Skip to main content
Top
Published in: BMC Pulmonary Medicine 1/2017

Open Access 01-12-2017 | Research article

Detection of ALK fusion transcripts in FFPE lung cancer samples by NanoString technology

Authors: Adriane F. Evangelista, Maicon F. Zanon, Adriana Cruvinel Carloni, Flávia E. de Paula, Mariana Andozia Morini, Maressa Ferreira-Neto, Iberê Cauduro Soares, Jose Elias Miziara, Pedro de Marchi, Cristovam Scapulatempo-Neto, Rui M. Reis

Published in: BMC Pulmonary Medicine | Issue 1/2017

Login to get access

Abstract

Background

ALK-rearranged lung cancers exhibit specific pathologic and clinical features and are responsive to anti-ALK therapies. Therefore, the detection of ALK-rearrangement is fundamental for personalized lung cancer therapy. Recently, new molecular techniques, such as NanoString nCounter, have been developed to detect ALK fusions with more accuracy and sensitivity.

Methods

In the present study, we intended to validate a NanoString nCounter ALK-fusion panel in routine biopsies of FFPE lung cancer patients. A total of 43 samples were analyzed, 13 ALK-positive and 30 ALK-negative, as previously detected by FISH and/or immunohistochemistry.

Results

The NanoString panel detected the presence of the EML4-ALK, KIF5B-ALK and TFG-ALK fusion variants. We observed that all the 13 ALK-positive cases exhibited genetic aberrations by the NanoString methodology. Namely, six cases (46.15%) presented EML-ALK variant 1, two (15.38%) presented EML-ALK variant 2, two (15.38%) presented EML-ALK variant 3a, and three (23.07%) exhibited no variant but presented unbalanced expression between 5’/3’ exons, similar to other positive samples. Importantly, for all these analyses, the initial input of RNA was 100 ng, and some cases displayed poor RNA quality measurements.

