| BackgroundLung cancer is the leading cause of cancer-related death worldwide which poses a major threat to human life and health due to the best time for treatment was usually missed when final diagnosed. Non-small cell lung cancer (NSCLC), which accounts for about80%to85%in all kinds of lung cancers has a dismal5-year survival rate less than15%. Therefore, it is important to screen reliable clinical bio-makers for predicting the prognosis of NSCLC patients and development of individualized treatment.MicroRNAs (miRNAs) as a focus in studies on oncomolecularbiology, are found to play key roles in tumorigenesis and tumor progression. Hundreds of miRNAs are determined to be present in serum or plasma which have significant disease specificity and display correlation with states of diseases. Thus, miRNAs have considerable applicable value in cancer diagnosis, treatment and prognosis monitoring.In our previous study, a novel monoclonal antibody NJ001specific to non-small cell lung cancer was successfully prepared by immunization with human lung adenocarcinoma cell line SPC-A1. Then, SPC-A1cells were treated with NJ001for different time for microarray expression analysis, and the results indicated that miR-638showed the most significant up-regulation in a time dependent manner during the apoptosis process. Here, this study was to explore the possibility of miR-638as a prognostic indicator to predict NSCLC prognosis with chemotherapy.PurposeIn this study, on the basis of altered expression of miR-638in human lung adenocarcinoma cell line SPC-A1during the apoptosis process affected by cisplatin in vitro, we further studied serum miR-638expression of clinical non-small cell lung cancer (NSCLC) patients during chemotherapy and analyzed the relationship between its expression and clinicopathological parameters as well as the prognosis for patients with different serum miR-638expression patterns.MethodsSPC-Al cell line was treated with different concentrations of cisplatin for different time in vitro, quantitative real-time PCR was performed to analyze miR-638expression in cultured supernatants and SPC-A1cells and flow cytometry was used to detect SPC-A1apoptosis rates. In clinic, paired serum samples before and after chemotherapy of89cases of NSCLC patients during the hospitalization from July2009to March2013in the First Affiliated Hospital of Nanjing Medical University were collected. Serum miR-638expression were determined by real-time PCR and correlation of the expression patterns after chemotherapy with NSCLC clinicopathological features(age, gender, smoking history, TNM stage and lymph node metastasis) were evaluated as well as the clinical value of miR-638for predicting NSCLC prognosis.ResultsCell apoptosis rates of SPC-A1with cisplatin treatment detected by flow cytometry showed that cisplatin induced significant increase of SPC-A1apoptosis rates in vitro in a dose-and time-dependent manner when the concentration was2.5μg/ml or5μg/ml. Real-time PCR results demonstrated that miR-638expression in cultured supernatants and cells elevated in a time dependent manner in the process of SPC-A1apoptosis with the prolonged time when the concentration of cisplatin was2.5μg/ml or5μg/ml. When the concentration was2.5μg/ml, miR-638expression in the culture supernatants and cells elevated after treatment for72h, and the fold change was3.90and4.49respectively than that of the paired untreated group (P <0.05); when the concentration was5μg/ml, miR-638expression elevated after treatment for24h and increased with the prolonged time, the increase of miR-638expression in culture supernatants for24h,48h and72h was1.65,1.81,2.74fold change respectively compared to the paired untreated group while the increase in cell was1.75,10.21,6.89fold change respectively which showed statistically significant difference (P<0.05). Correlation analysis indicated that miR-638expression levels in culture supernatants (DDP2.5μg/ml, r=0.808P<0.001; DDP5μg/ml, r=0.711P=0.001) or cell (DDP2.5μg/ml, r=0.823P<0.001; DDP5μg/ml, r=0.744P<0.001) showed positive correlation with apoptosis rates of SPC-A1respectively. Serum miR-638detection of89cases of NSCLC patients with the first-line chemotherapy (cisplatin) illustrated that78cases had changes of serum miR-638expression in which43cases showed serum miR-638up-regulation while35cases showed down-regulation. Serum miR-638expression pattern after chemotherapy was not correlated with the following clinicopathological features such as age, gender, smoking history or TNM stage. However, significant correlation of miR-638expression pattern with lymph node metastasis was observed which suggested that the positive rate of lymph node metastasis in patients with serum miR-638up-regulation was significantly lower than that in patients with serum miR-638down-regulation (74.4%vs94.3%,%2=5.483, P=0.019). Kaplan-Meier survival analysis according to serum miR-638expression patterns after chemotherapy showed that overall survival rate of patients with serum miR-638up-regulation was significantly higher than those with serum miR-638down-regulation (P=0.008). Cox proportional hazards model analysis showed that serum miR-638was an independent risk factor for prognosis of NSCLC patients.ConclusionCisplatin induced increase in SPC-A1cells apoptosis in vitro accompanied with miR-638up-regulation in cultured supernatants and cells. Serum miR-638expression pattern was correlated with lymph node metastasis in NSCLC and could predict the prognosis of NSCLC patients. Serum miR-638was an independent risk factor for NSCLC prognosis which could has a potential prognostic value in predicting NSCLC prognosis. |
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