Qifeng Sun

Syk is low-expressed in non-small-cell lung cancer and inversely correlates with patient's survival

The protein tyrosine kinases (PTKs) are a group of enzyme proteins that can phosphorylate substrate protein tyrosine residues, and are involved in many signal transduction pathways. They also play an important role in the control of cell differentiation, proliferation, and spreading. A recent study has found that Syk, as a tumor suppressor of PTKs, is closely related to tumor invasion and metastasis [1]. Syk has been also showed to have potential inhibitory effect in breast, gastric, and pancreatic cancers [2–4]. Sung et al. [5] found that in the mouse mammary gland, loss of one Syk allele profoundly increased proliferation, ductal branching, and invasion of epithelial cells through the mammary fat pad during puberty, and that mammary carcinoma developed after 1 year. An increasing number of clinical studies have revealed a correlation between reduced Syk expression and increased risk of metastasis formation, and Syk is assigned as a potential new prognostic marker in different tumor types [6]. In this study, we examined the expression of Syk in non-small-cell lung cancer (NSCLC) and analyzed its association with the prognosis. The study approved by the Ethics Committee of the Second Hospital of Shandong University included 70 patients (54 males, 16 females; median age of 61 years, ranged from 41 to 76 years) with NSCLC that were diagnosed in the Department of Thoracic Surgery, the Second Hospital of Shandong University. In 70 cases, 28 were diagnosed as adenocarcinoma, 36 squamous-cell carcinoma and 6 large-cell carcinoma. According to Primary Lung Cancer Diagnostic and Treatment Practices (2001), 13 cases belonged to stage I, 33 stage II, 19 stage III, and 5 stage IV. None of the patients received radiation or chemotherapy before surgery. All patients were followed up from January 2007 to December 2010. Tumor tissues were taken from primary tumor area, avoiding necrosis and inflammatory areas. Adjacent tissues were from 2 cm next to the tumor material, and normal lung tissues from 5 cm next to tumor margin. Pathological examination showed no carcinoma cells. Syk expression was immunohistochemically examined using mouse anti-human Syk monoclonal antibody (NeoMarkerS, Fremont, USA) under a light microscope. Three fields of cells were counted, respectively at 400 magnification to determine whether the cells were positive for Syk, and the percentage of stained cells was averaged. Specimens were regarded as Syk negative if ,5% of the cells were stained, 5%–25% as ‘þ’, 25%– 50% as ‘þþ’, and .50% as ‘þþþ’ according to many previous reports [7,8]. The results showed that brown particles could be seen in the cytoplasm in Syk-positive cells under a light microscope (Fig. 1). Syk expression rates were 5.7%, 95.7%, and 100% in tumor, adjacent lung cells, and normal lung cells, respectively. The positive rates were 5.6% in squamous-cell carcinoma, 3.6% in adenocarcinoma, and 16.7% in largecell carcinoma (P 1⁄4 0.394). Syk expression rate in NSCLC tumor cells was significantly lower compared with those in normal lung tissue and adjacent lung tissue (P, 0.05), while there was no difference between adjacent lung tissue and normal lung tissue. These results are similar to previous reports about Syk expression in other types of malignant tumors [4]. However, unlike those reported in previous studies, the Syk expression rates among patients of different pathologic types, differentiation, and clinical stages did not reveal any statistically significant differences. A previous study showed [6] that Syk expression was negatively correlated with differentiation and clinical stages of malignant tumors such as pancreatic cancer, esophageal squamous cell carcinoma, and endometrial cancer. We also noticed that the positive rate of Syk expression in our study was much lower than those reported in previous literatures. This may be explained by the following factors: very low expression of Syk in NSCLC, small sample size, and experimental techniques. Large sample volume and other experimental methods are needed for further study. Other associations between Syk expression and lymph node metastasis, tumor size, and tumor node metastasis have been reported [9], and the results also need further validation. In our study, 47 patients survived .3 years, and Syk expression in this group (8.57%) was higher than that with Acta Biochim Biophys Sin 2013, 45: 149–151 | a The Author 2012. Published by ABBS Editorial Office in association with Oxford University Press on behalf of the Institute of Biochemistry and Cell Biology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences. DOI: 10.1093/abbs/gms102. Advance Access Publication 2 December 2012

