Development of a semi-quantitative ligation-based assay for Hepatitis B

总结/摘要 虽然疫苗可以预防，但慢性B型肝炎病毒（HBV）感染仍然是全球公共卫生问题 问题：HBV引起的肝硬化和肝癌在美国约有2,000人死亡， 全球每年有62万人。在全球范围内，世卫组织报告了2.57亿人，即人口的3.5%， 2015年，全世界的慢性HBV感染者。未经治疗的慢性病毒性肝炎可进展为 危及生命的并发症，如肝硬化或肝细胞癌。与其他病毒合并感染， 如艾滋病毒也很常见。一旦受影响的人被诊断出来，患者将被长期安置在 治疗HBV治疗的目标是通过持续的治疗来阻止或逆转肝脏炎症和纤维化。 抑制HBV复制。然而，为了有效，治疗需要持续监测，因为 对治疗的抵抗可能会发展，特别是在不顺从的患者中。 虽然免疫测定是诊断HBV患者的标准方法，但核酸扩增试验（NAAT） 已经成为乙肝的标准治疗。由于病毒载量（VL）检测是最重要的指标， 治疗反应，VL应在基线时测量所有患者，然后定期测量，特别是 在接受治疗的患者中。VL通常用精密仪器测量，目前没有 结合了联合收割机速度、准确性、低成本和Point Of- 在乎一个能提供病毒载量信息的真正的HBV POC NAAT将立即产生临床影响。 我们建议开发一种准确、快速、廉价的HBV病毒载量纸基分子检测方法 半定量与金标准NAAT一样准确，但成本要低得多， 在POC中执行。 我们已经开发了一种纸流分子诊断平台，其在几个等温条件下表现良好。 核酸扩增测定。我们将把这个平台的联合收割机创新与一种新颖的、可调的半导体激光器相结合， 基于目标扩增子的可预测分层的定量测定，所述目标扩增子基于其丰度。在这 开发计划，我们将专注于开发和优化的测定，以及初始步骤， 使系统便于携带，为今后工作的全面整合奠定基础。这项工作有可能 改变我们如何管理慢性HBV患者的护理。

Summary/Abstract While vaccine-preventable, chronic infection with hepatitis B virus (HBV) remains a global public health problem: HBV-induced liver cirrhosis and liver cancer kill about 2,000 people in the United States and roughly 620,000 people worldwide each year. Globally, WHO reported 257 million people, or 3.5% of the population, were living with chronic HBV infection in the world in 2015. Untreated chronic viral hepatitis can progress to life-threatening complications, such as cirrhosis or hepatocellular carcinoma. Coinfections with other viruses, such as HIV, are also common. Once affected individuals are diagnosed, patients are placed on long-term treatment. The goals of HBV therapy are to stop or reverse liver inflammation and fibrosis, through sustained suppression of HBV replication. However, to be effective, therapies require continuous monitoring, since resistance to treatment can develop, especially in non-compliant patients. While immunoassays were the norm for diagnosing HBV patients, nucleic acid amplification tests (NAATs) have become standard of care for HBV. Since viral load (VL) testing is the most important indicator of response to therapy, VL should be measured in all patients at baseline and then on a regular basis, especially in patients on treatment. VL is usually measured with sophisticated instruments, and there currently are no methods for monitoring HBV viral load that combine speed, accuracy, low cost, and the option for Point-Of- Care. A truly POC NAAT for HBV that provided viral load information would have an immediate clinical impact. We propose to develop an accurate, rapid, and inexpensive paper-based molecular test for HBV Viral Load semi-quantification that is as accurate as gold standard NAATs but far less expensive, and that can be implemented at the POC. We have developed a paperfluidic molecular diagnostic platform that performs well with several isothermal nucleic acid amplification assays. We will combine innovations from this platform with a novel, tunable semi- quantitative assay based on predictable stratification of target amplicons based on their abundance. In this developmental program we will focus on development and optimization of the assay, and the initial steps to make the system portable, setting the stage for full integration in future work. This work has the potential to change how we manage care for chronic HBV patients.