Conclusions

In this study, we reported the great utility of NanoString technology in the assessment of ALK fusions in routine lung biopsies of FFPE specimens.
Appendix
Available only for authorised users
Literature
1.
go back to reference Morris SW, Kirstein MN, Valentine MB, Dittmer KG, Shapiro DN, Saltman DL, et al. Fusion of a kinase gene, ALK, to a nucleolar protein gene, NPM, in non-Hodgkin’s lymphoma. Science. 1994;263:1281–4.CrossRefPubMed Morris SW, Kirstein MN, Valentine MB, Dittmer KG, Shapiro DN, Saltman DL, et al. Fusion of a kinase gene, ALK, to a nucleolar protein gene, NPM, in non-Hodgkin’s lymphoma. Science. 1994;263:1281–4.CrossRefPubMed
2.
go back to reference Soda M, Choi YL, Enomoto M, Takada S, Yamashita Y, Ishikawa S, et al. Identification of the transforming EML4-ALK fusion gene in non-small-cell lung cancer. Nature. 2007;448:561–6.CrossRefPubMed Soda M, Choi YL, Enomoto M, Takada S, Yamashita Y, Ishikawa S, et al. Identification of the transforming EML4-ALK fusion gene in non-small-cell lung cancer. Nature. 2007;448:561–6.CrossRefPubMed
3.
go back to reference Wong DW-S, Leung EL-H, So KK-T, Tam IY-S, Sihoe AD-L, Cheng L-C, et al. The EML4-ALK fusion gene is involved in various histologic types of lung cancers from nonsmokers with wild-type EGFR and KRAS. Cancer. 2009;115:1723–33.CrossRefPubMed Wong DW-S, Leung EL-H, So KK-T, Tam IY-S, Sihoe AD-L, Cheng L-C, et al. The EML4-ALK fusion gene is involved in various histologic types of lung cancers from nonsmokers with wild-type EGFR and KRAS. Cancer. 2009;115:1723–33.CrossRefPubMed
4.
go back to reference Takeuchi K, Choi YL, Togashi Y, Soda M, Hatano S, Inamura K, et al. KIF5B-ALK, a novel fusion oncokinase identified by an immunohistochemistry-based diagnostic system for ALK-positive lung cancer. Clin Cancer Res. 2009;15:3143–9.CrossRefPubMed Takeuchi K, Choi YL, Togashi Y, Soda M, Hatano S, Inamura K, et al. KIF5B-ALK, a novel fusion oncokinase identified by an immunohistochemistry-based diagnostic system for ALK-positive lung cancer. Clin Cancer Res. 2009;15:3143–9.CrossRefPubMed
5.
go back to reference Rikova K, Guo A, Zeng Q, Possemato A, Yu J, Haack H, et al. Global Survey of Phosphotyrosine Signaling Identifies Oncogenic Kinases in Lung Cancer. Cell. 2007;131:1190–203.CrossRefPubMed Rikova K, Guo A, Zeng Q, Possemato A, Yu J, Haack H, et al. Global Survey of Phosphotyrosine Signaling Identifies Oncogenic Kinases in Lung Cancer. Cell. 2007;131:1190–203.CrossRefPubMed
6.
go back to reference Roskoski R. Anaplastic lymphoma kinase (ALK): structure, oncogenic activation, and pharmacological inhibition. Pharmacol Res. 2013;68:68–94.CrossRefPubMed Roskoski R. Anaplastic lymphoma kinase (ALK): structure, oncogenic activation, and pharmacological inhibition. Pharmacol Res. 2013;68:68–94.CrossRefPubMed
7.
go back to reference Mossé YP, Wood A, Maris JM. Inhibition of ALK signaling for cancer therapy. Clin Cancer Res. 2009;15:5609–14.CrossRefPubMed Mossé YP, Wood A, Maris JM. Inhibition of ALK signaling for cancer therapy. Clin Cancer Res. 2009;15:5609–14.CrossRefPubMed
8.
go back to reference Gaughan EM, Costa DB. Genotype-driven therapies for non-small cell lung cancer: focus on EGFR, KRAS and ALK gene abnormalities. Ther Adv Med Oncol. 2011;3:113–25.CrossRefPubMedPubMedCentral Gaughan EM, Costa DB. Genotype-driven therapies for non-small cell lung cancer: focus on EGFR, KRAS and ALK gene abnormalities. Ther Adv Med Oncol. 2011;3:113–25.CrossRefPubMedPubMedCentral
9.
