Molecular diagnosis & therapy · 2021
Detecting ALK Rearrangement with RT-PCR: A Reliable Approach Compared with Next-Generation Sequencing in Patients with NSCLC.
Сравнительное исследованиеСтатьяПоддержка исследования: вне правительства СШАНазвание и аннотация: Перевод пока не подготовлен
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Kuang Yukun, Xu Peihang, Wang Jiyu, Zheng Yifan, Sun Xue, Li Zimu, Gan RunJing, Li Huixia, Guo Yubiao, Yao Fei, Zhu Changbin, Ke Zunfu, Tang Kejing
Аннотация
BACKGROUND: Precise detection of anaplastic lymphoma kinase (ALK) rearrangement guides the application of ALK-targeted tyrosine kinase inhibitors (ALK-TKIs) in patients with non-small-cell lung cancer (NSCLC). Next-generation sequencing (NGS) has been widely used in clinics, but DNA-based NGS used to detect fusion genes has delivered false-negative results. However, fusion genes can be successfully detected at the transcription level and with higher sensitivity using RNA-based reverse transcription polymerase chain reaction (RT-PCR).
OBJECTIVE: This study compared the performance of RT-PCR and NGS in the detection of echinoderm microtubule-associated protein-like 4 (EML4)-ALK fusion in Chinese patients with NSCLC.
METHODS: Formalin-fixed paraffin-embedded tissues from 153 patients who were pathologically diagnosed as having NSCLC were collected from November 2017 to October 2019. Both DNA/RNA-based NGS and RNA-based RT-PCR were used to detect EML4-ALK fusion. For samples with discordant ALK status results, fluorescence in situ hybridization (FISH) or Sanger sequencing was used to further confirm the ALK status.
RESULTS: In total, 124 samples were successfully analyzed using both NGS and RT-PCR. For 118 samples, results were consistent between NGS and RT-PCR, with 25 reported as ALK fusion positive and 93 as ALK fusion negative, achieving a concordance rate of 95.16%. Among the six samples with disconcordant results, five were positive using RT-PCR but negative using NGS, and one was positive using NGS but negative using RT-PCR. Four of six cases with disconcordant results (three RT-PCR positive and one NGS positive) were successfully validated using either FISH or Sanger sequencing.
CONCLUSIONS: Compared with NGS, RT-PCR appears to be a reliable method of detecting EML4-ALK fusion in patients with NSCLC.
OBJECTIVE: This study compared the performance of RT-PCR and NGS in the detection of echinoderm microtubule-associated protein-like 4 (EML4)-ALK fusion in Chinese patients with NSCLC.
METHODS: Formalin-fixed paraffin-embedded tissues from 153 patients who were pathologically diagnosed as having NSCLC were collected from November 2017 to October 2019. Both DNA/RNA-based NGS and RNA-based RT-PCR were used to detect EML4-ALK fusion. For samples with discordant ALK status results, fluorescence in situ hybridization (FISH) or Sanger sequencing was used to further confirm the ALK status.
RESULTS: In total, 124 samples were successfully analyzed using both NGS and RT-PCR. For 118 samples, results were consistent between NGS and RT-PCR, with 25 reported as ALK fusion positive and 93 as ALK fusion negative, achieving a concordance rate of 95.16%. Among the six samples with disconcordant results, five were positive using RT-PCR but negative using NGS, and one was positive using NGS but negative using RT-PCR. Four of six cases with disconcordant results (three RT-PCR positive and one NGS positive) were successfully validated using either FISH or Sanger sequencing.
CONCLUSIONS: Compared with NGS, RT-PCR appears to be a reliable method of detecting EML4-ALK fusion in patients with NSCLC.
Предметные рубрики MeSH
AdultAgedAged, 80 and overAnaplastic Lymphoma KinaseCarcinoma, Non-Small-Cell LungCell Cycle ProteinsChinaFemaleGene RearrangementHigh-Throughput Nucleotide SequencingHumansIn Situ Hybridization, FluorescenceLung NeoplasmsMaleMicrotubule-Associated ProteinsMiddle AgedOncogene Proteins, FusionReproducibility of ResultsReverse Transcriptase Polymerase Chain ReactionSequence Analysis, DNASerine Endopeptidases