like BDQ alone although there is a trend toward fewer necrotic lesions in the BL combination. The 3-drug regimen is similar to HRZE in activity in the marmoset, but not superior to it. As with the BPaL regimen, we have investigated another drug combination that is likely headed for a human phase 2 trial this summer. The combination of Delamanid (D), BDQ (B), and an DprE2 inhibitor OPC-167832 (O) was tested in the marmosets in single, double and the triple drug combination. Each individual agent was highly effective promoting survival, reducing pathology, inflammation, and bacterial burdens in the lung and extrapulmonary organs. The combination of DO reduces residual pathology significantly more than either agent alone and the 3-drug combination DBO is significantly better than the individual agents in reducing bacterial burden and organ culture positivity rates. The analysis of the two-agent combinations and pathology measures are ongoing. We continue to study the oxazolidinone (OXA) antibiotics such as linezolid (LZD) which have shown significant therapeutic effects in patients with extensively drug-resistant (XDR) TB despite modest effects in rodents. One of these new amino-OXAs causes a significant rapid reduction in lesion pathology and inflammation compared with LZD as measured by FDG PET/CT. This activity was associated with lesion type and physical distribution into the lesions. Together with the Gates Foundation's TB Drug Accelerator scientists we have engaged in developing novel OXAs that are TB-selective and less toxic than LZD. We continue to evaluate the PET/CT imaging data from marmosets from treated with other classes of antibiotics from partners in the TBDA program including diarylquinolines, quinolines, imidazopyridines, nitroimidazoles, among others. These classes of antibiotics are being explored as composing new regimens for treatment of MTB and understanding the specific contribution of each one to activity including consideration of spatial distribution and the kinetics of accumulation in lesions to avoid temporal and spatial black holes of monotherapy. With each new drug candidate, we test for in vivo efficacy with our academic and industry partners, we continue to assess the candidates penetration into granulomas and cavities in our model animals to correlate the information with any observed efficacy. We continue to explore host-directed therapy (HDT) as a method to increase drug efficacy by increasing agent delivery to the site of infection in the rabbit model of Mtb. In 2022 modeling results from the Savic lab, BDQ penetration is increased in fibrotic lesions at steady state in HDT-treated rabbits but not in other lesion types. These experiments have also been conducted with moxifloxacin, but the results of the modeling are not yet available. In host RNA analysis (2022) from the control and HDT treatment indicates that signaling is changed during the HDT treatment with significant changes in expression of extracellular matrix components as predicted as well as changes in regulation of signal transduction, cell communication and signaling. These data are being explored to see what effect the HDT treatment had on the TB lesions.

