Application Of The Method To Detect SCCA1 MRNA By Real-time Quantitative RT-PCR In Lung Squamous Cell Cancer

Objectives The incidence and mortality of lung cancer was increased year by year,which make serious burden for people.It's crucial to explore and establish method for specific markers of circulating tumor cell in lung cancer.SCCA belonging to ovserpin/clade B serpin family is highly expressed in squamous cells,also known as squamous cell carcinoma antigen,including SCCA1 and SCCA2.They are encoded by two separate but highly homologous genes.However,SCCA1 and SCCA2 show distinct properties and substrate specificities.Recently,it was demonstrated that the expression level of SCCA is associated with the development of squamous cell carcinoma.In patients with lung cancer,it demonstrated a higher level SCCA1 in lung cancer when compared to benign lung disease or healthy people.Circulating tumor cells are tumor cells that enter the cycle.Once circulating tumor cells are detected in the blood,it means tumor formation or metastasis.Due to the circulating tumor cells in the blood is rarely difficult to detect,expression level of the circulating tumor cell is consistent with the mRNA expression level.Therefore,the detection of nucleic acid in circulating tumor cells has broad application prospects.The objective of this study was to investigate a quantitative method for detecting squamous cell carcinoma antigen 1(SCCA 1)mRNA by real-time quantitative RT-PCR,and explore the relationship between the expression level of SCCA1 in peripheral blood of lung cancer patients and clinical characteristics.The application of this method to detect the circulating tumor cells in the peripheral blood of lung squamous cell cancer patients will also be evaluated.Methods Totally 28 cases lung cancer patients with peripheral blood samples,who were all in hospital between 2/1/2014 and 10/31/2014 and confirmed by pathological examination,were included in the study(11 cases were confirmed distal metastasis,17cases not diagnosed metastasis),10 cases pneumonia patients,10 healthy donors.RNA extraction,and the SCCA 1 mRNA was detected by the real-time quantitative RT-PCR in peripheral blood samples.Quantitative standard curve was calculated by regression analysis.Gene expression with different pathological stages in lung cancer was performed by K Independent Samples Test of multiple independent sample.The positive detection rate was performed by Kruskal-Wallis H test of multiple independent sample rates.ORIGIN 7.5 software was used to draw scatter plots and analyze.Results 1.The method to detect the SCCA 1 mRNA using the real-time quantitative RT-PCR was successfully established by defining the probes,primers,standards and controls.The sensitivity and specificity of this method was high.2.All of the peripheral blood samples from 11 patients with metastatic lung cancer were positive for SCCA 1mRNA,9 of 17 patients with non-metastatic lung cancer were positive for SCCA 1mRNA in the peripheral blood samples,all of the blood samples from 10 pneumonia patients and 10 healthy donors were negative for SCCA 1 mRNA,which had statistically significant difference(?~2=36.456,p<0.05).3.The expression of SCCA1mRNA was relevant to TNM stage,there was a tendency for them to increase with the stage increasing,which had statistically significant difference(?~2=36.456,p<0.05).Conclusion 1.The quantitative RT-PCR assay for SCCA 1 mRNA could be used for the detection of circulating tumor cells in the peripheral blood of lung cancer patients,which would be valuable for the diagnosis of lung squamous cell cancer.2.The expression of SCCA1 mRNA in peripheral blood of patients with squamous cell carcinoma is correlated with tumor metastasis and clinical stage,which is a better parameter of circulating tumor cells in lung squamous cell cancer.