go back to reference Shaw AT, Yeap BY, Mino-Kenudson M, Digumarthy SR, Costa DB, Heist RS, et al. Clinical features and outcome of patients with non-small-cell lung cancer who harbor EML4-ALK. J Clin Oncol. 2009;27:4247–53.CrossRefPubMedPubMedCentral Shaw AT, Yeap BY, Mino-Kenudson M, Digumarthy SR, Costa DB, Heist RS, et al. Clinical features and outcome of patients with non-small-cell lung cancer who harbor EML4-ALK. J Clin Oncol. 2009;27:4247–53.CrossRefPubMedPubMedCentral
10.
go back to reference McKeage K. Alectinib: A Review of Its Use in Advanced ALK-Rearranged Non-Small Cell Lung Cancer. Drugs. 2014;75:75–82.CrossRef McKeage K. Alectinib: A Review of Its Use in Advanced ALK-Rearranged Non-Small Cell Lung Cancer. Drugs. 2014;75:75–82.CrossRef
13.
go back to reference Lira ME, Kim TM, Huang D, Deng S, Koh Y, Jang B, et al. Multiplexed gene expression and fusion transcript analysis to detect ALK fusions in lung cancer. J Mol Diagn. 2013;15:51–61.CrossRefPubMed Lira ME, Kim TM, Huang D, Deng S, Koh Y, Jang B, et al. Multiplexed gene expression and fusion transcript analysis to detect ALK fusions in lung cancer. J Mol Diagn. 2013;15:51–61.CrossRefPubMed
14.
go back to reference Kulkarni MM. Digital multiplexed gene expression analysis using the NanoString nCounter system. Curr Protoc Mol Biol. 2011;25:Unit25B.10.PubMed Kulkarni MM. Digital multiplexed gene expression analysis using the NanoString nCounter system. Curr Protoc Mol Biol. 2011;25:Unit25B.10.PubMed
15.
go back to reference Yamane LS, Scapulatempo-Neto C, Alvarenga L, Oliveira CZ, Berardinelli GN, Almodova E, et al. KRAS and BRAF mutations and MSI status in precursor lesions of colorectal cancer detected by colonoscopy. Oncol Rep. 2014;32:1419–26.PubMed Yamane LS, Scapulatempo-Neto C, Alvarenga L, Oliveira CZ, Berardinelli GN, Almodova E, et al. KRAS and BRAF mutations and MSI status in precursor lesions of colorectal cancer detected by colonoscopy. Oncol Rep. 2014;32:1419–26.PubMed
16.
go back to reference Becker AP, Scapulatempo-Neto C, Carloni AC, Paulino A, Sheren J, Aisner DL, et al. KIAA1549: BRAF Gene Fusion and FGFR1 Hotspot Mutations Are Prognostic Factors in Pilocytic Astrocytomas. J Neuropathol Exp Neurol. 2015;74:743–54.CrossRefPubMedPubMedCentral Becker AP, Scapulatempo-Neto C, Carloni AC, Paulino A, Sheren J, Aisner DL, et al. KIAA1549: BRAF Gene Fusion and FGFR1 Hotspot Mutations Are Prognostic Factors in Pilocytic Astrocytomas. J Neuropathol Exp Neurol. 2015;74:743–54.CrossRefPubMedPubMedCentral
17.
go back to reference Reis-Filho JS, Pinheiro C, Lambros MBK, Milanezi F, Carvalho S, Savage K, et al. EGFR amplification and lack of activating mutations in metaplastic breast carcinomas. J Pathol. 2006;209:445–53.CrossRefPubMed Reis-Filho JS, Pinheiro C, Lambros MBK, Milanezi F, Carvalho S, Savage K, et al. EGFR amplification and lack of activating mutations in metaplastic breast carcinomas. J Pathol. 2006;209:445–53.CrossRefPubMed
18.
go back to reference Camidge DR, Kono SA, Flacco A, Tan A-C, Doebele RC, Zhou Q, et al. Optimizing the detection of lung cancer patients harboring anaplastic lymphoma kinase (ALK) gene rearrangements potentially suitable for ALK inhibitor treatment. Clin Cancer Res. 2010;16:5581–90.CrossRefPubMedPubMedCentral Camidge DR, Kono SA, Flacco A, Tan A-C, Doebele RC, Zhou Q, et al. Optimizing the detection of lung cancer patients harboring anaplastic lymphoma kinase (ALK) gene rearrangements potentially suitable for ALK inhibitor treatment. Clin Cancer Res. 2010;16:5581–90.CrossRefPubMedPubMedCentral
19.
go back to reference Kwak EL, Bang Y-J, Camidge DR, Shaw AT, Solomon B, Maki RG, et al. Anaplastic lymphoma kinase inhibition in non-small-cell lung cancer. N Engl J Med. 2010;363:1693–703.CrossRefPubMedPubMedCentral Kwak EL, Bang Y-J, Camidge DR, Shaw AT, Solomon B, Maki RG, et al. Anaplastic lymphoma kinase inhibition in non-small-cell lung cancer. N Engl J Med. 2010;363:1693–703.CrossRefPubMedPubMedCentral
