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

 描述（由申请人提供）：单细胞分析对于了解癌细胞如何对药物产生反应至关重要，因为药物反应本质上是异步的，并且转化细胞在遗传和表观遗传上都是异质的。虽然单细胞RNA-Seq正在成为一种用于评估表型异质性的高维和日益可扩展的工具，但细胞表型的许多方面不能仅从转录组推断。然而，如果我们能够大规模地将联合收割机单细胞成像和RNA-Seq相结合，那么我们就可以利用高含量的成像测定，获得细胞代谢、蛋白定位、翻译、细胞周期和细胞信号传导的许多表型观测值，并具有单细胞分辨率。现有的用于合并单细胞成像和测序的工具价格昂贵、通量低，并且与短期细胞培养和刺激不兼容。在这里，我们将通过将我们用于单细胞RNA-Seq和成像的高度可扩展的微流体平台与一种新的条形码策略相结合来解决这个问题，该策略将使我们能够将汇集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，以验证我们在几种癌细胞系上的单细胞药物刺激，成像和RNA-Seq的集成平台，其中化合物用于靶向和细胞毒性治疗。

 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.