Rapid Sample-to-Answer Diagnosis of Kaposi's Sarcoma Across Sub-Saharan Africa using KS-COMPLETE

摘要 在本提案中，我们将开发、制造和执行一个多地点的撒哈拉以南非洲临床 KS-COMPLENT的验证-第一个真正的护理点样本到答案诊断系统， 卡波西肉瘤（KS）。我们最近在非洲的大规模研究表明，KS可以通过以下方式诊断： 在皮肤活检中定量卡波西肉瘤疱疹病毒（KSHV）DNA具有高灵敏度和特异性。 这些努力也导致了TINY的发展-一个强大的，易于使用的，无基础设施的，点- 用于KSHV DNA定量的护理（OTT）技术-目前正在多个站点部署 评价这项工作还揭示了广泛采用基于皮肤活检的皮肤活检的关键挑战， 从皮肤活检中提取DNA所需的时间和手动步骤-可能长达4小时。 KS-COMPLENT将是第一个用于皮肤穿刺活检的"直接LAMP"诊断系统。类似 直接到LAMP方法极大地简化了对其它样品基质的PCR诊断， 皮肤的胶原性质使得这对活组织检查是一个挑战。KS-COMPLENT将通过以下方式解决此问题： 我们的"SLICER"技术将自动将穿刺活检加工成更小的"微芯"， 可以通过我们的"直接LAMP"方法在TINY中直接进行DNA定量。这种方法将 将得出结果的时间缩短到60分钟左右，消除了目前所有的手动和密集的样品处理 步骤，并兼容成本，鲁棒性，基础设施和简单性的要求，在低收入国家的运作。 该系统的临床验证将通过我们在非洲建立的KS临床站点网络进行。通过 在项目结束时，我们将交付12个KS-Complete系统，并进行多中心临床验证。 KS是撒哈拉以南非洲男性和女性最常见的癌症之一。KS很难区分 其他皮肤病，特别是在非洲，那里训练有素的病理学家有限， 免疫组织化学几乎不存在。早期和更准确的诊断将赋予许多 临床获益。对于患有KS的患者，它避免了难以获得，缓慢和不可靠的需求 组织病理学检查，并允许在更早的临床阶段进行检测，从而获得更好的临床结果。患者 使用模仿者，快速排除KS可以及时重新定位诊断过程， 潜在的毒性化疗。我们的直接LAMP诊断测试可能会产生重大影响， KS诊断为目前诊断的多种其他病毒、分支杆菌和真菌相关皮肤病 通过传统的病理学可以过渡到这种方法，并最终护理点。

Abstract In this proposal, we will develop, manufacture, and perform a multi-site sub-Saharan African clinical validation of KS-COMPLETE — the first true point-of-care sample-to-answer diagnostic system for Kaposi's sarcoma (KS). Our recent large-scale studies in Africa have shown that KS can be diagnosed through quantification of Kaposi's sarcoma herpesvirus (KSHV) DNA in a skin biopsy with high sensitivity and specificity. These efforts have also resulted in the development of TINY — a robust, easy-to-use, infrastructure-free, point- of-care (PoC) technology for KSHV DNA quantification — which is being currently deployed in a multi-site evaluation. The work has also revealed that the key challenge to widespread adoption of skin biopsy-based PoC systems is the time and manual steps required to extract DNA from a skin biopsy — which can be up to 4 hours. KS-COMPLETE will be the first “direct-to-LAMP” diagnostic system for skin punch biopsies. Similar direct-to-LAMP methods have greatly simplified PoC diagnostics for other sample matrices but the solid-phase, collagenous nature of skin has made this a challenge for biopsies. KS-COMPLETE will address this issue with our “SLICER” technology that will automatically process a punch biopsy into smaller “micro-cores” on which we can directly perform DNA quantification in TINY through our “direct-to-LAMP” approach. This approach will reduce the time to result to around 60 minutes, eliminate all the current manual and intensive sample processing steps, and is compatible with cost, robustness, infrastructure, and simplicity requirements for operation in LMICs. Clinical validation of the system will be done through our established network of KS clinical sites in Africa. By the end of the project, we will deliver 12 KS-Complete systems and conduct a multi-site clinical validation. KS is one of the most common cancers in men and women in sub-Saharan Africa. KS is difficult to distinguish from other skin conditions, particularly in Africa where access to trained pathologists is limited and immunohistochemistry is practically non-existent. Early-stage and more accurate diagnosis would confer many clinical benefits. For patients who have KS, it obviates the need for the difficult to obtain, slow, and unreliable histopathology and allows for detection at earlier clinical stages resulting in better clinical outcomes. For patients with mimickers, rapid exclusion of KS allows for timely re-orienting of the diagnostic process and prevents use of potentially toxic chemotherapy. Our direct-to-LAMP diagnostic test could have significant impact beyond the diagnosis of KS as multiple other viral, mycobacterial and fungal-related skin diseases currently diagnosed through traditional pathology could be transitioned to this method and ultimately the point of care.