20.
go back to reference Wynes MW, Sholl LM, Dietel M, Schuuring E, Tsao MS, Yatabe Y, et al. An international interpretation study using the ALK IHC antibody D5F3 and a sensitive detection kit demonstrates high concordance between ALK IHC and ALK FISH and between evaluators. J Thorac Oncol. 2014;9:631–8.CrossRefPubMedPubMedCentral Wynes MW, Sholl LM, Dietel M, Schuuring E, Tsao MS, Yatabe Y, et al. An international interpretation study using the ALK IHC antibody D5F3 and a sensitive detection kit demonstrates high concordance between ALK IHC and ALK FISH and between evaluators. J Thorac Oncol. 2014;9:631–8.CrossRefPubMedPubMedCentral
21.
go back to reference Wilson FH, Johannessen CM, Piccioni F, Tamayo P, Kim JW, Van Allen EM, et al. A functional landscape of resistance to ALK inhibition in lung cancer. Cancer Cell. 2015;27:397–408.CrossRefPubMedPubMedCentral Wilson FH, Johannessen CM, Piccioni F, Tamayo P, Kim JW, Van Allen EM, et al. A functional landscape of resistance to ALK inhibition in lung cancer. Cancer Cell. 2015;27:397–408.CrossRefPubMedPubMedCentral
22.
go back to reference Heuckmann JM, Balke-Want H, Malchers F, Peifer M, Sos ML, Koker M, et al. Differential protein stability and ALK inhibitor sensitivity of EML4-ALK fusion variants. Clin Cancer Res. 2012;18:4682–90.CrossRefPubMed Heuckmann JM, Balke-Want H, Malchers F, Peifer M, Sos ML, Koker M, et al. Differential protein stability and ALK inhibitor sensitivity of EML4-ALK fusion variants. Clin Cancer Res. 2012;18:4682–90.CrossRefPubMed
23.
go back to reference Crystal AS, Shaw AT. Variants on a theme: a biomarker of crizotinib response in ALK-positive non-small cell lung cancer? Clin Cancer Res. 2012;18:4479–81.CrossRefPubMed Crystal AS, Shaw AT. Variants on a theme: a biomarker of crizotinib response in ALK-positive non-small cell lung cancer? Clin Cancer Res. 2012;18:4479–81.CrossRefPubMed
24.
go back to reference Rogers T-M, Arnau GM, Ryland GL, Huang S, Lira ME, Emmanuel Y, et al. Multiplexed transcriptome analysis to detect ALK, ROS1 and RET rearrangements in lung cancer. Sci Rep. 2017;7:42259.CrossRefPubMedPubMedCentral Rogers T-M, Arnau GM, Ryland GL, Huang S, Lira ME, Emmanuel Y, et al. Multiplexed transcriptome analysis to detect ALK, ROS1 and RET rearrangements in lung cancer. Sci Rep. 2017;7:42259.CrossRefPubMedPubMedCentral
25.
go back to reference Ali SM, Hensing T, Schrock AB, Allen J, Sanford E, Gowen K, et al. Comprehensive Genomic Profiling Identifies a Subset of Crizotinib-Responsive ALK-Rearranged Non-Small Cell Lung Cancer Not Detected by Fluorescence In Situ Hybridization. Oncologist. 2016;21:762–70.CrossRefPubMed Ali SM, Hensing T, Schrock AB, Allen J, Sanford E, Gowen K, et al. Comprehensive Genomic Profiling Identifies a Subset of Crizotinib-Responsive ALK-Rearranged Non-Small Cell Lung Cancer Not Detected by Fluorescence In Situ Hybridization. Oncologist. 2016;21:762–70.CrossRefPubMed
26.
Metadata
Title
Detection of ALK fusion transcripts in FFPE lung cancer samples by NanoString technology
Authors
Adriane F. Evangelista
Maicon F. Zanon
Adriana Cruvinel Carloni
Flávia E. de Paula
Mariana Andozia Morini
Maressa Ferreira-Neto
Iberê Cauduro Soares
Jose Elias Miziara
Pedro de Marchi
Cristovam Scapulatempo-Neto
Rui M. Reis
Publication date
01-12-2017
Publisher
BioMed Central
Published in
BMC Pulmonary Medicine / Issue 1/2017
Electronic ISSN: 1471-2466
DOI
https://doi.org/10.1186/s12890-017-0428-0

Other articles of this Issue 1/2017

BMC Pulmonary Medicine 1/2017 Go to the issue