ABSTRACT
Radiation therapy is a widely used treatment for esophageal cancer. However, radiation resistance might result in a poor prognosis. Overexpression of HER2 has been related to adaptive radiation resistance. Pyrotinib is a HER2 inhibitor that shows an anti-tumor effect in breast cancer. The present study aims to explore the influence of pyrotinib combined with radiotherapy on HER2-positive esophageal cancer cells and explore the underlying mechanism. We screened two cell lines (TE-1 and KYSE30) that highly express HER2 from several human esophageal cancer cell lines. Cells were treated with pyrotinib or/and radiation. Cell proliferation, cell cycle distribution, and cell migration were measured. The protein levels involved in cell cycle and DNA repair were measured by Western blot. Results showed that pyrotinib inhibited HER2 activation and exerted an anti-proliferative effect in TE-1 and KYSE30 cells. Furthermore, it enhanced the anti-proliferative effect of radiation in these two cell lines. These effects might be via inhibiting HER2 phosphorylation, inducing G0/G1 arrest, and reducing EMT and DNA repair. Our results indicated that pyrotinib sensitivitied HER2 positive esophageal cancer cells to radiation treatment through various mechanisms. These findings may provide a new therapeutic strategy for treating HER2 positive esophageal cancer.
Subject(s)
Acrylamides/pharmacology , Aminoquinolines/pharmacology , Esophageal Neoplasms/radiotherapy , Radiation Tolerance/drug effects , Radiation-Sensitizing Agents/pharmacology , Receptor, ErbB-2/antagonists & inhibitors , Cell Cycle Checkpoints/drug effects , Cell Cycle Checkpoints/radiation effects , Cell Line, Tumor , Cell Proliferation/drug effects , Cell Proliferation/radiation effects , Cyclin D1/metabolism , Cyclin-Dependent Kinase 4/metabolism , DNA Repair/drug effects , DNA Repair/radiation effects , Epithelial-Mesenchymal Transition/drug effects , Epithelial-Mesenchymal Transition/radiation effects , Esophageal Neoplasms/metabolism , Esophageal Neoplasms/pathology , Humans , Phosphorylation , Receptor, ErbB-2/metabolism , Signal TransductionABSTRACT
Background: Esophageal squamous cell carcinoma (ESCC) is a common cancer with poor prognosis. The molecular pathogenesis underlying ESCC remains to be explored. Leucine-rich É-2-glycoprotein 1 (LRG1) has been implicated in the pathogenesis of various cancer types, however its role in ESCC is unknown. Materials and Methods: Data from the public database was analyzed to address the expression of LRG1 in ESCC. Gain-of-function studies were performed in select ESCC cell lines by over-expression or addition of recombinant LRG1, while loss-of-function studies achieved by small interfering RNA mediated knockdown. Wound healing and transwell assays were conducted to investigate ESCC cell migration and invasion upon manipulating LRG1 levels. Western blot and Immunofluorescence staining were used to examine the changes in epithelial to mesenchymal transition (EMT) and TGFß signaling pathway. Results: LRG1 mRNA levels were found to be significantly down-regulated in patients with ESCC as well as in several ESCC cell lines. Silencing of LRG1 promoted, while overexpression of LRG1 inhibited ESCC cell migration and invasion. In line with this, Silencing of LRG1 enhanced, while overexpression of LRG1 reduced TGFß signaling and EMT of ESCC cells. Conclusion/Significance: LRG1 suppresses ESCC cell migration and invasion via negative modulation of TGFß signaling and EMT. Down-regulation of LRG1 in ESCC patients may favor tumor metastasis and disease progression.
ABSTRACT
Aims: Pyrotinib is a newly developed irreversible pan-ErbB receptor tyrosine kinase inhibitor for treatment of human epidermal growth factor receptor 2 (HER2)-positive cancers, and clinic trials of pyrotinib in treatment of HER2-positive gastric cancer (GC) are underway. Exosomes are tiny vesicles secreted by cancer cells and take essential roles in the progression of carcinoma. Whether pyrotinib application has any effect on the cancer cell-released exosomes has not been studied. The aim of our work was to address if pyrotinib treatment impacts the effect of HER2-positive GC cell-derived exosomes on endothelial cell (EC) progression. Methods: Isolation of exosomes released by HER2-positive NCI-N87 and MKN45 lines after pyrotinib treatment was performed. Then, human umbilical vein endothelial cells (HUVECs) were incubated with different concentrations of exosomes to address their proliferation by 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium (MTS). Effect of pyrotinib-treated exosomes at concentration of 10 µg/mL was compared to that without pyrotinib treatment over 96-hr time course. Transwell assay and wound-healing assay were carried out by incubating with exosomes released by NCI-N87 and MKN45 cells with/without pyrotinib treatment over 24-hr time course. The aforementioned experiments were done under same conditions in order to evaluate the combined effect of apatinib and pyrotinib on HUVEC motility and invasive capacity. Results: We showed that HUVEC proliferation, motility and invasive capacity were further enhanced upon incubation with exosomes released by pyrotinib-treated GC cell lines, compared to those without pyrotinib treatment. Significantly, this effect was counteracted by the vascular endothelial growth factor receptor (VEGFR)-2 inhibitor apatinib which inhibits EC progression. Conclusion: Our study suggests that pyrotinib application on HER2-positive GC produces stronger exosomes that promote the proliferation and motility of vascular ECs, and combination of pyrotinib with apatinib provides potentially better therapy.
