Pre-clinical Development of an Advanced Genexpert Test for Drug Resistant Mtb

先进的 Genexpert 耐药 Mtb 检测的临床前开发

• 批准号: 8691594
• 负责人: David Alland
• 金额: $ 154.98万
• 依托单位: RBHS-NEW JERSEY MEDICAL SCHOOL
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-04-01 至 2019-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8691594
• 关键词: Accounting; Advanced Development; Algorithms; Amikacin; Aminoglycoside Antibiotics; Aminoglycosides; Antitubercular Agents; Automation; Biological Assay; Chemistry; Clinical; Clinical Trials; Color; Complement; Computer software; Country; Decision Making; Detection; Development; Disease; Drug Resistant Tuberculosis; Drug resistance; Drug resistance in tuberculosis; Dyes; Extreme drug resistant tuberculosis; Fluoroquinolones; Future; Genetic; Grant; Hand; Incidence; Infection Control; Institution; Kanamycin; Laboratories; Methods; Modification; Molecular; Moxifloxacin; Multi-Drug Resistance; Mutation; Mycobacterium tuberculosis; Patients; Performance; Pharmaceutical Preparations; Phase; Predisposition; Preparation; Procedures; Quality Control; Reagent; Reporting; Resistance; Rifampicin resistance; Rifampin; Risk; Robotics; Sampling; Sentinel; Sputum; Temperature; Testing; Therapeutic; Time; Tracer; Tuberculosis; United States National Institutes of Health; Validation; Work; assay development; base; cost effective; expectation; fluoroquinolone resistance; improved; inhibitor/antagonist; innovation; internal control; isoniazid; killings; manufacturing process; melting; mortality; novel; pathogen; pre-clinical; preclinical study; prevent; public health relevance; reagent testing; research and development; research clinical testing; resistance mutation; tuberculosis drugs

DESCRIPTION (provided by applicant): The need for rapid and sensitive methods to detect drug-resistant tuberculosis (TB) has never been greater. The incidence of multi-drug-resistant (MDR) and extensively-drug resistant (XDR) TB is increasing at an alarming rate. Even in the absence of MDR and XDR TB, rapid Fluoroquinolone (FQ) resistance testing will be increasingly needed for patients who are treated with the Moxifloxacin-based regimes that are expected to partially replace Isoniazid (INH)-based treatment in the future. Our group developed the Xpert MTB/RIF (Xpert) assay, the first ever self-contained, near-patient assay that can perform TB detection and also detect resistance to the drug Rifampicin (RIF). This test is transforming TB detection worldwide by making TB detection simple and rapid. However in regards to drug resistance detection, the Xpert assay is limited to identifying patients who potentially have MDR by using RIF-resistance as a surrogate. The Xpert assay cannot tell whether a patient with RIF resistance remains INH-susceptible, and thus could still be treated with an INH-containing regime. Nor can it identify which RIF-resistant patients can be treated with a FQ and/or aminoglycoside as opposed to those who have XDR TB. Thus, patients are at risk for being either over treated or undertreated, with potentially serious consequences. Furthermore, patients with undiagnosed XDR can remain infectious for prolonged periods, further spreading their disease. Here, we propose to combine highly innovative 10-color real-time PCR detection (that makes use of novel large Stokes-shift dyes), innovative three phase PCR fluidics, sloppy molecular beacon probes, the existing GeneXpert platform and basic Xpert assay cartridge to create a new assay that identifies resistance to INH, the FQs, amikacin and kanamycin, important types of resistance not detected by the Xpert assay. This new "Xtend-XDR" assay will complement the current Xpert assay, making it possible to select appropriate treatment (and infection control) approaches in patients identified with TB. The Xtend-XDR assay will use the same testing procedures and testing platform as the Xpert, and largely be restricted to patients who already test TB positive (and RIF resistant) in a primary Xpert test. These features will make it highly cost effective. We have developed proof of principal for all essential assay components. In the current proposal, we will finalize assay development, and create manufacturing processes, quality control, and assay software. Finally, we will perform the analytic and pre-clinical studies needed to prepare for clinical trials. These activities are exactly the type requested in the current partnership RFA. Our work will be performed in the following specific aims: 1. Finalize all assay parameters and internal control components in wet format. 2. Adapt assay for robotic assembly and modify assembly line for cartridge needs. 3. Develop analytic software and a simple interface for clinical operators. 4. Perform analytic and pre-clinical studies in preparation for clinical trials.

描述（由申请人提供）：对快速、灵敏的方法来检测耐药结核病（TB）的需求从未如此强烈。耐多药（MDR）和广泛耐药（XDR）结核病的发病率正在以惊人的速度增加。即使不存在耐多药和广泛耐药结核病，接受基于莫西沙星的方案治疗的患者也将越来越需要快速氟喹诺酮（FQ）耐药性检测，预计未来该方案将部分取代基于异烟肼（INH）的治疗。 我们的小组开发了 Xpert MTB/RIF (Xpert) 检测，这是第一个独立的、靠近患者的检测，可以进行结核病检测，还可以检测对药物利福平 (RIF) 的耐药性。该测试使结核病检测变得简单、快速，正在改变全世界的结核病检测。然而，在耐药性检测方面，Xpert 检测仅限于通过使用 RIF 耐药性作为替代来识别可能患有 MDR 的患者。 Xpert 检测无法判断 RIF 耐药的患者是否仍然对 INH 敏感，因此仍然可以接受含有 INH 的治疗方案。它也无法确定哪些 RIF 耐药患者可以接受 FQ 和/或氨基糖苷类药物治疗，而不是那些患有广泛耐药结核病的患者。因此，患者面临着过度治疗或治疗不足的风险，并可能造成严重后果。此外，未经诊断的 XDR 患者可能会长时间保持传染性，从而进一步传播疾病。 在这里，我们建议将高度创新的 10 色实时 PCR 检测（利用新型大斯托克斯位移染料）、创新的三相 PCR 流体、草率分子信标探针、现有的 GeneXpert 平台和基本的 Xpert 检测盒结合起来，以创建一种新的检测方法，可识别对 INH、FQ、阿米卡星和卡那霉素的耐药性，这是 Xpert 检测未检测到的重要耐药类型。这种新的“Xtend-XDR”检测将补充当前的 Xpert 检测，使得为结核病患者选择适当的治疗（和感染控制）方法成为可能。 Xtend-XDR 检测将使用与 Xpert 相同的检测程序和检测平台，并且主要仅限于在主要 Xpert 检测中已检测出 TB 阳性（和 RIF 耐药）的患者。这些功能将使其具有极高的成本效益。 我们已经为所有重要的检测组件开发了原理证明。在当前的提案中，我们将完成检测开发，并创建制造工艺、质量控制和检测软件。最后，我们将进行为临床试验做准备所需的分析和临床前研究。这些活动正是当前合作关系 RFA 中所要求的类型。我们的工作将按照以下具体目标进行： 1. 以湿法形式最终确定所有测定参数和内部控制组件。 2. 调整检测以适应机器人组装，并根据卡盒需求修改组装线。 3. 为临床操作人员开发分析软件和简单的界面。 4. 进行分析和临床前研究，为临床试验做准备。