American Association for Cancer Research
10780432ccr120677-sup-fig1.pdf (7.83 MB)

Supplementary Figure 1 from The Use of Quantitative Real-Time Reverse Transcriptase PCR for 5′ and 3′ Portions of ALK Transcripts to Detect ALK Rearrangements in Lung Cancers

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journal contribution
posted on 2023-03-31, 17:44 authored by Rui Wang, Yunjian Pan, Chenguang Li, Haichuan Hu, Yang Zhang, Hang Li, Xiaoyang Luo, Jie Zhang, Zhaoyuan Fang, Yuan Li, Lei Shen, Hongbin Ji, David Garfield, Yihua Sun, Haiquan Chen

PDF file, 8019K, Heatmaps of normalized Exon Array expression values for ALK in lung adenocarcinomas.



Purpose: Approximately 3% to 7% of non–small cell lung cancers (NSCLC) harbor an ALK fusion gene, thus defining a tumor group that may be responsive to targeted therapy. The breakpoint in ALK consistently occurs at exon 20 and EML4 or other fusion partners, thus driving a strong expression of ALK kinase domain and resulting in an unbalanced expression in 5′ and 3′ portions of ALK transcripts. We have developed a rapid and accurate method by simultaneously detecting the expression in 5′ and 3′ portions of ALK mRNA.Experimental Design: Quantitative real-time reverse transcriptase PCR (qRT-PCR) was used to examine expression levels of the 5′ and 3′ portions of ALK transcripts in177 NSCLCs, in which EGFR, KRAS, HER2, and BRAF mutations were absent. If unbalanced ALK mRNA expression was seen, ALK rearrangement was assumed to exist. ALK FISH was used to confirm the accuracy of qRT-PCR. RT-PCR and 5′ RACE coupling sequencing identified the fusion variants.Results: Real-time RT-PCR showed excellent sensitivity and specificity (100% and 100%, respectively) for detection of ALK rearrangements in resected specimens. In addition, six novel ALK fusion variants were identified, including one KIF5B-ALK (E17;A20) and five EML4-ALK variants (E6a;A19, E6a/b ins 18;A20, E17b ins 39;A20, E10a/b, E13;A20, and E17 ins 65;A20).Conclusions: Real-time RT-PCR is a rapid and accurate method for diagnosing ALK-rearranged lung cancers. Coupling of 5′ RACE to this method should further facilitate rapid identification of novel ALK fusion genes. Clin Cancer Res; 18(17); 4725–32. ©2012 AACR.

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