Integrated Micro Scale transcriptional profiling of cell communication networks

描述(由申请人提供)：在癌症中，风险因素和遗传学的差异在患者层面上产生了不同的临床行为(疾病进展、治疗反应)。然而，尽管对治疗结果有重要影响，但很少有工具可以在细胞水平上以严格的方式研究患者样本，并有足够的重复来实现临床上有意义的评估。这一挑战在癌症转移的背景下尤其尖锐，在这种情况下，微小肿瘤建立在细胞组成与原发肿瘤显著不同的异位环境中。但很少有工具允许探测细胞间网络通信(例如，不同细胞类型之间的旁分泌信号)。对于需要大量样本的异质性研究来说，样本制备瓶颈尤其严重(例如，每个患者样本都不同，每个样本都需要核酸提纯)。因此，需要从大量样本(多个患者、治疗和重复)中进行培养(包括共培养)、治疗和分析物纯化。我们的目标是应用简单的微尺度技术对不同种类的患者样本进行细胞间调节。我们方法的基石是一个不混油的屏障，称为“相门”，它将样品处理的上游“脏”端(培养、处理、裂解)与下游的“净”端(分析物提纯和检测)和微室共培养分隔开来，从而提高了对旁分泌信号的敏感性。我们的初步数据预测，邻近的间质在调节关键的治疗靶点方面发挥着关键作用，实验能力(吞吐量、敏感性、区隔)的进步将提供新的工具和生物学见解，将促进基础和转化性癌症研究。在这里，我们将应用一个共培养和转录分析平台来剖析细胞网络通讯-特别是细胞间调节如何有助于核受体调节，并最终促进肿瘤进展和治疗耐药。针对癌症生物学中的一个重要领域(核受体反应)的这项技术的验证和先进发展将加速其使用，使以前不可行的研究成为可能。 公共卫生相关性：我们预计，这项拨款中提议的研究不仅将有助于阐明细胞间调节如何有助于核受体调节，而且最终将有助于肿瘤进展和治疗耐药。更广泛地说，这些研究将提供新的工具和生物学见解，促进基础和转化性癌症研究。

DESCRIPTION (provided by applicant): In cancer, differences in risk factors and genetics generate diverse clinical behaviors (disease progression, response to therapy) at the patient level. Yet, despite critical implications for therapeutic outcome, few tools are available to study patient samples at the cellular level in a rigorous way with sufficient replicates to enable clinically meaningful assessments. This challenge is particularly acute in the context of cancer metastasis, where micro-tumors establish themselves in ectopic environments which are significantly different in cellular make-up than primary tumor. But few tools allow one to probe intercell network communication (e.g. paracrine signals between different cell types). A sample preparation bottleneck is particularly acute for the study of heterogeneity requiring large numbers of samples (e.g. each patient sample is different, each requiring nucleic acid purification). Thus, there is a need to perform culture (including co-culture), treatment, and analyte purification from a large number of samples (multiple patients, treatments, and repeats). Our goal is to apply simple micro scale technology to profile intercell regulation on heterogeneous patient samples. The cornerstones of our approach are an immiscible oil barrier, termed the "Phase-Gate," that separates the upstream "dirty" side (culture, treatment, lysis) of sample processing from the downstream "clean" side (analyte purification and detection) and micro chamber co-culture with increased sensitivity to paracrine signaling. Our preliminary data predicts that neighboring stroma play a critical role in regulating key therapeutic targets and that advances in experimental capabilities (throughput, sensitivity, compartmentalization) will provide new tools and biological insights that will advance basic and translational cancer research. Here, we will apply a co-culture and transcriptional analysis platform to dissecting cell network communication - specifically how intercell regulation contributes to nuclear receptor regulation and ultimately to tumor progression and therapeutic resistance. The validation and advanced development of this technology directed towards an important area in cancer biology (nuclear receptor response) will accelerate its use enable studies not feasible previously. PUBLIC HEALTH RELEVANCE: We anticipate that the research proposed in this grant will not only help to elucidate how intercell regulation contributes to nuclear receptor regulation, but ultimately to tumor progression and therapeutic resistance. More broadly these studies will provide new tools and biological insights that will advance basic and translational cancer research.