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Detecting anaplastic lymphoma kinase (ALK) gene rearrangements with next-generation sequencing remains a reliable approach in patients with non-small-cell lung cancer

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Abstract

Next-generation sequencing (NGS) is rapidly becoming routine in clinical oncology practice to identify therapeutic biomarkers, including gene rearrangements in anaplastic lymphoma kinase (ALK). Our study investigated the concordance of ALK positivity evaluated by DNA-based NGS with orthogonal ALK testing methods such as fluorescence in situ hybridization (FISH), immunohistochemistry (IHC), and RNA-based NGS (RNA-NGS). Thirty-eight patients with lung adenocarcinoma who were detected with ALK rearrangements using DNA-NGS and also had adequate tissue samples submitted for FISH, IHC, and RNA-NGS, were included in this study. Of the 38 patients, RNA samples from 3 patients failed quality control for RNA-NGS. The concordance of ALK positivity was calculated relative to DNA-NGS results. The concordance rates were 97.1% (34/35) for RNA-NGS, 94.7% (36/38) for IHC, and 97.4% (37/38) for FISH. DNA-NGS detected single ALK rearrangements in 14 (35.0%) patients and complex ALK rearrangements in 26 (65.0%). RNA-NGS detected only single transcripts of the primary ALK fusions. A novel LANCL1-ALK (L7:A20) detected using DNA-NGS was detected as EML4-ALK (E13:A20) transcripts using RNA-NGS. Interestingly, patients with single ALK rearrangements were more likely to be detected with atypical isolated red signals (p < 0.001), while patients with complex ALK rearrangements were more likely to be detected with atypical split red and green signals less than 2 signal diameters apart (p < 0.001). Our study highlights the reliability of NGS in the accurate detection of specific ALK fusion variants and concomitant mutations that are crucial for individualized treatment decisions in patients with lung cancer.

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Data availability

All authors confirm adherence to the policy. All datasets used and/or analyzed during the current study are available from the corresponding author on reasonable request.

Abbreviations

ALK:

Anaplastic lymphoma kinase

EML4:

Echinoderm microtubule-associated protein-like 4

FDA:

Food and Drug Administration

FFPE:

Formalin-fixed, paraffin-embedded

FISH:

Fluorescence in situ hybridization

IHC:

Immunohistochemistry

NGS:

Next-generation sequencing

NSCLC:

Non-small-cell lung cancer

qRT-PCR:

Quantitative reverse transcriptase-polymerase chain reaction

TKI:

Tyrosine kinase inhibitor

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Acknowledgements

The authors would like to thank all the patients and their families for their cooperation and support. We also thank the investigators, study coordinators, operation staff, and the whole project team who worked on this project. We would also like to extend our utmost appreciation for the active support of the staff of Burning Rock Biotech, particularly Drs. Analyn Lizaso, Dejian Gu, Bing Li, Haiyan Li, Wenjie Sun, Zhou Zhang, and Jing Lin.

Funding

This study was supported by grants from the National Natural Science Foundation of China (82172991, 81502089 to Y.D., 81773109 to Z.Z.), the Natural Science Foundation of Jiangsu Province (BK20151024 to Y.D., BK20151582 to Z.Z.), and the Natural Science Research Program for Higher Education in Jiangsu Province (21KJB320015 to Y.D.).

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Authors and Affiliations

  1. Department of Pathology, the First Affiliated Hospital of Nanjing Medical University, Nanjing, 210029, China

    Ying Ding, Chen Miao, Xiao He & Zhi-Hong Zhang

  2. The First School of Clinical Medicine, Nanjing Medical University, Nanjing, 210029, China

    Chang Sun & Wei Su

  3. Department of Pathology, Nanjing First Hospital, Nanjing Medical University, Nanjing, 210000, China

    Jin-Song Wang

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  1. Ying Ding
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  2. Chang Sun
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  3. Wei Su
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Contributions

Zhi-Hong Zhang and Ying Ding conceived the study and drafted the manuscript. Zhi-Hong Zhang and Ying Ding acquired funding. All the authors collected and analyzed the clinical data and participated in the data interpretation, data analysis, and manuscript writing and editing. All the authors contributed in the revision and approved the final manuscript.

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Correspondence to Zhi-Hong Zhang.

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All procedures performed in studies involving human participants were performed in accordance with the Declaration of Helsinki. This study was approved by the Ethics Committee of the First Affiliated Hospital of Nanjing Medical University.

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The authors declare no competing interests.

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Ding, Y., Sun, C., Su, W. et al. Detecting anaplastic lymphoma kinase (ALK) gene rearrangements with next-generation sequencing remains a reliable approach in patients with non-small-cell lung cancer. Virchows Arch 481, 405–419 (2022). https://doi.org/10.1007/s00428-022-03339-y

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