Effect of methodological and biological variability on molecular profiling of extracellular vesicles in cancer detection

项目摘要 最近出现了一种新的旁分泌信号模式，其基础是发现细胞外 在正常和病理条件下，囊泡（EV）作为生物信息的细胞间传送器 例如癌症。我们和其他人已经证明EV及其货物可以调节基因表达并改变基因表达。 细胞在不同类型的细胞中发挥作用。此外，在诸如癌症的病理状况期间， EV的组合物改变宿主免疫应答以及同步继发性肿瘤的行为。 EV（即RNA、蛋白质、翻译后修饰、脂质， 代谢物）在健康和疾病中的作用对于理解EV的生物发生和效应器功能至关重要。 目前，EV作为与细胞内信号传导和疾病诊断相关的生物实体的研究正在进行中。 基于EV的生物发生和去除以稳态速率发生的假设， 主要由宿主的健康/患病状态改变。我们公布的结果表明，这不是 案子我们的数据表明，组织来源，数量，大小分布，以及蛋白质，脂质，代谢产物和 通过标准技术分离的EV的RNA组成不仅取决于血液收集方法， 也在当天的时间收集血液样品。因此，系统评估这些 EV谱中的可变性的因素和其它来源对于实现基础生物学、临床和生物学特性是重要的。 个性化医疗应用。 虽然这超出了这笔赠款的目的，但我们团队的长期目标是建立可复制的 血液采集和样本处理的方法，使我们能够识别特定的分子EV 在白天/夜晚周期的签名，以精确定位血液的理想，器官特异性时间的目的 这将提高早期癌症检测的可靠性和特异性。我们认为 建立生物学和方法学基线对于正确比较蛋白质组学绝对至关重要 和糖组学数据，以及解释从患者的 样品

Project Summary A novel paradigm in paracrine signaling has recently emerged based on the findings identifying extracellular vesicles (EVs) as intercellular conveyors of biological information both in normal and pathological conditions such as cancer. EVs and their cargo have been shown by us and others to regulate gene expression and alter cell function in various cell types. Moreover, during pathological conditions such as cancer, the number and compositions of EVs alter the host immune response as well as synchronize the behavior of secondary tumors. Isolation and molecular profiling of EVs (i.e. RNAs, proteins, post-translational modifications, lipids, metabolites) both in health and disease are critical for understanding EVs' biogenesis and effector functions. Currently, the study of EVs as biological entities relevant for intracellular signaling and disease diagnosis is based on the assumption that the biogenesis and removal of EVs happen at a steady state rate, being modified mostly by the healthy/diseased status of the host. Our published results show that that is not the case. Our data indicate that the tissue-origin, number, size distribution, as well as protein, lipid, metabolite and RNA composition of EVs isolated by standard techniques depend not just on the blood collection methods, but also on the time of the day the blood samples were collected. Therefore, systematic assessment of these factors and other sources of variability in EV profiles are important for enabling basic biology, clinical and personalized medicine applications. Although it is beyond the purpose of this grant, the long-term goal of our team is to establish reproducible methods for blood collection and sample processing that would allow us to identify the specific molecular EV signatures during the day/night cycle, with the aim of pinpointing the ideal, organ-specific times for blood collection, which would increase the reliability and specificity of early cancer detection. We believe that establishing biological and methodological baselines are absolutely vital for correctly comparing proteomics and glycomics data obtained in various studies, as well as interpreting the data obtained from patients' samples.