Apatinib preferentially inhibits PC9 gefitinib-resistant cancer cells by inducing cell cycle arrest and inhibiting VEGFR signaling pathway

Yong An Song, Ting Ma, Xue Yan Zhang, Xiang Song Cheng, Ayobami Matthew Olajuyin, Zhifu D Sun, Xiao Ju Zhang

Research output: Contribution to journalArticle

Abstract

Background: Lung cancer is one of the most common and deadly tumors around the world. Targeted therapy for patients with certain mutations, especially by use of tyrosine kinase inhibitors (TKIs) targeting epidermal growth factor receptor (EGFR), has provided significant benefit to patients. However, gradually developed resistance to the therapy becomes a major challenge in clinical practice and an alternative to treat such patients is needed. Herein, we report that apatinib, a novel anti-angiogenic drug, effectively inhibits obtained gefitinib-resistant cancer cells but has no much effect on their parental sensitive cells. Methods: Gefitinib-resistant lung cancer cell line (PC9GR) was established from its parental sensitive line (PC9) with a traditional EGFR mutation after long time exposure to gefitinib. Different concentrations of apatinib were used to treat PC9, PC9GR, and other two lung cancer cell lines for its anti-growth effects. RNA sequencing was performed on PC9, PC9GR, and both after apatinib treatment to detect differentially expressed genes and involved pathways. Protein expression of key cycle regulators p57, p27, CDK2, cyclin E2, and pRb was detected using Western blot. Xenograft mouse model was used to assess the anti-tumor activity of apatinib in vivo. Results: The established PC9GR cells had over 250-fold increased resistance to gefitinib than its sensitive parental PC9 cells (IC 50 5.311 ± 0.455 μM vs. 0.020 ± 0.003 μM). The PC9GR resistance cells obtained the well-known T790M mutation. Apatinib demonstrated much stronger (~ fivefold) growth inhibition on PC9GR cells than on PC9 and other two lung cancer cell lines, A549 and H460. This inhibition was mostly achieved through cell cycle arrest of PC9GR cells in G1 phase. RNA-seq revealed multiple changed pathways in PC9GR cells compared to the PC9 cells and after apatinib treatment the most changed pathways were cell cycle and DNA replication where most of gene activities were repressed. Consistently, protein expression of p57, CDK2, cyclin E2, and pRb was significantly impacted by apatinib in PC9GR cells. Oral intake of apatinib in mouse model significantly inhibited establishment and growth of PC9GR implanted tumors compared to PC9 established tumors. VEGFR2 phosphorylation in PC9GR tumors after apatinib treatment was significantly reduced along with micro-vessel formation. Conclusions: Apatinib demonstrated strong anti-proliferation and anti-growth effects on gefitinib resistant lung cancer cells but not its parental sensitive cells. The anti-tumor effect was mostly due to apatinib induced cell cycle arrest and VEGFR signaling pathway inhibition. These data suggested that apatinib may provide a benefit to patients with acquired resistance to EGFR-TKI treatment.

Original languageEnglish (US)
Article number117
JournalCancer Cell International
Volume19
Issue number1
DOIs
StatePublished - May 2 2019

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Cell Cycle Checkpoints
Neoplasms
Lung Neoplasms
Epidermal Growth Factor Receptor
Cyclins
Growth
Protein-Tyrosine Kinases
Mutation
apatinib
gefitinib
Therapeutics
RNA Sequence Analysis
Cell Line
Angiogenesis Inhibitors
G1 Phase
DNA Replication
Heterografts
Genes
Cell Cycle
Proteins

Keywords

  • Apatinib
  • Cell cycle arrest
  • Gefitinib-resistance
  • PC9 cells
  • VEGF2 pathway

ASJC Scopus subject areas

  • Oncology
  • Genetics
  • Cancer Research

Cite this

Apatinib preferentially inhibits PC9 gefitinib-resistant cancer cells by inducing cell cycle arrest and inhibiting VEGFR signaling pathway. / Song, Yong An; Ma, Ting; Zhang, Xue Yan; Cheng, Xiang Song; Olajuyin, Ayobami Matthew; Sun, Zhifu D; Zhang, Xiao Ju.

In: Cancer Cell International, Vol. 19, No. 1, 117, 02.05.2019.

Research output: Contribution to journalArticle

Song, Yong An ; Ma, Ting ; Zhang, Xue Yan ; Cheng, Xiang Song ; Olajuyin, Ayobami Matthew ; Sun, Zhifu D ; Zhang, Xiao Ju. / Apatinib preferentially inhibits PC9 gefitinib-resistant cancer cells by inducing cell cycle arrest and inhibiting VEGFR signaling pathway. In: Cancer Cell International. 2019 ; Vol. 19, No. 1.
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AU - Ma, Ting

AU - Zhang, Xue Yan

AU - Cheng, Xiang Song

AU - Olajuyin, Ayobami Matthew

AU - Sun, Zhifu D

AU - Zhang, Xiao Ju

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AB - Background: Lung cancer is one of the most common and deadly tumors around the world. Targeted therapy for patients with certain mutations, especially by use of tyrosine kinase inhibitors (TKIs) targeting epidermal growth factor receptor (EGFR), has provided significant benefit to patients. However, gradually developed resistance to the therapy becomes a major challenge in clinical practice and an alternative to treat such patients is needed. Herein, we report that apatinib, a novel anti-angiogenic drug, effectively inhibits obtained gefitinib-resistant cancer cells but has no much effect on their parental sensitive cells. Methods: Gefitinib-resistant lung cancer cell line (PC9GR) was established from its parental sensitive line (PC9) with a traditional EGFR mutation after long time exposure to gefitinib. Different concentrations of apatinib were used to treat PC9, PC9GR, and other two lung cancer cell lines for its anti-growth effects. RNA sequencing was performed on PC9, PC9GR, and both after apatinib treatment to detect differentially expressed genes and involved pathways. Protein expression of key cycle regulators p57, p27, CDK2, cyclin E2, and pRb was detected using Western blot. Xenograft mouse model was used to assess the anti-tumor activity of apatinib in vivo. Results: The established PC9GR cells had over 250-fold increased resistance to gefitinib than its sensitive parental PC9 cells (IC 50 5.311 ± 0.455 μM vs. 0.020 ± 0.003 μM). The PC9GR resistance cells obtained the well-known T790M mutation. Apatinib demonstrated much stronger (~ fivefold) growth inhibition on PC9GR cells than on PC9 and other two lung cancer cell lines, A549 and H460. This inhibition was mostly achieved through cell cycle arrest of PC9GR cells in G1 phase. RNA-seq revealed multiple changed pathways in PC9GR cells compared to the PC9 cells and after apatinib treatment the most changed pathways were cell cycle and DNA replication where most of gene activities were repressed. Consistently, protein expression of p57, CDK2, cyclin E2, and pRb was significantly impacted by apatinib in PC9GR cells. Oral intake of apatinib in mouse model significantly inhibited establishment and growth of PC9GR implanted tumors compared to PC9 established tumors. VEGFR2 phosphorylation in PC9GR tumors after apatinib treatment was significantly reduced along with micro-vessel formation. Conclusions: Apatinib demonstrated strong anti-proliferation and anti-growth effects on gefitinib resistant lung cancer cells but not its parental sensitive cells. The anti-tumor effect was mostly due to apatinib induced cell cycle arrest and VEGFR signaling pathway inhibition. These data suggested that apatinib may provide a benefit to patients with acquired resistance to EGFR-TKI treatment.

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KW - Cell cycle arrest

KW - Gefitinib-resistance

KW - PC9 cells

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