Development of a chemical reaction sensor array platform for label-free, real-time kinetic analysis of enzyme-substrate reactions to enable high-throughput drug discovery

项目摘要 全球制药公司用于研发的支出持续增加 以目前估计的10亿美元和15年以上的时间开发一种新药。药物发现努力 对候选人选择的下游成功至关重要，并严重依赖高通量筛选 (HTS)以确定可行的线索。目前用于HTS药物发现分析的方法，如荧光法或 基于发光的方法存在固有的技术缺陷和限制，导致错误的结果 导致失败的程序。此外，目前的大多数高温超导方法不能修改为超高- 吞吐量和使用终点分析，而不是实时动力学方法，可以提供更多 彻底评估分子间的相互作用。使用实时动力学分析和更高的处理能力 药物发现计划可以极大地提高成功率，同时减少成本和时间。 INano Bio正在开发一种新型的全耗尽指数耦合(FDEC)场效应管(FET) 用于高温超导的基于生物传感器的纳米传感器多路电子药物发现平台。我们的nMEDD 平台将提供(I)实时酶反应监测；(Ii)超高检测灵敏度；(Iii)超高温超导 可伸缩性；以及(Iv)高度自动化兼容性。FDEC FET传感器以电子方式监控 通过与溶液中的离子直接反应而产生的电荷或电势，从而可以确定动力学反应 实时检测和定量测量抑制剂或激活剂对 酶促反应。作为对该技术平台的初步验证，我们正专注于为 用于治疗阿尔茨海默病(AD)的药物研发。具体地说，我们将专注于开发监测 Tau的磷酸化和去磷酸化，以筛选tau磷酸化的调节子，AS Tau的胞浆聚集物与AD的病理有关。 该Fast-Track计划的总体目标是开发INanoBio的nMEDD平台作为HTS方法 为了药物发现的目的而监测酶反应。为了实现这一目标，第一阶段计划将是 重点开发了一种小规模的多重传感器和FDEC FET检测tau磷酸化和 去磷酸化以验证多路传感器的性能。成功完成第一阶段将 产生具有适用于1536孔板格式的间隔的传感器芯片并能够提供 芯片响应稳定，可检测tau的磷酸化和去磷酸化。第二阶段 该计划将专注于通过完成以下工作来进一步展示FDEC FET技术的商业潜力 用于1536孔磷酸化检测的FDEC FET复合传感器及检测系统的研制 以及tau的去磷酸化，此外还验证了该检测方法并证明了其更高的 吞吐量。拟议方案的成功完成将为以下方面提供nMEDD技术的验证 药物发现，以支持合作努力，并为将技术扩展到其他疾病领域提供信息。

Project Summary Global expenditures by pharmaceutical companies for research and development continue to increase each year with a current estimate of $1B USD and upwards of 15 years to develop a new drug. Drug discovery efforts are critical to the downstream success of candidate selection and relies heavily on high-throughput screening (HTS) to identify viable leads. Current methods for HTS drug discovery assays, such as fluorescence-, or luminescence-based methods, suffer from inherent technical drawbacks and limitations that result in false results leading to failed programs. Additionally, the majority of current HTS methods are not amendable to ultra high- throughput and use end-point assays as opposed to real-time kinetic approaches that can provide a more thorough assessment of molecular interactions. The use of real-time kinetic assays and higher throughput in drug discovery programs could greatly enhance success rates while reducing cost and time. INanoBio is developing a novel fully depleted exponentially coupled (FDEC) field effect transistor (FET) biosensor-based nanosensor multiplexed electronic drug discovery platform (nMEDD) for HTS. Our nMEDD platform will offer (i) real time enzymatic reaction monitoring; (ii) ultra-high sensitivity of detection; (iii) ultra HTS scalability; and (iv) high automation compatibility. The FDEC FET sensors electronically monitor changes in charge or potential by directly reacting with ions in solution, thus allowing for the determination of kinetic reaction rates to inform real-time detection and quantitative measurement of the effects of inhibitors or activators on the enzymatic reaction. As an initial validation of the technology platform, we are focusing on developing nMEDD for use in drug discovery for Alzheimer’s disease (AD). Specifically, we will focus on developing assays to monitor tau phosphorylation and dephosphorylation to enable screening for modulators of tau phosphorylation, as cytosolic aggregates of tau have been linked to AD pathology. The overall goal of this Fast-Track program is to develop INanoBio’s nMEDD platform as a HTS method for monitoring enzymatic reactions for drug discovery purposes. To achieve this goal, the Phase I program will be focused on development of a small-scale multiplex sensor and FDEC FET assays for tau phosphorylation and dephosphorylation to validate the performance of the multiplex sensor. Successful completion of Phase I will result in a sensor chip with spacing that is applicable to a 1536-well plate format and is capable of providing a stable response across the chip for detection of tau phosphorylation and dephosphorylation. The Phase II program will focus on further demonstrating the commercial potential of the FDEC FET technology by completing fabrication of an FDEC FET multiplex sensor and detection system for 1536-well detection of phosphorylation and dephosphorylation of tau, in addition to validation of the assay and demonstration of its potential for higher throughput. Successful completion of the proposed program will provide validation of the nMEDD technology for drug discovery to support partnering efforts and inform expansion of the technology into other disease areas.