Smartphone Enabled Point-of-Care Detection of Serum Markers of Liver Cancer

摘要 肝细胞癌（HCC）是世界上第六大常见癌症，并且在低-和高-年龄组中高度流行。 中等收入国家（LMIC）。仅中国就占HCC诊断的50%以上。手术切除是 主要和唯一的治疗选择，但只有30-40%的新诊断患者是这样的候选人 治疗这是因为HCC的发展通常是无声的，诊断发生相对较晚， 当疾病已经转移或变得太大而无法手术切除时。因此，早期诊断 是取得积极临床结果的关键，因为在早期切除-当肿瘤很小， 非转移性-将导致受影响患者的治愈率显著增加。基于这一假设， 本提案目的是开发一种用于肝细胞癌的基于血液的即时检测（POCT （HCC）。我们开发的POCT“D4 POCT”非常适合LMIC的血液检测 设置. D4 POCT使用聚合物刷包被的抗体（Ab）微阵列， 背景结合产生定量结果，并在30分钟内达到皮摩尔灵敏度。所有 试剂是喷墨打印的，并储存在D4 POCT盒上，不需要冷藏。在直接 将手指针刺血液应用到卡盒上，分析物捕获和检测自动发生，生成 通过将磁带盒放置在磁性连接到智能设备的小设备中获得的可量化信号 我们将开发一种多路复用的D4 POCT， 能够同时检测甲胎蛋白（AFP）、dickkopf-1（DKK-1）和骨桥蛋白（OPN）， 将为HCC诊断提供高灵敏度和特异性。我们选择的标记物是基于最近的临床 有证据表明，同时测量AFP、DKK-1和OPN作为一个多重组， 为HCC的早期和准确诊断提供了一种有前途的策略。本提案的创新之处在于 这是几十年来第一次完全自下而上地重新设计“黄金标准”临床诊断-三明治 免疫测定-作为一种独立的即时检测微阵列测定， 智能手机该项目将由浙江杜克大学的研究人员组成的联合体进行 省人民医院和我们的行业合作伙伴Immucor，一家领先的诊断公司， 获得了这项技术的权利。在杜克对D4 POCT进行初始确认后， 性能里程碑，我们将证明在LMIC中使用的技术功能和临床潜力 在中国广州浙江省人民医院使用患者衍生的手指针刺 血液样本该项目的影响是广泛的：超越了一个新的，低成本的，临床验证 尽管用于HCC的护理点诊断的诊断技术已经被广泛使用，但是该诊断技术可以用于靶向任何 抗体对可用的分析物，并因此广泛适用于所有用于 癌症或其他疾病的诊断和预后。

ABSTRACT Hepatocellular cancer (HCC) is the sixth most common cancer worldwide, and is highly prevalent in low- and middle-income countries (LMIC). China alone accounts for >50% of HCC diagnoses. Surgical removal is the primary and only curative option, yet only 30-40% of newly diagnosed patients are candidates for such treatment. This is because the development of HCC is usually silent and diagnosis occurs relatively late, when the disease has either metastasized or grown too large for surgical removal. Therefore, early diagnosis is the key to achieving positive clinical outcomes, as removal at early-stage —when the tumor is small and not metastatic— would result in a marked increase in cures in affected patients. Motivated by this hypothesis, the objective of this proposal is to develop a blood-based point-of-care test (POCT) for hepatocellular cancer (HCC). The POCT we have developed, the “D4 POCT”, is ideally suited for blood-based testing in LMIC settings. The D4 POCT uses polymer brush-coated antibody (Ab) microarrays that have virtually no background binding, yields quantitative results, and achieves picomolar sensitivity within 30 minutes. All reagents are inkjet-printed and stored on D4 POCT cassettes, which do not require refrigeration. Upon direct application of fingerstick blood onto a cassette, analyte capture and detection occur automatically, generating a quantifiable signal obtained by placing the cassette in a small device that magnetically attaches to a smart phone, which images and analyzes microarrays via on-board App. We will develop a multiplexed D4 POCT capable of simultaneously detecting alpha-fetoprotein (AFP), dickkopf-1 (DKK-1) and osteopontin (OPN) that will provide high sensitivity and specificity for HCC diagnosis. Our choice of markers is based on recent clinical evidence showing that the simultaneous measurement of AFP, DKK-1, and OPN as a multiplexed panel provides a promising strategy for early and accurate diagnosis of HCC. The innovation of this proposal is the first complete bottom-up redesign in decades of the “gold standard” clinical diagnostic—the sandwich immunoassay—as a self-contained point-of-care microarray assay that can be imaged and interrogated by a smart phone. This project will be carried out by a consortium of investigators from Duke University, Zhejiang Provincial People's hospital and our industry partner, Immucor, a leading diagnostics company that has acquired the rights to this technology. After initial validation of the D4 POCT at Duke by meeting specific performance milestones, we will demonstrate technical functionality and clinical potential for use in LMIC settings at the Zhejiang Provincial People's hospital in Guangzhou China using patient-derived finger-stick blood samples. The impact of this project is broad: beyond a new, low-cost, and clinically validated technology for the point-of-care diagnosis of HCC, this diagnostic technology can be utilized to target any analyte for which Ab pairs are available, and is hence broadly applicable to all immunoassays used for diagnosis and prognosis for cancer or other diseases.