Abstract
Purpose
IHC, FISH, and NGS are common methods of ALK evaluation in NSCLC. The purpose of this study was to investigate whether ALK false positives or false negatives occurred more often in daily routines. An approach to identify ALK fusion was then proposed.
Materials and methods
We analyzed 1815 cases of NSCLC, including 83 (4.6%) ALK IHC positives. Total 182 samples (62 ALK+ and 120 ALK−) were examined via FISH, RT-ddPCR, NGS, RT-qPCR and RNAscope to confirm ALK status.
Results
One ALK FISH false negative was found, which harbored two genomic rearrangements involved in EML4–ALK (exon 13:exon 20) fusion. One ALK IHC false negative was confirmed depending on a rare ALK FISH-positive pattern and ALK RNAscope positive but ALK fusion was not found via NGS. In addition, an atypical ALK FISH-positive pattern was observed in an IHC-positive case with chromosome 2 inversion leading to EML4–ALK (exon 6:exon 20) fusion. EML4–ALK fusion was determined in one case with an atypical FISH patterns by RT-qPCR. Rare complicated genomic rearrangements involved in a novel ALK fusion of EML4–ALK (exon 7:exon 14) were distinguished in an ALK IHC and FISH double-positive case.
Conclusion
False negative of ALK IHC, FISH and NGS results were found in our cohort, but none was false ALK positive. False ALK negatives should be more concerned than false positives. ALK rearrangements with cryptic ALK fusion patterns could be identified using our algorithm. Non-squamous non-small cell lung cancer was recommended for priority detection.
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Abbreviations
- ALK:
-
Anaplastic lymphoma kinase
- NSCLC:
-
Non-small cell lung cancer
- FISH:
-
Fluorescence in situ hybridization
- IHC:
-
Immunohistochemistry
- FDA:
-
Food and Drug Administration
- CFDA:
-
China Food and Drug Administration
- SFDA:
-
State Food and Drug Administration of China
- NMPA:
-
China National Medical Products Administration
- RT-qPCR:
-
Quantitative reverse transcription-PCR
- CAP:
-
College of American Pathologists
- IASLC:
-
International Association for the Study of Lung Cancer
- AMP:
-
Association for Molecular Pathology
- NGS:
-
Next-generation sequencing
- RT-ddPCR:
-
Reverse transcription droplet digital PCR
- FIATA:
-
Fusion-induced asymmetric transcription assay
- FFPE:
-
Formalin-fixed paraffin-embedded
- RNAscope:
-
A method of RNA in situ hybridization
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Acknowledgements
We thank Dr. Linping Lu and Dr. Lingxiang Zhu from TargetingOne Co. and Dr. Xiaoqian Ma and Chao Ren from Geneplus Co. for providing the technical help of RT-ddPCR and NGS testing, respectively.
Funding
This study was supported by the foundation from the National Key Research and Development Program of China (No. 2017YFC1309004), Chinese Academy of Medical Sciences (CAMS) Initiative for Innovative Medicine (No. 2017-I2M-1–005) and CAMS Innovation Fund for Medical Sciences (No. 2019-I2M-2–002).
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All procedures performed in studies involving human participants were in accordance with the ethical standards of the institutional and/or national research committee and with the 1964 Helsinki declaration and its later amendments or comparable ethical standards.
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The retrospective study was approved by the institutional review board of Peking Union Medical College Hospital with a waiver for the need to obtain informed consent.
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Liu, Y., Wu, S., Shi, X. et al. ALK detection in lung cancer: identification of atypical and cryptic ALK rearrangements using an optimal algorithm. J Cancer Res Clin Oncol 146, 1307–1320 (2020). https://doi.org/10.1007/s00432-020-03166-1
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DOI: https://doi.org/10.1007/s00432-020-03166-1