RNA-based analysis of ALK fusions in non-small cell lung cancer cases showing IHC/FISH discordance
Abstract Background Rearrangements of the anaplastic lymphoma kinase (ALK) belong to the promising targets in the therapy of advanced non-small cell lung cancer (NSCLC) and are predominantly detected by immunohistochemistry (IHC) and/or fluorescence in-situ hybridization (FISH). However, both method...
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| Format: | Article |
| Language: | English |
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BMC
2018-11-01
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| Series: | BMC Cancer |
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| Online Access: | http://link.springer.com/article/10.1186/s12885-018-5070-6 |
| _version_ | 1811218849285013504 |
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| author | Claudia Vollbrecht Dido Lenze Michael Hummel Annika Lehmann Markus Moebs Nikolaj Frost Philipp Jurmeister Leonille Schweizer Udo Kellner Manfred Dietel Maximilian von Laffert |
| author_facet | Claudia Vollbrecht Dido Lenze Michael Hummel Annika Lehmann Markus Moebs Nikolaj Frost Philipp Jurmeister Leonille Schweizer Udo Kellner Manfred Dietel Maximilian von Laffert |
| author_sort | Claudia Vollbrecht |
| collection | DOAJ |
| description | Abstract Background Rearrangements of the anaplastic lymphoma kinase (ALK) belong to the promising targets in the therapy of advanced non-small cell lung cancer (NSCLC) and are predominantly detected by immunohistochemistry (IHC) and/or fluorescence in-situ hybridization (FISH). However, both methods occasionally produce discordant results, especially in so-called borderline (BL) cases, showing ALK FISH-positive signals in 10–20% of the tumor nuclei around the cutoff (15%). This leads to a diagnostic and thus to a therapeutic dilemma. Methods We selected 18 unequivocal (12 ALK IHC/FISH-negative; 6 ALK IHC/FISH-positive) and 15 equivocal samples with discordant results between FISH (Abbott, Vysis LSI ALK Dual Color) and IHC (Ventana, D5F3), including cases with FISH-BL results, for further RNA based-analysis. To detect ALK rearrangement at the transcriptional level, RNA was analyzed using a targeted multiplex-PCR panel followed by IonTorrent sequencing and by direct transcript counting using a digital probe-based assay (NanoString). Sensitivity of both methods was defined using RNA obtained from an ALK-positive cell line dilution series. Results Cases with unequivocal IHC/FISH results showed concordant data with both RNA-based methods, whereas the three IHC-negative/FISH-positive samples were negative. The four IHC-negative/FISH-BL-negative cases, as well as the five IHC-negative/FISH-BL-positive samples showed negative results by massive parallel sequencing (MPS) and digital probe-based assay. The two IHC-positive/FISH-BL-positive cases were both positive on the RNA-level, whereas a tumor with questionable IHC and FISH-BL-positive status displayed no ALK fusion transcript. Conclusions The comparison of methods for the confirmation of ALK rearrangements revealed that the detection of ALK protein by IHC and ALK fusion transcripts on transcriptional level by MPS and the probe-based assay leads to concordant results. Only a small proportion of clearly ALK FISH-positive cases are unable to express the ALK protein and ALK fusion transcript which might explain a non-responding to ALK inhibitors. Therefore, our findings led us to conclude that ALK testing should initially be based on IHC and/or RNA-based methods. |
| first_indexed | 2024-04-12T07:15:14Z |
| format | Article |
| id | doaj.art-af72e0ed2bf34fe08603e87cd561159e |
| institution | Directory Open Access Journal |
| issn | 1471-2407 |
| language | English |
| last_indexed | 2024-04-12T07:15:14Z |
| publishDate | 2018-11-01 |
| publisher | BMC |
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| series | BMC Cancer |
| spelling | doaj.art-af72e0ed2bf34fe08603e87cd561159e2022-12-22T03:42:28ZengBMCBMC Cancer1471-24072018-11-011811910.1186/s12885-018-5070-6RNA-based analysis of ALK fusions in non-small cell lung cancer cases showing IHC/FISH discordanceClaudia Vollbrecht0Dido Lenze1Michael Hummel2Annika Lehmann3Markus Moebs4Nikolaj Frost5Philipp Jurmeister6Leonille Schweizer7Udo Kellner8Manfred Dietel9Maximilian von Laffert10Charité-Universitätsmedizin Berlin corporate member of Freie Universität Berlin, Humboldt-Universität zu Berlin, and Berlin Institute of Health, Institute of PathologyCharité-Universitätsmedizin Berlin corporate member of Freie Universität Berlin, Humboldt-Universität zu Berlin, and Berlin Institute of Health, Institute of PathologyCharité-Universitätsmedizin Berlin corporate member of Freie Universität Berlin, Humboldt-Universität zu Berlin, and Berlin Institute of Health, Institute of PathologyCharité-Universitätsmedizin Berlin corporate member of Freie Universität Berlin, Humboldt-Universität zu Berlin, and Berlin Institute of Health, Institute of PathologyCharité-Universitätsmedizin