| c-Met is the cell surface receptor for hepatocyte growth factor(HGF). HGF-induced activation of c-Met results in a complex genetic program referred to as "invasive growth."It consists of a series of physiological processes including proliferation, invasion, and angiogenesis. It usually occurs inembryonic development, postnatal hepatic, repair of cardiac injury repair and pathologically during oncogenesis.Dysregulation of HGF/c-Met signal axis has been observed in a wide range of human malignancies, including bladder, breast, cervical, colorectal, gastric, head and neck, liver, lung, ovarian, pancreatic, prostate, renal, and thyroid cancer, as well as in various sarcomas, hematopoietic malignancies,and melanoma. In lung cancer, overexpression of c-Met was observed in 40%-60% of patients, and 4% lung cancer patients were found Met gene amplifiation. Furthermore, Met gene amplifiation also was found to be an additional mechanism of acquired EGFR-TKI resistance. Amplifid Met results in overexpression and overactivation of c-Met and consequently triggers the activation of Her3 which activates downstream signal transducer molecules, such as Akt and Erk, independent of EGFR kinase activity. In clinic, Bean reported that Met gene amplifiation was detected in 22% of acquired EGFR-TKI resistant non-small cell lung cancers, and, compared with patients unexposed to EGFR kinase inhibitor, Gefiinib or Tarceva treatment was more likely to select Met gene amplifiation (21% versus 3%). Th observation provides c-Met as a target in lung cancer therapy. In fact, several Met-targeted molecules are under early clinical evaluation currently. Based on a phase II result of MetMab, a c-Met specifi antibody, patients with c-Met overexpression would benefi from Met-targeted therapy, suggesting that detection of c-Met status is critical for Mettargeted therapy. However, not all patients in clinic are suitable for biopsy; thus it is necessary to discover a surrogate marker to detect c-Met status.Soluble Met (s-Met) is generated via c-Met ectodomain shedding. c-Met is initially synthesized as a single-chain intracellular precursor and subsequently undergoes proteolytic processing at diffrent stages during intracellular traffiing, leading to the presentation of an a/β heterodimer at cell surface. Th 140 P KD chain of the complex can be proteolytically cleaved by cells constitutively and released to the surrounding environments. s-Met was found to exist in several cancer cells culture supernatants, and a signifiant and direct correlation has been established in preclinical cell line and mouse models between the malignant potential and rate of c-Met ectodomain shedding.Therefore, this study were investigated s-Met expression in lung cancer; and in vitro epidermal growth factor receptor tyrosine kinase inhibitor-resistant non-small cell lung cancer cells by siRNA c-Met and combined with anti-tumor drugs (erlotinib) jointly applied research to c-Met as a target of drug therapy combined with chemotherapy to get the role of EGFR-TKI-resistant non-small cell lung cancer.1 Soluble c-Met is a reliable and sensitive marker to detect c-Met expression level in lung cancerObjective:to study the expression of s-Met in non small cell lung cancer and EGFR in the treatment of non small cell lung cancer.Methods:Select 146 cases of histologically diagnosed lung cancer patients in our hospital from January 2013 to December 2013 stay thoracic surgical treatment of clinical data and tissue samples and blood samples were also collected blood samples of healthy volunteers and patients before surgery and postoperative blood samples. ELISA assay was used to detect soluble c-Met. Immunohistochemical detection was used to examined c-Met in lung cancer, and analyze the correlation between soluble c-Met and c-Met in lung cancer.Results:①In healthy volunteers and in patients with lung cancer in both soluble c-Met expression in healthy volunteers, the serum concentration of soluble c-Met larger range, and in the serum of patients with lung cancer are also widely distributed in the concentration range, far higher than the healthy Volunteers serum soluble c-Met expression. ② serum soluble c-Met lung cancer and lung cancer tissue concentration c-Met expression showed a good correlation. Overexpressed in lung cancer c-Met (IHC 2+/3+) in serum concentrations of soluble c-Met was significantly increased. However, n lung cancer patients with low expression of c-Met-positive (IHC 1+) in the serum concentration of soluble c-Met is not significantly increased. ③ soluble c-Met expression with different clinical types of lung cancer and express different. In well-differentiated lung cancer, the expression of soluble c-Met in a significant increase in the degree of differentiation and with reduced expression of soluble c-Met is further increased. In TNM staging, TNM stage level with increased expression of soluble c-Met is further increased; increased with the degree of malignancy, the serum of patients with lung cancer in high-level expression of soluble c-Met reduce increased. Soluble c-Met expression is reduced and the degree of negative correlation between the differentiation of lung cancer, and lung cancer staging were positively correlated. ④ When the expression of c-Met overexpression surgery for lung cancer patients after excision of serum to detect the expression of soluble c-Met, result in significantly reduced serum soluble expression of c-Met. C-Met expression in surgical resection of lung cancer patients with negative serum to detect the expression of soluble c-Met, the serum levels of soluble c-Met expression did not change significantly. C-Met expression in weakly positive lung cancer patients after surgical resection to detect the expression of serum soluble c-Met, the results of serum soluble c-Met expression levels are slightly lower, was not statistically significant. ⑤ performance of soluble c-Met (s-Met) determination of diagnosis of lung cancer, the results confirm that the threshold sensitivity (true positives/(true positives+false negatives)) amounted to 90.5%, while the critical value of specificity (true negative/(true negatives+false positives)) amounted to 90.3%.