Abstract
Abstract The goal of this study was to characterize the small RNA transcriptome and discover novel microRNAs (miRNAs) associated with lung cancer in human bronchial epithelial cells using massively parallel sequencing. To this end, small RNA (10-40 nts) was isolated from bronchial brushings obtained via bronchoscopy. Samples pooled from healthy non-smokers, healthy smokers, and smokers with and without lung cancer (n=3 per pool) were sequenced with ABI SOLiD, using extremely deep coverage for small RNA in human tissue, 60 million 36nt-long reads/pool. Reads were aligned to the human genome with Bowtie and novel miRNAs were predicted using the miRDeep algorithm. One of the predicted novel miRNAs was selected for experimental validation. In vitro expression of the putative novel microRNA precursor was assayed with and without the siRNA knock-down of Dicer, an enzyme that processes miRNA precursors into mature miRNAs. Tissue specificity of the putative novel miRNA was profiled across 23 human tissue types using qRT-PCR. Potential microRNA targets were identified using transient overexpression of the miRNA in a cell line combined with in silico algorithms. We identified 143 miRNA that are differentially expressed in the airways of smokers with lung cancer (FC > 2) in our cohort. The miRDeep algorithm identified 131 putative novel miRNAs across all samples, including 68 differentially expressed putative miRNA in the airways of lung cancer subjects (FC > 2). In vitro expression of the putative miRNA was reduced after siRNA knockdown of DICER, providing strong evidence that the novel transcript is truly a miRNA. Out of 23 human tissue types assayed, the microRNA is expressed almost exclusively in the respiratory tract, with highest expression in the bronchus and nose and moderate expression in lung. Importantly, the expression of the miRNA is decreased significantly in lung tumors compared to adjacent normal tissue. In addition, the expression of the novel miRNA is decreased in the bronchial epithelium of smokers with lung cancer as compared to smokers without lung cancer. Overexpression of the novel miRNA in lung cancer cell line has identified putative mRNA targets that are associated with apoptosis and cell proliferation. In summary, using next generation sequencing, we have identified airway miRNA expression profiles associated with lung cancer, and we discovered a novel miRNA whose expression is reduced in the airway and lung tissue of smokers with lung cancer. This miRNA may serve as a novel biomarker to identify smokers at high risk of developing lung cancer and may also provide new insights into the genomic regulatory networks that underlie lung cancer pathogenesis. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 3989. doi:10.1158/1538-7445.AM2011-3989
Published Version
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