| Lung cancer is one of the most common malignancies and has became one of a leading cause of cancer death in the United States. The main reasons for this is that the exact molecular mechanism responsible for lung cancer are still remain unclear. One of the most important reasons in cells that resulted in lung cancer is that the activation of oncogen and the inactivation of tumor suppressor genes(TSG). Recently, evidence has emerged that epigenetic changes play an important role in lung cancer's development and progression. Especially, the abnormal of DNA methylation modification that silenced tumor suppressor genes taking an essential role in the development of lung cancer. As a consequence, studying for new TSG that resulted in dysfunction in lung cancer's progression, and its role can elucidate the mechanism and instruct the therapeutic in lung cancer. The ADAM23 gene, a member of the ADAM protein family, is a transmembrane glycoproteins modulating a variety of cell-cell and cell-extracellular matrix interactions. In particular, it can activate the receptor of integrin avβ3 during metastatic progression, which can promote angiogenesis and accelerate tumor cell's growth and immigration. At present, it has been reported that ADAM23 as a TSG, its downregulation has closely correlated with malignant tumor, including breast caner, brain cancer and gastric cancers and so on. Its mechanism is associated with the abnormal of DNA promoter CpG island methylation modification. However, very little is known about the expression and its role as well as its relationship with promoter methylation in NSCLC.The aim of this study is to investigate the expression of ADAM23 in lung cancer and analyze the possible role of ADAM23 in NSCLC. Further study focus on its promoter methylation and the results could contribute to elucidate the mechanism of tumorigenesis. It could provide a new target for the treatment of lung cancer. And also provide a new prognosis factor for lung cancer. Objective:To investigate the expressions of ADAM23 and alphavbeta3(avβ3) in NSCLC and its correlation to clinicopathological characteristics. Then, we analyze the promoter methylation status of ADAM23 in non-small-cell lung cancer and its correlation with the expression of ADAM23, expecting to provide theoretical and experimental evidences for the role of ADAM23 in NSCLC.Methods:(1) The expressions of ADAM23 and avβ3 were detected by immunohistochemistry and RT-PCR in 52 paired NSCLC specimens and corresponding normal tissues as well as 8 benign pulmonary lesions and analyze its correlation with clinicopathological characteristics. Western blotting and RT-PCR assays were used to examine the expression of ADAM23 in lung squamous carcinomas cell (SK-MES-1) and lung adenocarcinomas cells(A549,SPC-A1,LTEP-a-2).(2) Methylation-specific PCR (MSP) was used to determine the ADAM23 gene promoter methylation status both in tissues and cell lines, then analyze the correlation with its expression.(3) NSCLC cells were treated with 5-aza-2'-deoxycytidine(5-Aza-2'-dC) to restore the ADAM23 expression and compare the changes of expression level and its promoter methylation with or without 5-Aza-2'-dC treatment.Results:(1) The positive rate of ADAM23 protein was significantly lower in NSCLC than that of tissues adjacent to carcinoma and benign lung tissues (38.5% vs 86.5% and 87.5%, P<0.05) whereas the positive rate of avβ3 protein was significantly higher in NSCLC than that of tissues adjacent to carcinoma and benign lung tissues (80.8% vs 26.9% and37.5%, P<0.05). ADAM23 expression had no correlation with patients'age, sex, tumor size and histological type but markedly related to tumor differentiation degree, lymph node metastasis and clinical stage in NSCLC; av(33 expression had no correlation with patients'age, sex, histological type and tumor differentiation degree but evidently related to tumor size, lymph node metastasis and clinical stage in NSCLC. The results of RT-PCR showed that there were no significantly different expression of ADAM23 in NSCLC and tissues adjacent to carcinoma, together with benign lung tissues in NSCLC; whereas the expression of av,β3 was significantly higher than that of adjacent normal tissues and benign lung tissues. The positive rates between ADAM23 and avβ3 were associated.(X2=9.026,γ=-0.417.P＜.05).(2) ADAM23 promoter methylation were observed in 21 of 52 (40.4%) lung cancer specimens and 4 of 52 (7.6%; 4/52) adjacent normal tissues as well as 0 of 8 benign pulmonary lesions (0/8). In cancer tissues of ADAM23 negative expression, the rate of ADAM23 gene methylation was 50.3%(17/32). ADAM23 expression and its promoter methylation were negatively associated (r=-0.328,p=0.017).(3) In four lung cancer cells, the expression level of ADAM23 was much higher in A549 and SK-MES-1 than in SPC-A1, LTEP-a-2 which show significantly lower or absent. Moreover, the results of MSP showed ADAM23 promoter methylation were observed in SPC-A1, LTEP-a-2 that methylated primer can amplify the products. While, as to SK-MES-1 and A549, only unmethylated primer can amplify the products.(4) After incubated lung cancer cells of SPC-A1,LTEP-a-2 with 5-Aza-2'-dC for three days, the expression of ADAM23 was significantly high and its level of methylation decreased.Conclusions:(1) ADAM23 is lower-expressed while avβ3 is over-expressed in NSCLC. They may play an important role in invasion and metastasis of NSCLC.(2) ADAM23 gene downregulation is closely correlated with promoter methylation in NSCLC and were negatively associated. We proposed that promoter aberrant methylation is probably becoming one of the most important mechanism that lead to silence the expression of ADAM23 (3) ADAM23 promote cancer cell invasion and metastasis via negatively modulates integrin avβ3 activation. |
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