⑥Whether it is the case with negative expression of c-Met in lung cancer tumor tissue, or tumor tissues of patients with lung cancer overexpression of c-Met, and tumor size were unrelated.Summary:Concentration of soluble c-Met and lung tissue expression evels of c-Met has a good correlation. And expression of soluble c-Met is reduced and the degree of negative correlation between the differentiation of lung cancer, and lung cancer staging were positively correlated. Tip increase in lung cancer soluble c-Met is overexpressed c-Met in lung tissue secreted into the blood circulation. Therefore, the concentration of soluble c-Met detection can be used as indicators of lung tissue expression of c-Met judgment.2 Exploration of the expression of c-Met mRNA and protein, soluble c-Met protein in NSCLC with EGFR-TKI-resistant cell linesObjective:To explore the expression of c-Met mRNA and protein, soluble c-Met protein expression in NSCLC with EGFR-TKI-resistant cell lines.Methods:Establish and in vitro culture erlotinib-resistant non-small cell lung cancer cell lines A549, siRNA c-Met-resistant cell lines transfected into by Real time PCR c-Met mRNA expression was detected by Western blot analysis c-Met protein expression, detection of expression of soluble c-Met ELISA method. To explore the correlation of soluble c-Met protein and c-Met expression in A549 cell supernatants.Results: ①Resistant A549 cell lines significantly increased c-Met mRNA expression, A549 cell line with the normal c-Met mRNA expression, compared with a statistically significant (P<0.05). When c-Met siRNA interference and transfected into A549 cell lines resistant, can significantly reduce the expression of c-Met mRNA in the EGFR-TKI-resistant cell lines A549, A549 cell lines compared with the resistance, with significant statistical significance (P<0.05), but compared with the normal mRNA expression in A549 cells c-Met, was not statistically significant (P>0.05). ②Resistant A549 cell lines significantly increased c-Met protein expression EGFR-TKI, A549 cell line with the normal c-Met protein expression, compared with a statistically significant (P<0.05). When c-Met siRNA interference and ransfected into A549 cell lines resistant, can significantly reduce the expression of c-Met protein in the EGFR-TKI-resistant cell lines A549, A549 cell lines compared with the resistance, with significant statistical significance (P<0.05), but compared with the normal c-Met protein expression in A549 cells c-Met, was not statistically significant (P>0.05).③Soluble c-Met protein expression compared with normal A549 cell supernatants c-Met, with a statistically significant (P<0.05). When c-Met siRNA interference and transfected into A549 cell lines resistant, can significantly reduce the expression of soluble c-Met protein in the EGFR-TKI-resistant A549 cell supernatant compared with drug-resistant cell line A549, with a statistically significant significance (P<0.05), while the A549 cell line with normal soluble c-Met expression supernatant comparison was not statistically significant (P> 0.05).Summary:Resistant A549 cell lines significantly increased expression of c-Met mRNA and protein, Soluble c-Met. The expression of s-Met and c-Met protein have good correlations in A549 cell supernatants.3 Effect of c-Met target therapy on sensitivity of NSCLC with EGFR-TKIs resistanceObjective:To exploration the effect of c-Met target therapy on sensitivity of NSCLC with EGFR-TKIs resistance.Methods:Establish erlotinib-resistant non-small cell lung cancer cell lines A549 in vitro culture, A549 cells were cultured resistant and divided into four groups:crotrol group, erlotinib group, siRNA c-Met group, the combined effects of the group(the role of erlotinib in siRNA c-Met-resistant transfected into A549 cells). Detect each group of cell proliferation, colony formation and invasion case differences. MTT cell proliferation was detected in each group situation; colony-forming cells to detect the formation of cell clones; Transwell chambers to detect the cell invasion ability to change the situation.Results: ①In a separate action to establish Jiarueluo erlotinib erlotinib-resistant A549 cells, can not affect tumor cell proliferation, imatinib-resistant A549 cell proliferation compared with erlotinib, was not statistically significant (P>0.05). When the use of siRNA c-Met, can inhibit tumor cell proliferation, gefitinib and erlotinib-resistant A549 cell proliferation compared with erlotinib role of tumor cells, with statistically significant (P <0.05). Further, the effect of erlotinib on siRNA c-Met resistant transfected into A549 cells, tumor cell proliferation is seen more significantly inhibited, and erlotinib-resistant A549 cells and the role of tumor erlotinib cell proliferation, compared with a statistically significant (P<0.05), but the role and siRNA c-Met group, the inhibitory effect more pronounced.②Similar results can be seen in the clone formation assay.③In a separate action to establish Jiarueluo erlotinib erlotinib-resistant A549 cells, can not affect the ability of tumor cell invasion. siRNA c-Met can inhibit the ability of tumor cell invasion. Further, the effect of erlotinib on siRNA c-Met-resistant transfected into A549 cells, can be seen to be more invasive ability of tumor cells significantly inhibited, and the role and siRNA c-Met group, inhibition and more apparent.Summary:c-Met as a therapeutic target may inhibit EGFR-TKI-resistant lung cancer cell proliferation, colony formation and invasion ability to inhibit c-Met combined with EGFR-TKI therapy may further inhibit EGFR-TKI-resistant lung cancer cell proliferation, colony formation and invasive ability, explain to c-Met as a therapeutic target to improve the resistance of tumor cells, tumor cells to enhance the sensitivity of EGFR-TKI.Conclusions:This topic through tumor tissue from patients with lung cancer and blood samples confirmed the expression of soluble c-Met may be used as an indicator to detect the concentration of c-Met expression in lung cancer tissues. Experimental study on the cellular level from confirmed to c-Met as a therapeutic target may inhibit EGFR-TKI-resistant lung cancer cell proliferation, colony formation and invasion ability to inhibit c-Met joint EGFR-TKI therapy may further inhibit EGFR-TKI resistance cancer cell proliferation, colony formation and invasion abilities, instructions for c-Met is a therapeutic target tumor cells could improve drug resistance of tumor cells to enhance the sensitivity of EGFR-TKI. The expression of s-Met and c-Met... |
