Enhancing Pediatric Diagnosis of Tuberculosis with FLOW Technology

摘要 虽然正确的诊断是有效治疗结核病的关键，但在资源不足的情况下，结核病检测存在特定的障碍。 设置，特别是对儿童。虽然有许多结核病诊断工具可用，但每种工具都存在严重的 缺点（困难、不准确、昂贵）限制了它们在低资源环境中的广泛实施 及其在儿童中的可行性。一种分枝杆菌细胞壁糖脂，脂阿拉伯甘露聚糖（LAM），作为诊断 病原体生物标志物已被广泛证实，一种床旁（POC）LAM检测（Alere Determine™ TB LAM）目前已投放市场。Alere的技术将LAM检测纳入基于尿液的侧流 尽管LFA检测试剂盒（LFA）易于使用，但其诊断灵敏度差限制了其对极高血糖患者的实用性。 LAM浓度（例如，晚期HIV疾病和CD 4计数<100的患者）。这种限制是 特别是在儿童中的问题，因为他们的少杆菌结核病的发病率较高（当LAM浓度 可能更低）。 在这项研究中，我们将弥合儿童结核病诊断需求未得到满足与LAM使用之间的差距 作为一个有前途的解决方案，通过应用一种新兴的技术，称为“流动”，以提高灵敏度的 拉姆·拉法耶。Alere LFA仅从几滴尿液中测量LAM，而FLOW则从尿液中浓缩LAM。 将几mL尿液浓缩至约100 µL，然后通过LFA进行分析。重要的是，FLOW浓度 完全被动，除了基于LFA的分析外，几乎不需要或不需要额外的步骤，因此使其成为一个简单的 和用户友好的LFA“前端”，特别是在TB最突出的低资源设置中。作为 在南非进行的一项小型、仅限成人、首次人体（FIH）研究中证明了原理（由法案资助 和梅林达·盖茨基金会，见支持信），在Alere和（我们新的 Salus LFA导致临床灵敏度加倍。 在本FastTrack SBIR提案中，我们将优化和调整FLOW，用于儿科患者的结核病诊断 通过：目标1，优化FLOW技术，最大限度地提高结核病儿童的分析灵敏度;目标2 2、在我们的合作伙伴临床研究中进行的一项小型、仅限儿科的初步研究中评估临床敏感性 在目标3中，根据需要实施必要的变更和/或改进， 改进我们设备的收集到回答的工作流程，包括进一步开发Salus LFA。 此外，我们建议开发一种“可穿戴”适配器，面向非常年轻的人（例如0-4岁）， 这将插入我们的FLOW工作流程，并允许有效的样本收集和评估， 患者TB状态;在Aim 4中，使用我们的FLOW TB检测试剂盒返回南非， 研究的结果将为我们的商业化路线（目标5）奠定基础。重要的是我们 我已经确定了合作伙伴（Abbott Laboratories、Intuitive Biosciences，参见支持函）， 商业化进程，如果拟议工作的数据是有希望的。

ABSTRACT While proper diagnosis is key to effective TB treatment, there exist specific barriers to TB testing in low resource settings, especially for children. While a number of TB diagnostic tools are available, they each suffer from critical drawbacks (difficult, inaccurate, expensive) that limit their widespread implementation in low resource settings and their feasibility in children. A mycobacterial cell wall glycolipid, lipoarabinomannan (LAM), as a diagnostic pathogen biomarker has been widely demonstrated and one point-of-care (POC) LAM assay (Alere Determine™ TB LAM) is currently on the market. Alere’s technology incorporates LAM detection in a urine-based lateral flow assay (LFA) and while easy to use, its poor diagnostic sensitivity limits its utility to patients with extremely high LAM concentrations (e.g., patients with advanced HIV disease and CD4 counts <100). This limitation is particularly problematic in children, given their higher incidence of paucibacillary TB (when LAM concentration is likely even lower). In this study we will bridge the gap between the unmet need for TB diagnostics in children and the use of LAM as a promising solution by applying an emerging technology, termed “FLOW”, to enhance the sensitivity of the LAM LFA. While the Alere LFA measures LAM from only a few droplets of urine, FLOW concentrates LAM from several mL of urine into ~100 µL, which is then assayed by the LFA. Importantly, FLOW concentration is completely passive, requiring little or no additional steps beyond LFA-based analysis, thus making it a simple and user friendly “front end” for LFAs, especially within low resource settings where TB is most prominent. As proof of principle, in a small, adult only, first-in-human (FIH) study performed in South Africa (funded by the Bill and Melinda Gates Foundation, see letter of support), running FLOW-concentrated urine on Alere and (our newly developed) Salus LFAs resulted in doubling of clinical sensitivity. In this FastTrack SBIR proposal we will optimize and adapt FLOW for the diagnosis of TB in pediatric patients by: In Aim 1, optimizing the FLOW technology to maximize analytical sensitivity among children with TB; In Aim 2, assessing clinical sensitivity in a small, pediatric only, pilot study performed at our partner clinical research site in South Africa; In Aim 3, implementing necessary changes and/or improvements as needed and develop an improved collection-to-answer workflow for our device, including furthering the development the Salus LFA. Additionally, we propose to develop a “wearable” adaptor, geared toward the very young (e.g. ages 0-4) and very sick, that will plug into our FLOW workflow and allow for efficient sample collection and assessment of patient TB status; and in Aim 4 employing our FLOW TB assay back to South Africa for a properly powered study, the results of which will provide a foundation for our route to commercialization (Aim 5). Importantly, we have already identified partners (Abbott Laboratories, Intuitive Biosciences, see letters of support) to complete the commercialization process if the data from the proposed work is promising.