Activation of HER family signaling as a mechanism of acquired resistance to ALK inhibitors in EML4-ALK-positive non-small cell lung cancer

Junko Tanizaki, Isamu Okamoto, Takafumi Okabe, Kazuko Sakai, Kaoru Tanaka, Hidetoshi Hayashi, Hiroyasu Kaneda, Ken Takezawa, Kiyoko Kuwata, Haruka Yamaguchi, Erina Hatashita, Kazuto Nishio, Kazuhiko Nakagawa

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Abstract

Purpose: Anaplastic lymphoma kinase (ALK) tyrosine kinase inhibitors (TKI) such as crizotinib show marked efficacy in patients with non-small cell lung cancer positive for the echinoderm microtubule-associated protein-like 4 (EML4)-ALK fusion protein. However, acquired resistance to these agents has already been described in treated patients, and the mechanisms of such resistance remain largely unknown. Experimental Design: We established lines of EML4-ALK-positive H3122 lung cancer cells that are resistant to the ALK inhibitor TAE684 (H3122/TR cells) and investigated their resistance mechanism with the use of immunoblot analysis, ELISA, reverse transcription and real-time PCR analysis, and an annexin V binding assay. We isolated EML4-ALK-positive lung cancer cells (K-3) from a patient who developed resistance to crizotinib and investigated their characteristics. Results: The expression of EML4-ALK was reduced at the transcriptional level, whereas phosphorylation of epidermal growth factor receptor (EGFR), HER2, and HER3 was upregulated, in H3122/TR cells compared with those in H3122 cells. This activation of HER family proteins was accompanied by increased secretion of EGF. Treatment with an EGFR-TKI induced apoptosis in H3122/TR cells, but not in H3122 cells. The TAE684-induced inhibition of extracellular signal-regulated kinase (ERK) and STAT3 phosphorylation observed in parental cells was prevented by exposure of these cells to exogenous EGF, resulting in a reduced sensitivity of cell growth to TAE684. K-3 cells also manifested HER family activation accompanied by increased EGF secretion. Conclusions: EGF-mediated activation of HER family signaling is associated with ALK-TKI resistance in lung cancer positive for EML4-ALK.

Original languageEnglish
Pages (from-to)6219-6226
Number of pages8
JournalClinical Cancer Research
Volume18
Issue number22
DOIs
Publication statusPublished - Nov 15 2012

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Non-Small Cell Lung Carcinoma
Epidermal Growth Factor
Protein-Tyrosine Kinases
Lung Neoplasms
Epidermal Growth Factor Receptor
MAP4
anaplastic lymphoma kinase
Phosphorylation
Annexin A5
Extracellular Signal-Regulated MAP Kinases
Reverse Transcription
Real-Time Polymerase Chain Reaction
Proteins
Research Design
Enzyme-Linked Immunosorbent Assay
Apoptosis

All Science Journal Classification (ASJC) codes

  • Oncology
  • Cancer Research

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Activation of HER family signaling as a mechanism of acquired resistance to ALK inhibitors in EML4-ALK-positive non-small cell lung cancer. / Tanizaki, Junko; Okamoto, Isamu; Okabe, Takafumi; Sakai, Kazuko; Tanaka, Kaoru; Hayashi, Hidetoshi; Kaneda, Hiroyasu; Takezawa, Ken; Kuwata, Kiyoko; Yamaguchi, Haruka; Hatashita, Erina; Nishio, Kazuto; Nakagawa, Kazuhiko.

In: Clinical Cancer Research, Vol. 18, No. 22, 15.11.2012, p. 6219-6226.

Research output: Contribution to journalArticle

Tanizaki, J, Okamoto, I, Okabe, T, Sakai, K, Tanaka, K, Hayashi, H, Kaneda, H, Takezawa, K, Kuwata, K, Yamaguchi, H, Hatashita, E, Nishio, K & Nakagawa, K 2012, 'Activation of HER family signaling as a mechanism of acquired resistance to ALK inhibitors in EML4-ALK-positive non-small cell lung cancer', Clinical Cancer Research, vol. 18, no. 22, pp. 6219-6226. https://doi.org/10.1158/1078-0432.CCR-12-0392
Tanizaki, Junko ; Okamoto, Isamu ; Okabe, Takafumi ; Sakai, Kazuko ; Tanaka, Kaoru ; Hayashi, Hidetoshi ; Kaneda, Hiroyasu ; Takezawa, Ken ; Kuwata, Kiyoko ; Yamaguchi, Haruka ; Hatashita, Erina ; Nishio, Kazuto ; Nakagawa, Kazuhiko. / Activation of HER family signaling as a mechanism of acquired resistance to ALK inhibitors in EML4-ALK-positive non-small cell lung cancer. In: Clinical Cancer Research. 2012 ; Vol. 18, No. 22. pp. 6219-6226.
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T1 - Activation of HER family signaling as a mechanism of acquired resistance to ALK inhibitors in EML4-ALK-positive non-small cell lung cancer

