Yidong Cheng

A lentiviral sponge for miRNA-21 diminishes aerobic glycolysis in bladder cancer T24 cells via the PTEN/PI3K/AKT/mTOR axis.

Cancer cells exhibit the ability to metabolise glucose to lactate even under aerobic conditions for energy. This phenomenon is known as the Warburg effect and can be a potential target to kill cancer cells. Several studies have shown evidence for interplay between microRNAs and key metabolic enzyme effecters, which can facilitate the Warburg effect in cancer cells. In the present study, a microRNA sponge forcibly expressed using a lentiviral vector was utilised to knock down miR-21 expression in vitro. qPCR and Western blot assays were performed to evaluate the expression of a regulatory factor related to aerobic glycolysis and the signalling pathway it regulates. In bladder cancer specimens, expression levels of glycolysis-related genes [glucose transporter (GLUT)1, GLUT3, lactic dehydrogenase (LDH)A, LDHB, hexokinase (HK)1, HK2, pyruvate kinase type M (PKM) and hypoxia-inducible factor 1-alpha (HIF-1α)] were higher in tumour tissues than in adjacent tissues, suggesting the role of glycolysis in bladder cancer. miR-21 inhibition in bladder cancer cell lines resulted in reduction in tumour aerobic glycolysis. Decrease in glucose uptake and lactate production was observed upon expression of the miR-21 sponge, which promoted phosphatase and tensin homologue (PTEN) expression, decreased phosphorylated AKT and deactivated mTOR. Furthermore, messenger RNA (mRNA) and protein expression levels of glycolysis-related genes were also lower in miR-21 sponge cells compared to miR-21 control cells. Our findings suggest that miR-21 acts as a molecular switch to regulate aerobic glycolysis in bladder cancer cells via the PTEN/phosphatidylinositol 3-kinase (PI3K)/AKT/mTOR pathway. Blocking miR-21 function can be an effective diagnostic and therapeutic approach either by itself or in combination with existing methods to treat bladder cancer.

MicroRNA-218 inhibits bladder cancer cell proliferation, migration, and invasion by targeting BMI-1.

MicroRNAs (miRNAs) are recognized as important molecules and have emerged as important gene regulators in tumorigenesis. Growing evidence suggested that miR-218 was a tumor suppressor in many human cancers. However, its underlying role in bladder cancer (BCa) remains unclear. The aim of this study was to explore the effect of miR-218 on the proliferation, migration, and invasion of BCa cells. We found that miR-218 was frequently downregulated in BCa tissues compared with normal adjacent tissues. In vitro and in vivo assays demonstrated that miR-218 overexpression in the BCa cells inhibited cell proliferation, migration, and invasion. Luciferase reporter assay showed that BMI-1 was a direct target of miR-218. In addition, we found that miR-218 regulated the expression of BMI-1 and its downstream target (PTEN) and participated in the phosphorylation of AKT. Our findings indicate that miR-218 functions as tumor suppressor in BCa, and the miR-218/BMI-1 axis may provide novel diagnostic and therapeutic strategies for the treatment of BCa.

Targeting protein kinase CK2 suppresses bladder cancer cell survival via the glucose metabolic pathway

Casein kinase 2 (CK2) is a constitutively active serine/threonine kinase that promotes cell proliferation and resists apoptosis. Elevated CK2 expression has been demonstrated in several solid tumors. The expression of CK2α in bladder cancer was elevated in tumor tissues compared with that in adjacent normal tissues. Amplified expression of CK2α was highly correlated with histological grade in bladder cancer(P = 0.024). Knockdown of CK2α in bladder cancer cell lines resulted in a reduction in tumor aerobic glycolysis, accompanied with lower phosphorylated AKT. Moreover, low CK2α levels suppressed cell growth, and similar results could be reproduced after treatment with CX-4945 with a dose-dependent response. CX-4945 inhibited migration and induced apoptosis. Furthermore, knockdown of CK2α decreased the tumorigenicity of bladder cancer cells in vivo. This study is the first to report that CK2 increases glucose metabolism in human bladder cancer. Blocking CK2 function may provide novel diagnostic and potential therapeutic.