该项目包括了解当前抗结核化疗如何使用最现代的技术发挥作用以及开发新的和改进的疗法和治疗方法的方法。该框架内的各个项目包括 (1) 使用 PET/CT 开发结构和功能成像技术，用于感染结核分枝杆菌 (Mtb) 的活体动物，(2) 开发先进的动物模型，用于在完全模拟结核病患者经历的条件下预测药物疗效，(3) 了解各种药物在结核病治疗动物模型中的活性，(4) 将动物模型中观察到的反应与病理学和治疗反应相关联 在人类结核病中观察到，(5) 开发评估药物体内分布、渗透和药代动力学的技术。 2021 年，我们开发了在 Mediso PET/CT 扫描仪中对小鼠进行中等成像的能力，使用的腔室每次成像可容纳 4 只小鼠。 2022 年，我们利用这种能力来表征由细胞因子治疗刺激的小鼠病变，以招募中性粒细胞并测试它们对吡嗪酰胺的敏感性。 我们的大多数 PET/CT 研究均使用 18F-2-氟-2-脱氧葡萄糖 (FDG) 对结核病动物模型中结核病灶中的真核细胞的代谢进行成像。然而，我们更喜欢一种可以作为 PET 放射性示踪剂在体内内源性标记 Mtb 的小分子。多年来，我们一直致力于使用在 Mtb 细胞壁外部表达的 Mtb 抗原 85 酶，将外源提供的 18F 2-脱氧海藻糖 (FDT) 作为单霉菌酸盐或二霉菌酸盐掺入细胞壁内或细胞壁附近。在非人类灵长类动物中使用 FDT 进行 Mtb 成像，成功地实现了 TB 相关病变的特异性成像，并监测狨猴的治疗效果。这是一个有希望的迹象，表明与 FDG 相比，FDT 将能够更准确地指示治疗成功或失败。我们正在继续在狨猴身上开发这种探针。 过去，我们通过向受感染的症状狨猴提供标准结核病治疗（RIF、INH、PZA 和 EMB）来探索狨猴模型是否准确反映了对治疗的反应，并证明狨猴表现出与人类相似的治疗结果。作为早期杀菌活性 (EBA) 和配对 PET/CT 临床试验的对应项，NexGen EBA 在南非进行； NCT02371681，2018-2020 年，我们在随机感染 Mtb 的狨猴中重复了治疗组和观察结果。在该研究中，标准方案被解构，每种药物单独或成对组合给药，以测量药物对微生物和放射学标记物的影响，以及两种 4 药物方案 HRZE 和 MRZE，其中 M 是莫西沙星。 2022 年，我们根据 2 个月而非 2 周（典型 EBA 研究的持续时间）CT 和 FDG-PET 数据中的病灶体积疾病确定 MRZE 和 HRZE 组之间的治疗反应存在显着差异。当应用 PET/CT 图像评分系统时，我们发现 HRZE 处理的狨猴的病理学解决速度明显更快，并且解决率的提高导致疾病残留体积显着不同。 2021-2022 年，我们在大约 35 只狨猴中解构了 Bedaquiline (BDQ, B)、Pretomenid (Pa) 和 LZD (L) 的 Nix-TB 治疗方案以及所有药物组合。此前，我们发现 BDQ 在稳定状态下的狨猴（和兔子）结核病灶中显着积累，包括在干酪腔中，这可能解释了其在难以治疗的广泛耐药结核病患者中的出色疗效。合作者模拟的较低的、更接近人类的 BDQ 剂量与人类中观察到的 Cavg 非常接近，但在对肺组织进行消毒方面不如我们的原始剂量有效，这表明如果可以耐受，较高的 BDQ 对人类的作用可能会更有效。尽管 BDQ 单独使用时比 LZD 更有效，但每种药物都对狨猴中的 MTB 具有活性。我们发现 BPaL 方案在狨猴中非常活跃，但并不比 BPa 更有效。事实上，在某些措施中，将 LZD 添加到 Pa 中似乎是拮抗的，并且将 LZD 添加到 BDQ 中就像单独使用 BDQ 一样，尽管 BL 组合中有减少坏死病变的趋势。 3 种药物方案在狨猴中的活性与 HRZE 相似，但并不优于它。 与 BPaL 方案一样，我们研究了另一种药物组合，该药物组合可能会在今年夏天进行人体 2 期试验。 Delamanid (D)、BDQ (B) 和 DprE2 抑制剂 OPC-167832 (O) 的组合在狨猴中以单药、双药和三药组合进行了测试。每种药物都非常有效地促进生存，减少肺部和肺外器官的病理、炎症和细菌负担。 DO 组合比单独使用任一药物更能显着减少残留病理，并且 3 种药物组合 DBO 在降低细菌负荷和器官培养阳性率方面显着优于单独药物。对两种药物组合和病理学测量的分析正在进行中。 我们继续研究恶唑烷酮 (OXA) 抗生素，例如利奈唑胺 (LZD)，尽管对啮齿动物的影响不大，但它对广泛耐药 (XDR) 结核病患者显示出显着的治疗效果。通过 FDG PET/CT 测量，与 LZD 相比，这些新的氨基 OXA 之一可显着快速减少病变病理和炎症。 这种活性与病变类型和病变的物理分布有关。我们与盖茨基金会的结核病药物加速器科学家一起致力于开发新型 OXA，该药物具有结核病选择性且比 LZD 毒性更低。 我们继续评估接受 TBDA 计划合作伙伴提供的其他类别抗生素（包括二芳基喹啉、喹啉、咪唑并吡啶、硝基咪唑等）治疗的狨猴的 PET/CT 成像数据。正在探索这些类别的抗生素，以制定治疗 MTB 的新方案，并了解每种抗生素对活性的具体贡献，包括考虑空间分布和病灶中积累的动力学，以避免单一疗法的时空黑洞。对于每一种新候选药物，我们都会与学术和行业合作伙伴一起测试其体内功效，并继续评估候选药物对模型动物肉芽肿和空腔的渗透情况，将信息与任何观察到的功效相关联。 我们继续探索宿主定向治疗（HDT）作为一种通过增加结核分枝杆菌兔模型中药物递送至感染部位来提高药物疗效的方法。根据 Savic 实验室的 2022 年建模结果，在 HDT 治疗的兔子的稳定状态下，BDQ 渗透在纤维化病变中增加，但在其他病变类型中没有增加。这些实验也用莫西沙星进行，但建模结果尚未公布。在对对照和 HDT 治疗的宿主 RNA 分析 (2022) 中，信号传导在 HDT 治疗期间发生了变化，如预测的那样，细胞外基质成分的表达发生了显着变化，并且信号转导、细胞通讯和信号传导的调节也发生了变化。正在探索这些数据，以了解 HDT 治疗对结核病灶有何影响。