The Centre for Systems Biology, Lunenfeld-Tanenbaum Research Institute, Mount Sinai Hospital, Toronto, Ontario, Canada; Department of Molecular Genetics, University of Toronto, Toronto, Ontario, Canada.
Department of Surgery, Division of Urology, University of Toronto, Mount Sinai Hospital and University Health Network, Toronto, Ontario, Canada.
Eur Urol. 2014 Dec;66(6):982-6. doi: 10.1016/j.eururo.2014.07.045. Epub 2014 Sep 6.
Molecular profiling of individual cancers is key to personalised medicine. While sequencing technologies have required stringent sample collection and handling, recent technical advances offer sequencing from tissues collected in routine practice and tissues already stored in archives. In this paper, we establish methods for whole-transcriptome RNA sequencing (RNA-seq) from formalin-fixed paraffin-embedded tissues. We obtain average RNA-seq reads of >100 million per sample using the Illumina HiSeq2000 platform. We find high concordance with results from matching fresh frozen samples (>0.8 Spearman correlation). For validation, we compared low- and high-grade bladder cancer transcriptomes in 49 tumour samples after transurethral resection of bladder tumour. We found 947 differentially expressed protein-coding genes. While high-grade lesions exhibited distinct intertumour transcriptome heterogeneity, the transcriptome of low-grade tumours was homogeneous.
In this report, we show that it is now possible to use universally available bladder cancer samples that have been fixed in formalin to perform high-quality transcriptome analysis. This ability will facilitate the development of transcriptome-wide tests based on gene expression correlated with clinical outcome.
对个体癌症进行分子谱分析是实现个性化医疗的关键。虽然测序技术需要严格的样本采集和处理，但最近的技术进步提供了从常规实践中收集的组织和存档组织中进行测序的方法。在本文中，我们建立了从福尔马林固定石蜡包埋组织中进行全转录组 RNA 测序（RNA-seq）的方法。我们使用 Illumina HiSeq2000 平台获得了每个样本平均超过 1 亿个 RNA-seq 读段。我们发现与匹配的新鲜冷冻样本结果高度一致（>0.8 Spearman 相关系数）。为了验证，我们比较了 49 例经尿道膀胱肿瘤切除术的低级别和高级别膀胱癌肿瘤样本的转录组。我们发现了 947 个差异表达的蛋白编码基因。虽然高级别病变表现出明显的肿瘤间转录组异质性，但低级别肿瘤的转录组是同质的。
在本报告中，我们表明现在可以使用已用福尔马林固定的普遍可用的膀胱癌样本进行高质量的转录组分析。这种能力将有助于开发基于与临床结果相关的基因表达的转录组全谱测试。
