The Role of Integrin β1 Up-regulated in Resistance of Non-small Cell Lung Cancer to EGFR TKI
|Course||Department of Respiratory Medicine|
|Keywords||Resistance Gefitinib Β1 integrin Non - small cell lung cancer|
Background and Purpose: In many countries, lung cancer is due to cancer, the leading cause of death. Due to the lack of early stage lung cancer symptoms, about 70% -80% of lung cancer patients are diagnosed at an advanced stage. The most advanced lung cancer patients were treated with platinum-based chemotherapy, but is limited to first-line chemotherapy efficacy. And effective second-line treatment of non-small cell lung cancer is not ideal, PFS only three months. With NSCLC the molecular pathogenesis gradually clarify, for these molecular pathogenesis of molecular targeted therapy has become possible. NSCLC molecular target for treatment is mostly concentrated in the tumor cells, the epidermal growth receptor (epidermal growth factor receptor, EGFR). Inhibitors (EGFR TKI) is a receptor tyrosine kinase inhibitor, which compete with ATP epidermal growth factor receptor-specific binding sites, thus inhibiting the activity of tyrosinase, EGFR activation has been used in the clinical treatment of patients with non-small cell lung cancer mutations. However, many patients in the medication for some time after, that the emergence of resistance phenomenon even sicker. Study the mechanisms of resistance to this drug has become a hot research in recent years. Resistance mechanisms mature EGFR T790M mutation and amplification of the proto-oncogene c-met. Recent studies show that the disintegrin adjustable epithelial cell adhesion to the extracellular matrix (extracellular matrix ECM) proteins, induction of tyrosine phosphorylation, partial activation of EGFR, EGF can also be to enhance the expression of the integrin α2β1. PC9 cell line, and on this basis gefitinib resistant cell line induced when analyzed, found that the majority of drug-resistant strains of collagen adhesion capacity is much higher than PC9, suggesting that cell adhesion molecules and extracellular matrix may be involved in non gefitinib resistance [. Another study found that the analysis PC9 and resistant cell lines found resistant cell lines β1 integrin levels significantly increased β1 integrin may be associated with acquired resistance to gefitinib group of this study is aimed at analyzing the PC9 cell lines on the basis of establish stable expression the β1 integrin subcellular strains PC9/D6 and the empty vector transfected cell lines PC9/PCD, further confirmed further from the level of in vivo or in vitro expression of integrin β1 upward impact gefitinib sensitivity Immunohistochemical detection the level of protein expression in the transplanted tumor tissue, to better understand the The integrin β1 impact of EGFR TKI sensitivity, provide a strong basis for the search for new mechanisms of resistance. The first part of upregulation of integrin β1 gefitinib inhibit cell proliferation purposes: evaluation of three NSCLC cell lines PC9 (EGFR mutation), and based on liposomes stably transfected cell lines β1 integrin PC9/D6 stably transfected pcDNA 3.1 and liposome the Blank control strains PC9/PCD gefitinib efficacy of Nepal, to explore the expression of integrin β1 raised with gefitinib gefitinib resistant. Methods: PC9, PC9/PCD and PC9/D6 three cells, 0.5μmol / L gefitinib after alternate Niger role, was detected by MTT cell proliferation of the three cells at different time points. Results: 1. Gefitinib lung adenocarcinoma PC9 cell lines significantly inhibited, and this inhibition in a time-dependent (P lt; 0.05) 2. Gefitinib on the control strains PC9/PCD significantly inhibited, and this inhibition showed a time-dependent manner (P lt; 0.05). And the PC9 cell lines and PC9/PCD of cell lines at the same point in time there is no statistically significant difference in the rate of cell proliferation. 3 gefitinib high expression the integrin β1 cell strains PC9/D6 of the inhibition was significantly weakened. Compare with the PC9 cell lines and PC9/PCD of cell lines found to have a statistically significant difference (P lt; 0.01) compared to the first two cells at different time points PC9/D6 cell proliferation rate Description β1 integrin expression raised to reduce the sensitivity of the cell lines to gefitinib. , But gefitinib still part to suppress PC9/D6 cell proliferation, visible integrin β1 may be involved in the resistance of gefitinib part. The second part of β1 integrin upregulation and gefitinib inhibit tumor growth Objective: To investigate the integrin β1 upregulation gefitinib inhibit tumor growth. Method: the establishment of the PC9, PC9/PCD, PC9/D6 three cells in nude mice xenograft model, while the treatment group and the control group (3mg/kg gefitinib Benny and 1% Tween-80), and observed tumor growth depicting tumor growth curves, to the end of treatment, peel the tumor tissue, known as tumor inhibitory rate was calculated at the same time. Results: 1. Gefitinib PC9 cell line xenografts in nude mice was inhibited. To the end of treatment, the tumor weight of the treatment group and the control group (71.7 ± 8.12) mg (1364.86 ± 259.173) mg, the inhibition rate was 95.16%. 2 nude mouse xenograft model antitumor effect of gefitinib the PC9/PCD cell lines significantly. To the end of the 28 days of treatment, the inhibition rate was 93.54% 3. Gefitinib PC9 cell lines in nude mice xenograft model antitumor effect is relatively weak, and even in the given gefitinib treatment, while the treatment group nude mice is still in a growth state, only relative to the control group, the growth rate slow down. With the PC9 and PC9/PCD treatment group compared with a significant difference (P lt; 0.05). The third part of the transplanted tumor morphological examination of their organizations integrin β1 expression of EGFR, pEGFR Objective: To observe the treatment and control groups transplanted tumor morphological changes detected tissue integrin β1, EGFR, pEGFR expression. Method: peel the tumor tissue by 4% paraformaldehyde, pathology, at the same time take part slice hematoxylin - eosin staining; I slice immunohistochemical assay integrin β1, EGFR, pEGFR expression. Results: 1.PC9, PC9/PCD treatment group compared with the control group, the tumor cells are arranged more loose, the nucleus was lightly stained. But PC9/D6 group treatment group compared with the control group, the tumor cells were still dense, nucleus showed no lightly stained. Integrin β1 expression intensity in PC9/D6 group was significantly higher than that the PC9 and its the blank control strains PC9/PCD (P lt; 0.01), three cell transplant tumor treatment group compared with the control group, the β1 integrin There was no statistically significant difference. Three cells in the transplanted tumor EGFR expression was no significant difference has nothing to do with whether to give gefitinib treatment. Three cells transplanted tumors receiving gefitinib therapy pEGFR have different degrees of decline, the control group and the treatment group were significantly different (P lt; 0.01). PEGFR the treatment group PC9/D6 expression intensity was significantly higher than PC9, PC9/PCD (P lt; 0.01). CONCLUSION: Transfection of β1 integrin the cell PC9/D6 cell proliferation rate was significantly higher than PC9/PCD and PC9 group. Gefitinib PC9/D6 nude mice transplanted tumor growth inhibition is the most obvious, the inhibition rate was 61.38%, with PC9/PCD, PC9 group with statistically significant differences PC9/PCD PC9 inhibition rate were 93.54% 95.16%. Β1 integrin expression the PC9/D6 nude mice than PC9/PCD, PC9. The EGFR in express no statistical difference between the three. Three groups of nude mice by treatment with gefitinib, pEGFR expression have different degrees of decline, but the pEGFR of PC9/D6 treatment group expression group was significantly to higher than PC9/PCD PC9. In vivo or in vitro, β1 integrin upregulation reduce cell lines or xenograft tumor sensitivity to gefitinib. Prompted β1 integrin but a new resistance mechanisms involved in EGFR TKI resistance.