AU - Tanizaki, Junko

AU - Okamoto, Isamu

AU - Okabe, Takafumi

AU - Sakai, Kazuko

AU - Tanaka, Kaoru

AU - Hayashi, Hidetoshi

AU - Kaneda, Hiroyasu

AU - Takezawa, Ken

AU - Kuwata, Kiyoko

AU - Yamaguchi, Haruka

AU - Hatashita, Erina

AU - Nishio, Kazuto

AU - Nakagawa, Kazuhiko

PY - 2012/11/15

Y1 - 2012/11/15

N2 - Purpose: Anaplastic lymphoma kinase (ALK) tyrosine kinase inhibitors (TKI) such as crizotinib show marked efficacy in patients with non-small cell lung cancer positive for the echinoderm microtubule-associated protein-like 4 (EML4)-ALK fusion protein. However, acquired resistance to these agents has already been described in treated patients, and the mechanisms of such resistance remain largely unknown. Experimental Design: We established lines of EML4-ALK-positive H3122 lung cancer cells that are resistant to the ALK inhibitor TAE684 (H3122/TR cells) and investigated their resistance mechanism with the use of immunoblot analysis, ELISA, reverse transcription and real-time PCR analysis, and an annexin V binding assay. We isolated EML4-ALK-positive lung cancer cells (K-3) from a patient who developed resistance to crizotinib and investigated their characteristics. Results: The expression of EML4-ALK was reduced at the transcriptional level, whereas phosphorylation of epidermal growth factor receptor (EGFR), HER2, and HER3 was upregulated, in H3122/TR cells compared with those in H3122 cells. This activation of HER family proteins was accompanied by increased secretion of EGF. Treatment with an EGFR-TKI induced apoptosis in H3122/TR cells, but not in H3122 cells. The TAE684-induced inhibition of extracellular signal-regulated kinase (ERK) and STAT3 phosphorylation observed in parental cells was prevented by exposure of these cells to exogenous EGF, resulting in a reduced sensitivity of cell growth to TAE684. K-3 cells also manifested HER family activation accompanied by increased EGF secretion. Conclusions: EGF-mediated activation of HER family signaling is associated with ALK-TKI resistance in lung cancer positive for EML4-ALK.

AB - Purpose: Anaplastic lymphoma kinase (ALK) tyrosine kinase inhibitors (TKI) such as crizotinib show marked efficacy in patients with non-small cell lung cancer positive for the echinoderm microtubule-associated protein-like 4 (EML4)-ALK fusion protein. However, acquired resistance to these agents has already been described in treated patients, and the mechanisms of such resistance remain largely unknown. Experimental Design: We established lines of EML4-ALK-positive H3122 lung cancer cells that are resistant to the ALK inhibitor TAE684 (H3122/TR cells) and investigated their resistance mechanism with the use of immunoblot analysis, ELISA, reverse transcription and real-time PCR analysis, and an annexin V binding assay. We isolated EML4-ALK-positive lung cancer cells (K-3) from a patient who developed resistance to crizotinib and investigated their characteristics. Results: The expression of EML4-ALK was reduced at the transcriptional level, whereas phosphorylation of epidermal growth factor receptor (EGFR), HER2, and HER3 was upregulated, in H3122/TR cells compared with those in H3122 cells. This activation of HER family proteins was accompanied by increased secretion of EGF. Treatment with an EGFR-TKI induced apoptosis in H3122/TR cells, but not in H3122 cells. The TAE684-induced inhibition of extracellular signal-regulated kinase (ERK) and STAT3 phosphorylation observed in parental cells was prevented by exposure of these cells to exogenous EGF, resulting in a reduced sensitivity of cell growth to TAE684. K-3 cells also manifested HER family activation accompanied by increased EGF secretion. Conclusions: EGF-mediated activation of HER family signaling is associated with ALK-TKI resistance in lung cancer positive for EML4-ALK.

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