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Phosphoproteomic analysis identifies activated MET-axis PI3K/AKT and MAPK/ERK in lapatinib-resistant cancer cell line.

ABSTRACT: Lapatinib, a dual inhibitor of epidermal growth factor receptor (EGFR) and human epidermal growth factor receptor 2 (HER2) tyrosine kinases, has shown promising results as a growth inhibitor of HER2-positive cancer cells in vitro. However, similar to other EGFR-targeting drugs, acquired resistance to lapatinib by HER2-positive cancer cells remains a major clinical challenge. To elucidate resistance mechanisms to EGFR/HER2-targeting agents, we performed a systematic quantitative comparison of the phosphoproteome of lapatinib-resistant (LR) human gastric cancer cells (SNU216-LR) versus parental cells (SNU216) using a titanium dioxide (TiO2) phosphopeptide enrichment method and analysis with a Q-Exactive hybrid quadrupole-Orbitrap mass spectrometer. Biological network analysis of differentially expressed phosphoproteins revealed apparent constitutive activation of the MET-axis phosphatidylinositide 3-kinase (PI3K)/?-serine/threonine-protein kinase (AKT) and mitogen-activated protein kinase (MAPK)/extracellular signal-regulated kinase (ERK) signaling pathways in SNU216-LR. Inhibition of the PI3K/AKT and MAPK/ERK signaling pathways in SNU216-LR also leads to cell cycle arrest, confirming the biological network analysis. Lapatinib sensitivity was restored when cells were treated with several molecular targeting agents in combination with lapatinib. Thus, by integrating phosphoproteomic data, protein networks and effects of signaling pathway modulation on cell proliferation, we found that SNU216-LR maintains constitutive activation of the PI3K/AKT and MAPK/ERK pathways in a MET-dependent manner. These findings suggest that pathway activation is a key compensatory intracellular phospho-signaling event that may govern gastric cancer cell resistance to drug treatment.

SUBMITTER: Lee YY

PROVIDER: S-EPMC3849569 | biostudies-literature | 2013 Nov

REPOSITORIES: biostudies-literature

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Phosphoproteomic analysis identifies activated MET-axis PI3K/AKT and MAPK/ERK in lapatinib-resistant cancer cell line.

Lee Yong Yook YY Kim Hwang-Phill HP Kang Min Jueng MJ Cho Byoung-Kyu BK Han Sae-Won SW Kim Tae-You TY Yi Eugene C EC

Experimental & molecular medicine 20131122

Lapatinib, a dual inhibitor of epidermal growth factor receptor (EGFR) and human epidermal growth factor receptor 2 (HER2) tyrosine kinases, has shown promising results as a growth inhibitor of HER2-positive cancer cells in vitro. However, similar to other EGFR-targeting drugs, acquired resistance to lapatinib by HER2-positive cancer cells remains a major clinical challenge. To elucidate resistance mechanisms to EGFR/HER2-targeting agents, we performed a systematic quantitative comparison of the ...[more]

PMID: 24263233

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Project description:BackgroundThe repair of cranio-maxillofacial bone defects remains a formidable clinical challenge. The Ets variant 2 (ETV2) transcription factor, which belongs to the E26 transformation-specific (ETS) family, has been reported to play a key role in neovascularization. However, the role of ETV2 in the osteogenesis of human dental pulp stem cells (hDPSCs) remains unexplored.MethodsTransgenic overexpression of ETV2 was achieved using a lentiviral vector, based on a Dox-inducible system. The effects of Dox-induced overexpression of ETV2 on the osteogenesis of hDPSCs were evaluated by quantitative real-time polymerase chain reaction (qRT-PCR), western blot, immunofluorescence staining, alkaline phosphatase (ALP) staining, and Alizarin Red S (ARS) staining. Additionally, RNA-sequencing (RNA-Seq) analysis was performed to analyze the underlying mechanisms of ETV2-induced osteogenesis. Additionally, the role of ETV2 overexpression in bone formation in vivo was validated by animal studies with a rat calvarial defect model and a nude mice model.ResultsOur results demonstrated that ETV2 overexpression significantly upregulated the mRNA and protein expression levels of osteogenic markers, markedly enhanced ALP activity, and promoted matrix mineralization of hDPSCs. Moreover, the results of RNA-Seq analysis and western blot showed that the ERK/MAPK and PI3K-Akt signaling pathways were activated upon transgenic overexpression of ETV2. The enhanced osteogenic differentiation of hDPSCs due to ETV2 overexpression was partially reversed by treatment with inhibitors of ERK/MAPK or PI3K-AKT signaling. Furthermore, the results of in vivo studies demonstrated that ETV2 overexpression improved bone healing in a rat calvarial defect model and increased ectopic bone formation in nude mice.ConclusionsCollectively, our results indicated that ETV2 overexpression exerted positive effects on the osteogenesis of hDPSCs, at least partially via the ERK/MAPK and PI3K/AKT signaling pathways.

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Phosphoproteomic analysis identifies activated MET-axis PI3K/AKT and MAPK/ERK in lapatinib-resistant cancer cell line.

Publications

Phosphoproteomic analysis identifies activated MET-axis PI3K/AKT and MAPK/ERK in lapatinib-resistant cancer cell line.

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