Large-Scale Integration of Single Cell RNA-Seq and High-Content Imaging for Analyzing Drug Response in Cancer

 描述(申请人提供)：单细胞分析对于了解癌细胞对药物的反应至关重要，因为药物反应天生是不同步的，转化的细胞在遗传和表观遗传上都是异质的。虽然单细胞RNA-Seq正在成为评估表型异质性的高维和日益可扩展的工具，但仅从转录组不能推断细胞表型的许多方面。然而，如果我们能够大规模地结合单细胞成像和RNA-Seq，那么我们就可以利用高含量的成像分析来获得关于细胞代谢、蛋白质定位、翻译、细胞周期和单细胞分辨率的细胞信号的大量表型观察。现有的用于合并单细胞成像和测序的工具价格昂贵、吞吐量低，并且与短期细胞培养和刺激不兼容。在这里，我们将通过将我们高度可扩展的单细胞RNA-Seq和成像微流控平台与一种新的条形码策略相结合来解决这个问题，该策略将允许我们将池库中的序列条形码与我们设备中的光学条形码相关联。将进行定量研究，以验证我们的单细胞药物刺激、成像和RNA-Seq集成平台在几种癌症细胞系上的有效性，这些细胞含有靶向和细胞毒治疗中使用的化合物。

 DESCRIPTION (provided by applicant): Single cell analysis is crucial for understanding how cancer cells respond to drugs because drug responses are inherently asynchronous and transformed cells are both genetically and epigenetically heterogeneous. While single cell RNA-Seq is emerging as a high-dimensional and increasingly scalable tool for assessing phenotypic heterogeneity, many aspects of cellular phenotype cannot be inferred from the transcriptome alone. However, if we can combine single cell imaging and RNA-Seq on a large-scale, then we can take advantage of high-content imaging assays with access to numerous phenotypic observables for cellular metabolism, protein localization, translation, cell cycle, and cell signalng with single cell resolution. Existing tools for merging single cell imaging and sequencing are expensive, low-throughput, and incompatible with short-term cell culture and stimulation. Here, we will address this issue by combining our highly scalable microfluidic platform for single cell RNA-Seq and imaging with a novel barcoding strategy that will allow us to associate sequence barcodes in a pooled library with optical barcodes in our device. Quantitative studies will be carried out to validate our integrated platform for single cell drug stimulation, imaging, and RNA-Seq on several cancer cell lines with compounds used in both targeted and cytotoxic therapies.