Berlin corporate member of Freie Universität Berlin, Humboldt-Universität zu Berlin, and Berlin Institute of Health, Institute of PathologyMedical Department, Division of Infectiology and Pneumology, Charité-Universitätsmedizin Berlin corporate member of Freie Universität Berlin, Humboldt-Universität zu Berlin, and Berlin Institute of HealthCharité-Universitätsmedizin Berlin corporate member of Freie Universität Berlin, Humboldt-Universität zu Berlin, and Berlin Institute of Health, Institute of PathologyCharité-Universitätsmedizin Berlin corporate member of Freie Universität Berlin, Humboldt-Universität zu Berlin, and Berlin Institute of Health, Institute of PathologyJohannes Wesling Klinikum Minden, Institute for PathologyCharité-Universitätsmedizin Berlin corporate member of Freie Universität Berlin, Humboldt-Universität zu Berlin, and Berlin Institute of Health, Institute of PathologyCharité-Universitätsmedizin Berlin corporate member of Freie Universität Berlin, Humboldt-Universität zu Berlin, and Berlin Institute of Health, Institute of PathologyAbstract Background Rearrangements of the anaplastic lymphoma kinase (ALK) belong to the promising targets in the therapy of advanced non-small cell lung cancer (NSCLC) and are predominantly detected by immunohistochemistry (IHC) and/or fluorescence in-situ hybridization (FISH). However, both methods occasionally produce discordant results, especially in so-called borderline (BL) cases, showing ALK FISH-positive signals in 10–20% of the tumor nuclei around the cutoff (15%). This leads to a diagnostic and thus to a therapeutic dilemma. Methods We selected 18 unequivocal (12 ALK IHC/FISH-negative; 6 ALK IHC/FISH-positive) and 15 equivocal samples with discordant results between FISH (Abbott, Vysis LSI ALK Dual Color) and IHC (Ventana, D5F3), including cases with FISH-BL results, for further RNA based-analysis. To detect ALK rearrangement at the transcriptional level, RNA was analyzed using a targeted multiplex-PCR panel followed by IonTorrent sequencing and by direct transcript counting using a digital probe-based assay (NanoString). Sensitivity of both methods was defined using RNA obtained from an ALK-positive cell line dilution series. Results Cases with unequivocal IHC/FISH results showed concordant data with both RNA-based methods, whereas the three IHC-negative/FISH-positive samples were negative. The four IHC-negative/FISH-BL-negative cases, as well as the five IHC-negative/FISH-BL-positive samples showed negative results by massive parallel sequencing (MPS) and digital probe-based assay. The two IHC-positive/FISH-BL-positive cases were both positive on the RNA-level, whereas a tumor with questionable IHC and FISH-BL-positive status displayed no ALK fusion transcript. Conclusions The comparison of methods for the confirmation of ALK rearrangements revealed that the detection of ALK protein by IHC and ALK fusion transcripts on transcriptional level by MPS and the probe-based assay leads to concordant results. Only a small proportion of clearly ALK FISH-positive cases are unable to express the ALK protein and ALK fusion transcript which might explain a non-responding to ALK inhibitors. Therefore, our findings led us to conclude that ALK testing should initially be based on IHC and/or RNA-based methods.http://link.springer.com/article/10.1186/s12885-018-5070-6Non-small cell lung cancer (NSCLC)Anaplastic lymphoma kinase (ALK)Fluorescence in-situ hybridization (FISH)Immunohistochemistry (IHC)Massive parallel sequencing (MPS)NanoString |
| spellingShingle | Claudia Vollbrecht Dido Lenze Michael Hummel Annika Lehmann Markus Moebs Nikolaj Frost Philipp Jurmeister Leonille Schweizer Udo Kellner Manfred Dietel Maximilian von Laffert RNA-based analysis of ALK fusions in non-small cell lung cancer cases showing IHC/FISH discordance BMC Cancer Non-small cell lung cancer (NSCLC) Anaplastic lymphoma kinase (ALK) Fluorescence in-situ hybridization (FISH) Immunohistochemistry (IHC) Massive parallel sequencing (MPS) NanoString |
| title | RNA-based analysis of ALK fusions in non-small cell lung cancer cases showing IHC/FISH discordance |
| title_full | RNA-based analysis of ALK fusions in non-small cell lung cancer cases showing IHC/FISH discordance |
| title_fullStr | RNA-based analysis of ALK fusions in non-small cell lung cancer cases showing IHC/FISH discordance |
| title_full_unstemmed | RNA-based analysis of ALK fusions in non-small cell lung cancer cases showing IHC/FISH discordance |
| title_short | RNA-based analysis of ALK fusions in non-small cell lung cancer cases showing IHC/FISH discordance |
| title_sort | rna based analysis of alk fusions in non small cell lung cancer cases showing ihc fish discordance |
| topic | Non-small cell lung cancer (NSCLC) Anaplastic lymphoma kinase (ALK) Fluorescence in-situ hybridization (FISH) Immunohistochemistry (IHC) Massive parallel sequencing (MPS) NanoString |
| url | http://link.springer.com/article/10.1186/s12885-018-5070-6 |
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