Identification of a potential tumor suppressor gene, UBL3, in non-small cell lung cancer

Objective: Oncogenes have been shown to be drivers of non-small cell lung cancer(NSCLC), yet the tumor suppressing genes involved in lung carcinogenesis remain to be systematically investigated. This study aimed to identify tumor suppressing ubiquitin pathway genes(UPGs) that were critical to lung tumorigenesis.Methods: The 696 UPGs were silenced by an siRNA screening in NSCLC cells;the potential tumor suppressing UPGs were analyzed, and their clinical significance was investigated.Results: We reported that silencing of 11 UPGs resulted in enhanced proliferation of NSCLC cells, and four UPGs(UBL3, TRIM22, UBE2 G2, and MARCH1) were significantly downregulated in tumor samples compared to that in normal lung tissues and their expression levels were positively associated with overall survival(OS) of NSCLC patients. Among these genes, UBL3 was the most significant one. UBL3 expression was decreased in tumor samples compared to that in paired normal lung tissues in 59/86(68.6%) NSCLCs, was correlated with TNM stage and sex of NSCLC patients, and was significantly higher in non-smoking patients than in smoking patients. Silencing UBL3 accelerated cell proliferation and ectopic expression of UBL3 suppressed NSCLC in vitro and in vivo.Conclusions: These results showed that UBL3 represented a tumor suppressor in NSCLC and may have potential for use in therapeutics and for the prediction of clinical outcome of patients.

The expression of Sox4 in female lung cancer patients and its clinical significance

Objective:To investigate the expression level of sex-determining region Y-related high mobility group box 4(Sox4)in female lung cancer patients and its clinical significance.Methods:213cases of female lung cancer patients who presented in the third affiliated hospital of Kunming medical university were selected for this study.Patients were performed radical surgery of lung cancer;the tumor and adjacent normal tissues were collected.The expression of Sox4 were determined by real-time PCR and immunohistochemical staining.The correlation between Sox4 expression and clinicopathological characteristics was analyzed.Results:The mRNA level of Sox4 was significantly higher in tumor tissues than that in adjacent normal tissues(P <0.05).The positive expression rate of SOX4 in carcinoma tissues was 54.46%,which was significantly higher than that in adjacent normal tissues(23.47%)(P <0.05).The positive expression rate of Sox4 in stageⅢ was 89.47%,which was significantly higher than that in stageⅠ-Ⅱ(P <0.05).The positive expression rate of SOX4 in lung cancer patients with lymph node metastasis was 90.91%,which was significantly higher than that in patients without lymph node metastasis(P <0.05).The positive expression rates of Sox4 in low,middle and high differentiation tissues were 69.05%,32.88% and 23.21% respectively,suggesting that the expression of Sox4 in female lung cancer tissues decreased with the increase of pathological differentiation grades.Conclusion:In female lung cancer,SOX4expression was significantly higher in tumor tissue than that of adjacent normal tissues and was correlated with clinical stage,lymph node metastasis and pathological differentiation.Sox4 expression detection may help determine the prognosis of women with lung cancer and provide a theoretical basis for molecular targeted therapy of lung cancer in women.

Identification of an E3 ligase-encoding gene RFWD3 in non-small cell lung cancer

In order to unveil ubiquitin pathway genes (UPGs) that are essential for non-small cell lung cancer (NSCLC) cell proliferation,we recently conducted a siRNA screening experiment to knockdown the expression of 696 UPGs found in the human genome in A549 and H1975 NSCLC cells.We found that silencing of one of the candidates,RFWD3 that encodes an E3 ubiquitin ligase essential for the repair of DNA interstrand cross-links inresponse to DNA damage,led to dramatic inhibition of NSCLC cell proliferation with significant Z-scores.Knockdown of RFWD3 suppressed colony forming activity of NSCLC cells.We further evaluated the significance of RFWD3 in NSCLCs and found that this gene was more elevated in tumor samples than in paired normal lung tissues and was inversely associated with the clinical outcome of patients with NSCLC.Moreover,RFWD3 expression was significantly higher in smokers than in non-smokers.These results show for the first time that RFWD3 is required for NSCLC cell proliferation and may have an important role in lung carcinogenesis.

TGF-β/Akt/Smad signaling regulates ionizing radiation-induced epithelial-mesenchymal transition in acquired radioresistant lung cancer cells

Objective:To define the properties of lung cancer cells that resisted conventionally fractionated radiation exposure.Methods:Acquired radioresistant lung cancer cell line A549 was constructed by X-ray irradiation with a clinical conventional fraction dose of 2 Gy daily during 30 fractions.Cell morphology,molecular markers,migration capacity and invasion potential were evaluated by the microscope,Western blot,immunofluorescence,wound healing test and transwell chamber assay,respectively.Results:Radioresistant A549 cells shifted from an epithelial to a mesenchymal morphology,termed as epithelial-mesenchymal transition(EMT),and was accompanied by decreased expressions of epithelial markers(F=4.568,P<0.05)and increased expression of mesenchymal markers(F=4.270,P<0.05),greater migratory and invasive capabilities(t=6.386,5.644,P<0.05).The expression of TGF-β,and phosphorylated levels of Akt and Smad3 were also enhanced(F=6.496,4.685,3.370,P<0.05).Furthermore,the EMT phenotype induced by radiation could be reversed through inhibition of TGF-β,Akt or Smad3,indicating a functional relationship be-tween them.Conclusions:EMT mediates acquired radioresistance of lung cancer cells induced by IR with clinical parameters,and the crosstalk mode of TGF-β/Akt/Smad signaling plays a critical regulatory role in this process.