Study Of EML4-ALK Fusion Gene In The Primary And Metastases Tumor Tissues And The Serum Of VEGF In Non-small Cell Lung Cancer Patients

Objective1 Using quantitative real-time polymerase chain reaction(qRT-PCR) technology to detect the express status of EML4-ALK fusion gene in 119 cases of primary tumors and 71 cases of metastases in non-small cell lung cancer including 15 case of patients with primary tumors and matching metastases tumors.2 Analyzed the relationship between the expression of EML4-ALK fusion gene in NSCLC primary tumor and metastases tumors tissues and the clinical pathological characteristics as well as the serum levels of vascular endothelial growth factor(VEGF) of NSCLC patients with baseline data.Methods1 Collected the fresh or paraffin tissue specimens from 119 cases of primary tumors and 71 metastases tumors of NSCLC patients, who were diagnosed in Beijing military region general hospital and the 304 hospital affiliated to the Chinese people’s liberation army general hospital from February 2013 to November 2014. Both 15 cases of primary lung tissue specimens and matched metastases tissue samples were included. Collected all specimens of the serum of NSCLC patient(175 cases expect 15 matched specimens).2 Analyzed the relationship between the expression status of EML4-ALK fusion gene and the clinical pathologic features of 119 cases of primary tumors tissues and 71 cases of metastases tissues of NSCLC patients by using the qRT-PCR technique to detect the EML4-ALK fusion gene.3 By using enzyme-linked immunosorbent (ELISA) to detect the serum levels of VEGF, analyzed the relationship between it and the EML4-ALK fusion gene of NSCLC patients.Results1 EML4-ALK fusion gene positive status in the primary and metastases tissues NSCLC patientsIn 175 tumor tissue samples of NSCLC patients,13 cases with EML4-ALK fusion gene is positive, positive rate was 7.4%(13/175). There were 10 of 119 primary tumors samples that had EML4-ALK fusion gene and the positive rate was 8.4%(10/119). There were 5 cases of 71 metastatic tumor samples that had EML4-ALK fusion gene and the positive rate was 7.1%(5/71). There were 40 cases of lymph nodes samples,10 cases of brain tissue and bone tissue samples,8 cases of body surface soft tissue,1 case of live tissue samples, 1case of colon tissue and pleural tissue sample, of which had positive-EML4-ALK fusion gene tissues including lymph node, bone, brain and pleura whose positive rate were 2.5%(1/40),20%(2/10),10%(1/10) and 100%(1/1) respectively.2 The connection between EML4-ALK fusion gene status in the primary tumors and metastases tumors of NSCLC patients and their clinical pathological features(1) The positive rate of EML4-ALK fusion gene of 119 cases of primary tumor samples of NSCLC patients in smoking group and non-smoking group were 1.8% and 14.3% respectively(P=0.034), in adenocarcinoma group and non-adenocarcinoma group were 14.1% and 1.9% respectively (P=0.044), both the difference between two group had statistical significance. The positive rate of EML4-ALK fusion gene in male group and female group were 5.1% and 11.7% respectively (P=0.335), in the age>60 years old group and age≤60 years old group were 6.6% and 10.3% respectively (P=0.679), in low differentiation group and middle-high differentiation group were 4.0% and 11.6% respectively (P=0.225), in the stage of Ⅰ-Ⅱ and Ⅲ-Ⅳ were 7.5% and 8.9%(P=0.547). However, statistical analysis showed there were no relationship in sex group, age group and differentiation group about EML4-ALK fusion gene positive rate and gender, pathologic types(P>0.05).(2) The positive rate of EML4-ALK fusion gene of 71 cases of metastases tumor samples of NSCLC patients in smoking group and non-smoking group were 3.4% and 9.5% respectively, in adenocarcinoma group and non-adenocarcinoma group were 7.4% and 5.9% respectively, in male group and female group were 7.5% and 6.5% respectively, in the age>60 years old group and age≤60 years old group were 6.7% and 7.3% respectively, in low differentiation group and middle-high differentiation group were 6.3% and 7.7% respectively, in stage of Ⅲ and Ⅳ were 13.3% and 5.4%. All the difference were not statistically significant(P>0.05).3 The relationship between the serum levels of VEGF and the EML4-ALK fusion gene in primary tumors and metastases tumors of NSCLC patients The serum levels of VEGF of 175 cases of NSCLC patients except 15 pairs samples was significantly higher normal. The levels of serum VEGF in Ⅰ-Ⅱ stage and Ⅲ-Ⅳ stage of NSCLC patients was 177.42±81.88pg/ml and 401.74±215.06 pg/ml respectively, the difference were statistically significant (P<0.05). The serum levels VEGF of 10 cases of EML4-ALK fusion gene-positive and 109 cases of EML4-ALK fusion gene-negative were positive were 346.92±211.25pg/ml and 393.59± 222.24pg/ml in 119 cases of the primary tumor tissues of NSCLC patients. The serum levels of VEGF of 5 cases of EML4-ALK fusion gene-positive and 66 cases of EML4-ALK fusion gene-negative were positive were 403.31±128.73pg/ml and 363.88±208.30pg/ml in 71 cases of the primary tumor tissues of NSCLC patients, statistical analysis showed that EML4-ALK fusion gene had nothing to do with the serum levels of VEGF (P> 0.05).4 The EML4-ALK fuion gene expression status in the primary tumor tissues and their matched metastatic tumor tissues of NSCLC patientsThere were 15 cases of primary lung cancer tissues matching 9 cases of lymph node tissues,2 cases of brain tissues,2 cases of bone tissues,1 case of pleural tissue and 1 case of skin soft tissue in 119 cases of primary tumor tissues and 71 cases of metastatic tumor tissues of NSCLC patients. Of 15 pairs of specimens, we detected the 2 cases of primary lung tissue matching pleural tissue and bone tissue respectively. Their EML4-ALK fusion genes were all positive and consistent positive rate were 100%(2/2), the remaining 13 cases matching organization whose EML4-ALK fusion genes negative rate were 100%(13/13). The two patients whose EML4-ALK fusion gene were positive were women with 57 and 71 years old respectively without smoking history. Their tissues pathology were adenocarcinoma or containing adenocarcinoma.Conclusion1 The qRT-PCR technology can accurately detect the status of EML4-ALK fusion gene of NSCLC patients. The overall positive rate of EML4-ALK fusion gene of NSCLC patients was 7.4%. The positive rate in the primary tumor tissues and the metastatic tumor tissues were 8.4% and 7.1% respectively and the latter is slightly lower than the primary. Matching specimens shows there is no difference between EML4-ALK fuion gene in primary tumor tissues and it in metastatic tumor tissues. If the primary tumor tissue is difficult to obtain, we can consider the metastases tissue in order to provide treatment basis for those EML4-ALK fuion gene-positive NSCLC patients who use ALK inhibitors.2 EML4-ALK fusion gene expression status of the primary tumor tissues are related to patients’s pathologic type and smoking history, mainly in adenocarcinoma and non-smokers, but not related to gender, age and the degree of differentiation. The results suggest the clinical pathological characteristics of EML4-ALK fusion gene-positive NSCLCL patients in China.3 The serum levels of VEGF of NSCLC patients obviously is higher than reference value and is associated with the stage of NSCLC. The later stage, the higher the serum levels of VEGF. At the same time, the results show that the positive or negative status of EML4-ALK fusion gene in tumor tissues has no significant correlation with serum levels of VEGF. It hits indirectly that their signal pathways may have less association in the occurrence and development process of NSCLC.