Rapid, low-cost mRNA analysis system for cancer companion diagnostics

描述（由申请人提供）：核酸检测构成了新兴分子诊断（“MDx”）中一个庞大且不断增长的部分。伴随诊断是MDx的一个极其重要的方面，因为它有可能使用基于基因组的个性化方法来告知和伴随疾病的治疗。分子诊断的下一步将需要开发敏感和特定的仪器，以低成本和低复杂性提供疾病生物标志物的重复测试。LiquiLume诊断公司的长期目标是开发基于光流体的解决方案，光流体是集成光学和微流体在单芯片级系统上的结合。该应用程序的目的是展示LiquiLume设想的配套诊断仪器，Optofluidic RNA生物标志物测试系统（orbit）的关键功能。orbit是一种平台技术，在单分子水平上引入基于光流体波导的检测，用于无扩增核酸检测。作为第一个代表性的例子，我们将展示临床标本中癌症生物标志物的无扩增监测。本应用程序的目标将通过完成以下具体目标来实现：(1)在临床样品中检测核酸（mRNA）；(2)多点激发核酸检测，均采用orbit光流芯片。在第一个目标下，我们将展示来自临床样品材料的mRNA癌症生物标志物可以被识别和定量。具体来说，我们将展示在接受抗癌药物易普利姆单抗治疗的黑色素瘤患者外周血单个核细胞（PBMCs）中检测五种mRNA生物标志物。第二个目标将通过干涉波导结构引入多点光激发，以提高信本比。将演示比单点激励提高100倍。该项目的创新之处在于首次展示了用于癌症免疫治疗的集成光流体技术，并引入了用于光信噪比增强的干涉激励。这项工作意义重大，因为它将引入一种新型仪器，该仪器具有金标准聚合酶链反应方法的特异性和敏感性，但复杂性和成本都要低得多。轨道原理的演示不仅适用于癌症的伴随诊断，而且为更广泛的分子诊断仪器的快速病原体和生物标志物检测提供了蓝图。

DESCRIPTION (provided by applicant): Nucleic acid tests comprise a large and growing segment of emerging molecular diagnostics ("MDx"). Companion diagnostics is an extremely important aspect of MDx due its potential to inform and accompany treatment of diseases using a genome-based, personalized approach. Taking the next step in molecular diagnostics will require the development of sensitive and specific instruments that provide repeated testing of disease biomarkers at low cost and low complexity. The long-term goal of LiquiLume Diagnostics Inc. is to develop solutions to these challenges based on optofluidics - the combination of integrated optics and microfluidics on a single chip-scale system. The objective of this application is to demonstrate key capabilities of LiquiLume's envisioned companion diagnostic instrument, Optofluidic RNA Biomarker Testing System (ORBITS). The ORBITS is a platform technology introducing optofluidic waveguide- based detection on the single molecule level for amplification-free nucleic acid detection. As a first, representative example, we will demonstrate amplification-free monitoring of cancer biomarkers in clinical specimens. The objectives of this application will be accomplished by completing the following specific aims: (1) Nucleic acid (mRNA) detection in clinical samples; and (2) Multi-spot excitation nucleic acid detection, both using ORBITS optofluidic chips. Under the first Aim, we will show that mRNA cancer biomarkers from clinical sample material can be identified and quantitated. Specifically, we will demonstrate detection of five mRNA biomarkers in peripheral blood mononuclear cells (PBMCs) from melanoma cancer patients treated with the anti-cancer drug ipilimumab. The second Aim will introduce multi-spot optical excitation via interferometric waveguide structures for enhancing signal-to-background ratios. A 100x improvement over single-spot excitation will be demonstrated. The project's innovative aspects lie in the first demonstration of integrated optofluidic technology for cancer immunotherapy, and the introduction of interferometric excitation for optical signal-to-noise enhancement. The proposed work is significant because it will introduce a new type of instrument that features the specificity and sensitivity of gold standard polymerase chain reaction methods at much lower complexity and cost. Demonstration of the ORBITS principle will not only be applicable to companion diagnostics of cancer, but also serve as the blueprint for a broader class of molecular diagnostic instruments for rapid pathogen and biomarker detection. PUBLIC HEALTH RELEVANCE: Quantitative monitoring of tens of biomarkers is an essential component of cancer immunotherapy and other forms of emerging companion diagnostics. This application describes a novel approach to this problem using amplification-free detection of cancer mRNA biomarkers on an optofluidic chip. The proposed integrated diagnostic platform would optimize public health interventions by providing sensitive and accurate molecular diagnostics with low cost and complexity.