Microfluidic intact cell platform: A novel tool for oral cancer detection

口腔鳞状细胞癌(OSCC)夺走了美国数千人和数十万人的生命 每年在全球范围内。诊断口腔鳞状细胞癌的黄金标准是活检和组织病理学检查。 如果需要且经常需要对同一病变进行纵向评估，则痛苦、有创、昂贵、不切实际 在第三世界国家是不可用或不精确的。一种快速区分癌症和非癌症的工具 癌性病变和识别进展性或转化性病变可以允许早期干预，这 将改善结果，并消除对残留病变的患者进行不必要的活检的必要性 良性或尚未开始退化。因此，对快速、非侵入性、 口腔鳞癌的客观、经济的检测方法。我们和其他人已经报道了一种改变的表达模式 人β防御素3(HBD-3)，一种上皮细胞衍生的抗菌肽(AMP)，以及HBD-2，另一种 上皮细胞AMP是口腔鳞癌的早期事件。因此，当病变中HBD-3和HBD-2的比率 与对侧相比，可用于口腔鳞癌与口腔其他病变的鉴别 空洞。我们将这一比率称为防御素指数(BDI)。我们正在进行的78例患者的临床研究 可疑口腔病变表现出高度的敏感性(100%)和特异性(74%)的基础上的BDI 口腔癌性病变与非癌性病变的鉴别(P&lt；0.0001)与我们的BDI的高准确率(98%) 在分子检测的基础上，我们现在希望将我们的新平台从费力、耗时的ELISA 格式为基于成像的护理点(POC)设备，该设备利用微流控技术来量化BDI 预计周转时间为半小时。我们的微流控完整细胞分析(MICA)方法 用于口腔癌检测的POC设备是独一无二的；它利用捕获在微流控芯片中的完整上皮细胞 包括具有不同间距的微型制造的柱状阵列，以允许捕获上皮细胞。vt.在.的基础上 捕获后，细胞被渗透，并用荧光抗体标记以进行HBD比率分析。我们雇佣了 自动荧光成像和计算算法，以实现BDI的自动计算 得分。我们现在假设，可以有效检测口腔癌的ELISA格式可以是 配置为护理点MICA，保持其高精度，并使其更容易在全球使用。至 为了推动这一新的口腔癌检测方法的发现，我们提出了以下目标：1. 开发一种用于口腔癌检测的MICA POC设备的工作样机，配备细胞成像 和BDI计算能力。2.进行发现阶段研究，其中MICA POC和EL ISA，AS 在检测口腔癌的能力方面，将与病理学检查进行比较。这个 云母POC虽然不打算取代活组织检查，但在未来可以被部署到客观和非 无创性地确定谁确实需要活组织检查，监测现实世界实践中的口腔癌前病变和 在缺乏和/或不可靠病理学审查的低社会经济国家，满足未得到满足的主要需求。

Oral squamous cell carcinoma (OSCC) claims the lives of thousands in the U.S. and hundreds of thousands worldwide annually. A biopsy followed by histopathology, the gold standard for the diagnosis of OSCC, is painful, invasive, costly, not practical if longitudinal assessments of the same lesion are required and oftentimes is not available or imprecise in third world countries. A tool that quickly distinguishes cancerous from non- cancerous lesions and identifies progressive or transforming lesions could allow for early intervention, which would improve outcomes and negate the need for unnecessary biopsies in patients whose lesions remain benign or haven’t begun to degenerate. Therefore, there is an unmet need for a rapid, non-invasive, objective and cost-effective test for OSCC. We and others have reported that an altered expression profile of human beta defensin 3 (hBD-3), an epithelial cell derived antimicrobial peptide (AMP), and hBD-2, another epithelial cell AMP, is an early event in OSCC. Therefore, the ratio of hBD-3 and hBD-2 in the lesion, when compared to the contralateral site, could be exploited in distinguishing OSCC from other lesions of the oral cavity. We refer to this ratio as the beta defensin index (BDI). Our ongoing clinical study of 78 subjects with suspicious oral lesions demonstrated high sensitivity (100%) and specificity (74%) of the ELISA based BDI in distinguishing cancerous from noncancerous oral lesions (P<0.0001). With the high accuracy (98%) of our BDI based molecular assay, we now wish to advance our novel platform from the laborious, time consuming ELISA format into an imaging-based point-of-care (POC) device that utilizes microfluidic technology to quantify the BDI with an expected turnover time of half an hour. Our microfluidic intact cell assay (MICA) approach to developing a POC device for oral cancer detection is unique; it utilizes intact epithelial cells trapped in a microfluidic chip encompassing microfabricated pillar arrays with varying spaces to allow the capture of epithelial cells. Upon capture, the cells are permeabilized and labeled with fluorescent antibodies for hBD ratio analysis. We employ automated fluorescence imaging and computational algorithm to enable automated calculation of the BDI scores. We now hypothesize that the ELISA format that can effectively detect oral cancer, can be configured for point of care MICA, retaining its high accuracy and making it easier to use worldwide. To advance the discovery of this new approach for oral cancer detection, we propose the following aims: 1. Develop a working prototype of a MICA POC device for oral cancer testing equipped with cell imaging and BDI calculation capabilities. 2. Conduct a discovery phase study where MICA POC and ELISA, as independent assays, will be compared with pathology review in their ability to detect oral cancer. The MICA POC, while not intending to replace biopsy, could be deployed, in the future, to objectively and non- invasively determine who actually needs a biopsy, monitor oral premalignant lesions in real world practice and fulfill a major unmet need in low-socio economic countries where pathology review is lacking and/or unreliable.