Inhibitory Effects of Syk Transfection on Lung Cancer Cell Invasion

OBJECTIVE Spleen tyrosine kinase (Syk) is closely related to tumor invasion and metastasis, and has been shown to have potential inhibitory effects in tumors. In this study, we constructed a eukaryotic expression vector for Syk and analyzed its effects on invasive ability of the A549 non-small cell lung cancer cell line in vitro. METHODS A fragment of Syk was obtained by RT-PCR from human lung cancer cells and cloned into the expression vector pLNCXSyk. After restriction endonuclease digestion, PCR and DNA sequencing confirmation, the recombinant Syk expression plasmid was transfected into A549 human lung cancer cells using lipofectamine protocols. After selection, the cells stably expressed Syk. Detection of Syk expression of the cells by RT-PCR, and invasive ability were examined. RESULTS The eukaryotic expression plamid pLNCXSyk was constructed and expressed stably in the A549 human lung cancer cells. The RT-PCR results showed that Syk mRNA expression was upregulated significantly (P<0.05). Lower invasion through a basal membrane were apparent after transfection (P<0.05). CONCLUSIONS A eukaryotic expression plasmid to cause Syk expression in lung cancer cells can obviously inhibit their invasive ability in vitro.

Effect of Smac and Taxol on non-small-cell lung cancer

A series of structurally unique second mitochondria-derived activator of caspases (Smacs) that act as antagonists of the inhibitor of apoptosis proteins (IAPs) directly have been discovered. They play crucial roles in mitochondrial apoptosis pathways and promote chemotherapy-induced apoptosis. In this study, we constructed a eukaryotic expression vector pcDNA3.1/Smac and transfected it into A549 human lung cancer cells. Then we analyzed the cell invasive and cloning ability, as well as cell apoptosis induced by Taxol. The results showed that over-expressed Smac significantly inhibited A549 cell invasive and cloning ability and promoted apoptosis following Taxol treatment. This finding provides a potential approach for the biological therapy of lung cancer.

Effect of spleen tyrosine kinase on nonsmall cell lung cancer.

Aim of Study: To investigate the anti.tumor effect of spleen tyrosine kinase. (Syk) on the human nonsmall cell lung cancer cells in vitro and its mechanism. Materials and Methods: In this study, we constructed a eukaryotic expression vector pcDNA3.1D/V5-His-TOPO/Syk and transfected it into human nonsmall cell lung cancer cells A549. Then, we not only analyzed the expression of Syk in transfected cells and its invasion but also the expression of matrix metalloproteinase-9 (MMP-9). Results: The results showed that overexpressed Syk significantly inhibited A549 cell invasive ability by decreasing the expression of MMP-9. Conclusion: The overexpressed Syk plays an important role in tumor invasion and metastasis, and a negative role in human nonsmall cell lung cancer cells.

Involvement of syk and VEGF-C in invasion of lung adenocarcinoma A549 cells

BACKGROUND AND AIMS Lung cancer has become one of the most dangerous malignant tumors in the world nowadays, whose pathogenesis is complex involving multi-genes and multi-elements. This study aims to investigate the values of spleen tyrosine kinase (Syk) and vascular endothelial growth factor-C (VEGF-C) in lymphangiogenesis and metastasis of lung adenocarcinoma A549 cells. MATERIALS AND METHODS The pcDNA3.1-VEGF-C and pLNCX-syk were constructed and transfected into A549 cells. After cells with stable expression were sorted, the level of VEGF-C was tested by RT-PCR and immunohistochemistry and the mRNA of syk was tested by RT-PCR. The cell invasion assay was investigated by transwell chamber in vitro. Restriction enzyme digestion and gel electrophoresis demonstrated successful construction of the pcDNA3.1-VEGF-C. RESULTS RT-PCR and immunohistochemistry revealed higher expression of VEGF-C in VEGFC-construct-transfected A549 cells than that in controls (P < 0.05). Successful construction of the pLNCX-syk was demonstrated by restriction enzyme electrophoresis and sequencing. RT-PCR revealed Syk expression higher in syk-construct-transfected cells than in controls (P < 0.05). CONCLUSIONS The results indicate a potential link between the upregulation of Syk and VEGF-C expression and lung adenocarcinoma.