Identification of somatic mutations in non-small cell lung carcinomas using whole-exome sequencing

저자: Pengyuan Liu, Carl Morrison, Liang Wang, Donghai Xiong, Peter Vedell, Peng Cui, Xing Hua, Feng Ding, Yan Lu, Michael James, John D.Ebben, Haiming Xu, Alex A.Adjei, Karen Head, JaimeW.Andrae, Michael R.Tschannen, Howard Jacob, Jing Pan4, Qi Zhang, Francoise Van den Bergh, Haijie Xiao, Ken C.Lo, Jigar Patel, Todd Richmond, Mary-Anne Watt, Thomas Albert, Rebecca Selzer, Marshall Anderson, Jiang Wang, Yian Wang, Sandra Starnes, Ping Yang and Ming You

저널: Carcinogenesis. 2012 Jul;33(7):1270-6

사용한 제품: Dynabeads Streptavidin MyOne T1 (Cat# 65601, 65602)

Abstract

Lung cancer is the leading cause of cancer-related death, with non-small cell lung cancer (NSCLC) being the predominant form of the disease. Most lung cancer is caused by the accumulation of genomic alterations due to tobacco exposure. To uncover its mutational landscape, we performed whole-exome sequencing in 31 NSCLCs and their matched normal tissue samples. We identified both common and unique mutation spectra and pathway activation in lung adenocarcinomas and squamous cell carcinomas, two major histologies in NSCLC. In addition to identifying previously known lung cancer genes (TP53, KRAS, EGFR, CDKN2A and RB1), the analysis revealed many genes not previously implicated in this malignancy. Notably, a novel gene CSMD3 was identified as the second most frequently mutated gene (next to TP53) in lung cancer. We further demonstrated that loss of CSMD3 results in increased proliferation of airway epithelial cells. The study provides unprecedented insights into mutational processes, cellular pathways and gene networks associated with lung cancer. Of potential immediate clinical relevance, several highly mutated genes identified in our study are promising druggable targets in cancer therapy including ALK, CTNNA3, DCC, MLL3, PCDHIIX, PIK3C2B, PIK3CG and ROCK2.

제품 사용법

Capture of target genome

Exonic sequences from tumor (purity .70%) and normal DNAs were enriched using Agilent’s SureSelect technology for targeted exon capture, targeting 38 Mb of sequence from 212 911 exons and their flanking regions in  ~20,000 genes. Five hundred nanograms of the prepped library was incubated with whole exon biotinylated DNA capture baits supplied in the kit for 24 h at 65 C. The captured DNA hybrids were recovered using Dynabeads MyOne Streptavidin T1 from Dynal. The DNAwas eluted from the beads and desalted using Qiagen MinElute PCR purification columns. The purified capture products were then amplified using the SureSelect GA PCR primers (Agilent) for 12 cycles.