Vascular Endothelial Growth Inhibitor, a Cytokine of the Tumor Necrosis Factor Family, is Associated With Epithelial-Mesenchymal Transition in Renal Cell Carcinoma.

Previous studies have revealed that the activation of the epithelial-mesenchymal transition (EMT) endows metastatic properties upon cancer cells to promote invasion and migration. In this study, immunohistochemical analysis was performed in 50 cases of clear cell renal cell carcinoma (RCC) and paired normal kidney tissues. We detected the expression of vascular endothelial growth inhibitor (VEGI) and EMT markers (E-cadherin, fibronectin, and Slug) and recorded the clinical, pathologic, and follow-up (median follow-up: 79.0 mo) information. The expression of VEGI and E-cadherin was significantly lower in RCC tissues compared with normal kidney tissues (P<0.001). However, the expression of fibronectin and Slug was higher in RCC tissues (P<0.05). VEGI and EMT marker expression marginally differed in tumor size and stage. Significant differences were found in the pathologic grade (P<0.05). The Spearman correlation analysis suggested a positive correlation between VEGI and E-cadherin (r=0.451, P<0.01). A negative correlation was shown between VEGI and fibronectin (r=-0.465, P<0.01). There was also a negative correlation between VEGI and Slug (r=-0.758, P<0.01). During the 79.0 months (range, 7 to 119 mo) of follow-up, 6 patients died due to RCC, and the tumor-free survival rate was 88% (44/50). We did not find a significant correlation between VEGI/EMT markers (E-cadherin, fibronectin, and Slug) and overall survival (P>0.05). Our findings indicate that VEGI plays an important role in EMT in RCC. It suggests that VEGI may be investigated as a disease biomarker and therapeutic target in RCC.

Vascular endothelial growth inhibitor 174 and its functional domains inhibit epithelial-mesenchymal transition in renal cell carcinoma cells in vitro.

The present study was carried out to investigate the effects of vascular endothelial growth inhibitor 174 (VEGI174) and its functional domains (V7 and V8) on epithelial­mesenchymal transition (EMT) in renal cell carcinoma (RCC) cells in vitro. The RCC cell lines A498 and 786­O were used in this study. Based on our preliminary study, we selected full­length VEGI174 and its functional domains (V7 and V8) as the target genes in this study. Plasmids containing VEGI174, V7 or V8 transgenes were constructed and transfected into A498 and 786­O cell lines. Cytological activity was tested during cell culture. Quantitative PCR and western blot analysis were performed to determine the expression levels of EMT markers (E­cadherin, vimentin, ß­catenin and Slug). Overexpression of VEGI174, V7 or V8 did not have a significant influence on cell viability (P>0.05). The mRNA level of E­cadherin was significantly upregulated, while that of vimentin was downregulated in A498VEGIexp, A498V7exp, A498V8exp, 786­OVEGIexp, 786­OV7exp and 786­OV8exp cells compared with the cells containing the empty plasmid controls (P<0.05). The western blot results showed that changes in protein expression levels were consistent with the changes in mRNA expression. Both the mRNA and protein expression levels of ß­catenin and Slug were downregulated in the A498VEGIexp, A498V7exp, A498V8exp, 786­OVEGIexp, 786­OV7exp and 786­OV8exp cells. In conclusion, overexpression of VEGI174, V7 or V8 inhibited EMT in A498 and 786­O cells. Notably, V7 and V8 are two effective functional domains of VEGI174 that have the potential to be studied for peptide synthesis and the treatment of RCC.

Intra-tumour molecular heterogeneity of clear cell renal cell carcinoma reveals the diversity of the response to targeted therapies using patient-derived xenograft models.

Inter- and intra-tumour molecular heterogeneity is increasingly recognized in clear cell renal cell carcinoma (ccRCC). It may partially explain the diversity of responses to targeted therapies and the various clinical outcomes. In this study, a 56-year-old male ccRCC patient with multiple metastases received radical nephrectomy and resection of the metastatic tumour in chest wall. The surgical specimens were implanted into nude mice to establish patient-derived xenograft (PDX) models with KI2367 model derived from the primary tumour and KI2368 model from the metastastic tumour. The two modles were treated with Sorafenib, Sunitinib, Axitinib, combined Sorafenib/Sunitinib, or alternating therapy of Sorafenib and Sunitinib. Significant anti-tumour activity was found in KI2367 treated with Sorafenib/Sunitinib monotherapy, combined Sorafenib/Sunitinib, and alternating therapy of Sorafenib/Sunitinib (P<0.05) but not in that treated with Axitinib monotherapy. In contrast, KI2368 was significantly responsive to Sunitinib monotherapy, combined Sorafenib/Sunitinib therapy and alternating therapy of Sorafenib/Sunitinib but not responsive to Sorafenib and Axitinib monotherapy (P<0.05). RNAseq of the two models demonstrated that the expression levels of 1,725 genes including the drug targeted genes of PDGFA, PDGFB and PDGFRA were >5-fold higher in KI2367 than in KI2368 and the expression levels of 994 genes were > 5-fold higher in KI2368 than in KI2367. These results suggest the presence of intra-tumour molecular heterogeneity in this patient. This heterogeneity may influence the response to targeted therapies. Multiple biopsy, liquid biopsy and genomic analysis of intra- tumour molecular heterogeneity may help guide a more precise and effective plan in selecting targeted therapies for ccRCC patients.

Cobalt Chloride-induced Hypoxia Induces Epithelial-mesenchymal Transition in Renal Carcinoma Cell Lines.

OBJECTIVE: To establish epithelial-mesenchymal transition (EMT) models in renal cell carcinoma (RCC) cell lines. MATERIALS AND METHODS: The RCC cell lines A498 and 786-O were used in the experiment and CoCl2 was used to simulate hypoxia. Cells were cultured with different concentrations of CoCl2. Morphology and changes in cytoactivity were observed. After CoCl2 treatment, the expression of HIF-1α and the changes of EMT-related molecules (E-cadherin, fibronectin) were detected. RESULTS: Cell conjunctions of CoCl2-treated groups were loose and scattered compared to the control. CoCl2 did not promote or attenuate the viability of A498 cells at low dosage, but when the concentration of CoCl2 reached 250 µM, cell activity gradually declined. In contrast, CoCl2 induced 786-O cell proliferation in the range of 50 µ M-200 µ M, but inhibited cell growth at dosages higher than 200 µM. The expression of E-cadherin was significantly down-regulated, and fibronectin was up-regulated in both A498 and 786-O cell lines under CoCl2-simulated hypoxia in comparison with normoxic conditions (P<0.01). CONCLUSIONS: CoCl2-induced hypoxia could induce EMT in RCC cell lines. The models will help us further study the mechanisms of EMT and investigate novel therapeutic targets to inhibit tumor invasion and metastasis.