SBIR PA22-176 - Rapid Point-of-Care Detection of Human Papillomavirus

SBIR PA22-176 - 人乳头瘤病毒的快速护理点检测

• 批准号: 10761537
• 负责人: Zhenrong Zheng
• 金额: $ 27.58万
• 依托单位: ELECTRONUCLEICS, INC.
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-09-30 至 2024-09-29
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10761537
• 关键词: SBIR; PA22; 176; Rapid; Point

Project Summary The goal of this project is to demonstrate feasibility of a rapid (≤5 mins), sensitive system for the point-of-care (POC) detection of human papillomavirus (HPV) in human vaginal samples that integrates our patented detector of pathogens based on identification of species-specific, nucleic acid (NA) sequences. The detector relies on a patented electromechanical signal transduction mechanism that enables the low-cost, optics-free and amplification-free (e.g., no PCR) detection of DNA/RNA at ultralow concentration (~10-19 M). A compelling need persists for rapid (minutes), cost effective, POC NA detection devices for infectious disease diagnostics so as to facilitate prompt, well-informed therapies and counseling and to avoid potentially harmful actions including the inappropriate prescription of antibiotics. In earlier work, we demonstrated the detection of Neisseria gonorrhoeae (NG) with sensitivity of ~98% and specificity of ~100%. We currently are gathering similar data for detection of Chlamydia trachomatis (CT). Since HPV is the most common STI and the cause of slowly developing cervical cancers, it is a logical target for us to address in order to facilitate life-saving precancer screening. A key feature of our patented detector is the use of peptide nucleic acid (PNA) capture probes, which are uncharged polyamide analogs to NAs that share the same base chemistry. Since bead-PNA conjugates are designed to be charge neutral, they do not exhibit electrophoretic movement in the presence of a DC electric field. However, the substantial negative charge acquired upon capture of a target NA sequence makes the hybridized conjugate mobile. Electrophoresis of the bead-PNA conjugate with hybridized target NA to the mouth of a smaller diameter glass pore causes a significant increase in pore resistance, thereby resulting in a strong, sustained drop in measured ionic current. Nonspecifically bound NA is removed from the bead conjugate in the strong electric field at the pore mouth resulting in no sustained signal. Further, the opposing electroosmotic flow through the glass pore sweeps PNA-bead conjugates without hybridized target away from the pore mouth. In such a way, this simple conductometric device gives a highly selective (no observed false positives), binary response signaling the presence or absence of the target NA (and associated pathogen). This project focuses on three Aims: 1) Detection of highest risk HPV mRNA in media at ≤10-19 M (~60 copies/mL) and of HPV-infected human cells at ~5-10 cell/mL; 2) Development and testing of an integrated, lateral flow assay (LFA)-based prototype device for rapid (≤5 mins) detection of HPV-infected human cells at ~5-10 cells/mL; and 3) Demonstration of highest risk HPV detection in remnant clinical samples in 95% concordance with an approved commercial nucleic acid amplification test (NAAT). Successful achievement of these Aims will result in a device concept ready for manufacturable prototype development that is capable of HPV testing with competitive accuracy and with significant advantages of cost, rapidity, low power and robustness.

项目摘要 该项目的目的是证明快速（≤5分钟）的可行性，即护理点的敏感系统 （POC）在整合我们专利检测器的人类阴道样品中检测人乳头瘤病毒（HPV） 病原体基于规格特异性核酸（NA）序列的鉴定。检测器依靠 专利的机电信号转导机制，可实现低成本，无光学和 在超高浓度（〜10-19 m）下无扩增（例如，无PCR）检测DNA/RNA。令人信服的需求 持续存在快速（分钟），具有成本效益的POC NA检测设备，用于传染病诊断，以便 促进及时，信息良好的疗法和咨询，并避免可能采取可能有害的行动 在较早的工作中，我们证明了淋病奈瑟氏菌的检测 （ng）敏感性约为98％，特异性约为100％。我们目前正在收集类似的数据以检测 衣原体沙眼（CT）。由于HPV是最常见的性传播疾病，也是缓慢发展宫颈的原因 癌症，这是我们要解决的逻辑目标，以促进挽救生命的预科筛查。一个关键功能 我们专利的检测器的使用是使用肽核酸（PNA）捕获探针，这些探针是未加成的聚酰胺 对NA的类似物，共享相同的基础化学。由于珠-PNA偶联物的设计为充电 中性，在存在直流电场的情况下，它们不存在电泳运动。但是， 捕获目标NA序列后获得的实质性负电荷使杂交共轭物 移动的。珠-PNA的电泳与杂交靶Na偶联到较小直径的口 玻璃孔会显着增加孔的抗性，从而导致强劲，持续下降 测量的离子电流。非特异性绑定的Na从强电气中的珠子结合中删除 孔口处的场导致没有持续信号。此外，相反的电子流通过 玻璃孔糖pNA珠子偶联，而没有杂交靶标的远离孔口。这样， 这种简单的电导器设备具有高度选择性（没有观察到的假阳性），二进制响应 信号表明靶NA（以及相关病原体）的存在或不存在。这个项目重点是三个 目的：1）在≤10-19m（约60份/mL）和HPV感染的人类中检测媒体中最高风险的HPV mRNA 〜5-10细胞/mL的细胞； 2）基于集成的侧向流量测定（LFA）原型的开发和测试 在〜5-10细胞/mL的HPV感染人类细胞中快速检测（≤5分钟）的装置； 3）演示 残留的临床样本中最高风险的HPV检测与批准的商业核酸次数为95％的一致性 酸扩增测试（NAAT）。这些目标的成功实现将导致设备概念准备 能够以竞争精度进行HPV测试的制造原型开发 成本，速度，低功率和鲁棒性的显着优势。