Multiplexed analysis of exosomes in cancer nano therapy

 描述(由申请人提供)：检测治疗反应和耐药性的可靠标记物仍然是卵巢癌的主要临床挑战。这一点对于 一系列纳米疗法进入临床领域。一个很少探索的机会在于分析外切体，50-200纳米大小的囊泡不断进入血液循环。重要的是，这样的肿瘤来源的外切体是丰富的(癌症患者的外周血液中有1011个小泡/毫升)，随着时间的推移相当稳定，并且包含与亲代肿瘤中发现的蛋白质相同的蛋白质。目前的一个主要挑战和机遇是开发从临床样本中快速确定CMV丰度和组成的方法。我们最近开发了一种名为NPLEx(纳米等离子Exosome)的高灵敏度、基于纳米技术的护理点诊断方法，它包括在不透明金膜中制造的周期性纳米孔阵列，用于传输等离子成像(NAT Biotechnol 2014；32,490-5)。我们已经证明，i)这种分析非常敏感，ii)允许对外切体及其内部的数十种蛋白质进行剖析，从而可以进行细胞起源研究，以及iii)NPLEx分析可以进行复杂的治疗评估。该应用程序的目标是通过i)扩展NPLEx技术以允许蛋白质和RNA的并行分析，ii)在小鼠模型和在 诊所。我们假设，该方法在胞外分析中将比目前可能的更敏感和全面，并允许进行治疗评估。提出的集成图谱方法有可能改变纳米治疗试验、癌症研究和临床实践。它将使客观的治疗性读出成为可能，这些事件发生在传统的临床指标之前。它将促进在缺乏细胞的临床标本中广泛分析外切体，显著降低成本，并可以很容易地与其他下游分析相结合。

 DESCRIPTION (provided by applicant): Detection of reliable markers of therapy response and drug resistance remains a major clinical challenge in ovarian cancer. This is particularly true for a series of nanotherapeutics entering the clinical arena. One little explored opportunity lies in the analysis of exosomes, 50-200 nm sized vesicles continuously shed into the circulation. Importantly such tumor derived exosomes are abundant (1011 vesicles/ml of peripheral blood in cancer patients), are fairly stable over time and contain proteins reflective of those found in parent tumors. A current major challenge and opportunity is the development of methods for rapidly determining the abundance and composition of cMVs from clinical samples. We have recently developed a highly sensitive, nanotechnology-based, point-of-care diagnostic method termed nPLEX (nano Plasmonic Exosome) which comprises periodic nanohole arrays fabricated in an opaque gold film for transmission plasmon imaging (Nat Biotechnol 2014;32, 490-5). We have shown that i) such analyses are exquisitely sensitive, ii) allow profiling of dozens of proteins on and inside exosomes thus allowing cell of origin studies and iii) that nPLEX analysis allows sophisticated therapy assessment. The goal of this application is to advance the nPLEX technology by i) expanding it to allow parallel profiling of protein and RNA, ii) validating it as a therapeutic read-out of nanoparticle therapeutics in mouse models and in the clinic. We hypothesize that the approach will be more sensitive and comprehensive in exosomal analysis than is currently possible and allow treatment evaluation. The proposed integrated profiling method has the potential to transform nano therapeutic trials, cancer research and clinical practice. It will enable objective therapeutic read-outs, events that occur before conventional clinical metrics. It will facilitate extensive profiling of exosome in paucicellular clinical specimens, significantly reduce costs, and can be easily combined with other downstream analyses.