MiR-200c promotes bladder cancer cell migration and invasion by directly targeting RECK

Background Increasing evidence suggests that the dysregulation of certain microRNAs plays an important role in tumorigenesis and metastasis. MiR-200c exhibits a disordered expression in many tumors and presents dual roles in bladder cancer (BC). Therefore, the definite role of miR-200c in BC needs to be investigated further. Materials and methods Quantitative reverse transcription polymerase chain reaction was used to assess miR-200c expression. Cell invasion and migration were evaluated using wound healing and transwell assays. The luciferase reporter assay was used to identify the direct target of miR-200c. The expression of reversion-inducing cysteine-rich protein with kazal motifs (RECK) in BC tissues and adjacent nontumor tissues, as well as in BC cell lines, was detected through quantitative reverse transcription polymerase chain reaction, Western blot assay, and immunohistochemistry. Results The miR-200c expression was significantly upregulated in the BC tissues compared with the adjacent nontumor tissues. The downregulation of miR-200c significantly inhibited cell migration and invasion in the BC cell lines. The luciferase reporter assay showed that RECK was a direct target of miR-200c. The knockdown of RECK in the BC cell lines treated with anti-miR-200c elevated the previously attenuated cell migration and invasion. Conclusion Our findings indicated that miR-200c functions as oncogenes in BC and may provide a novel therapeutic strategy for the treatment of BC.

MicroRNA-218 Increases the Sensitivity of Bladder Cancer to Cisplatin by Targeting Glut1

Background/Aims: MicroRNA-218 (miR-218) is down-regulated in many malignancies that have been implicated in the regulation of diverse processes in cancer cells. However, the involvement of miR-218 in chemo-sensitivity to cisplatin and the precise mechanism of this action remained unknown in bladder cancer. Methods: qRT-PCR was used to detect miR-218 and its target Glut1 expression in bladder cancer cell lines T24 and EJ. CCK-8 method was utilized to measure the cell viability. IC 50 was calculated via a probit regression model. Glut1 was detected by western blotting for analysis of potential mechanism. Luciferase reporter assay was utilized to validate Glut1 as a direct target gene of miR-218. The intracellular level of GSH and ROS were determined using a commercial colorimetric assay kit and 2’, 7’-dichlorodihydro-fluorescein diacetate, respectively. Results: Over-expression of miR-218 significantly reduced the rate of glucose uptake and total level of GSH and enhanced the chemo-sensitivity of bladder cancer to cisplatin. Mechanistically, Glut1 was found to be a direct and functional target of miR-218. Up-regulation of Glut1 could restore chemo-resistance in T24 and EJ cells. On the contrary, knockdown of Glut1 could generate a similar effect as up-regulating the expression of miR-218. Conclusions: MiR-218 increases the sensitivity of bladder cancer to cisplatin by targeting Glut1. Restoration of miR-218 and repression of glut1 may provide a potential strategy to restore chemo-sensitivity in bladder cancer.

GSTP1 and GSTO1 single nucleotide polymorphisms and the response of bladder cancer patients to intravesical chemotherapy

SNPs may restrict cell detoxification activity and be a potential risk factor for cancer chemosensitivity. We evaluated the predictive value of these polymorphisms on the sensitivity of bladder cancer patients to epirubicin and mitomycin chemotherapy instillation as well as their toxicities. SNPs were analyzed by TaqMan genotyping assays in 130 patients treated with epirubicin and 114 patients treated with mitomycin. Recurrence-free survival (RFS) was estimated by the Kaplan-Meier method, and hazard ratios (HRs) and 95% confidence intervals (CIs) of the HRs were derived from multivariate Cox proportional hazard models. GSTP1 rs1695 and GSTO1 rs4925 were also associated with RFS in the epirubicin group. Patients carrying the GSTP1 AG+GG and GSTO1 AC+AA genotypes had an unfavorable RFS. Patients with the GSTP1 AA and GSTO1 CC genotypes had a reduced risk of recurrence after the instillation of epirubicin. In addition, patients with the GSTP1 rs1695 AA genotype had an increased risk of irritative voiding symptoms; while patients with the GSTO1 rs4925 CC genotype had a decreased risk of hematuria. Our results suggest that GSTP1 and GSTO1 polymorphisms are associated with epirubicin treatment outcomes as well as with epirubicin-related toxicity.

Urine microRNAs as biomarkers for bladder cancer: a diagnostic meta-analysis.

Background The diagnostic value of microRNA (miRNA) detection in patients with bladder cancer (BCa) is controversial. We performed a diagnostic meta-analysis to evaluate current evidence on the use of miRNA assays to diagnose BCa. Methods We systematically searched PubMed, Embase, and Web of Science for studies published before March 31, 2015. The pooled sensitivity, specificity, positive and negative likelihood ratios, diagnostic odds ratio, and area under the curve (AUC) were calculated to evaluate the overall test performance. Subgroup analyses were used to explore the between-study heterogeneity. Deeks’ funnel plot asymmetry test was used to test publication bias. We applied the software of RevMan 5.2 and Stata 11.0 to the meta-analysis. Results A total of 23 studies from nine articles were included in the meta-analysis, with a total of 719 patients and 494 controls. The pooled sensitivity and specificity were 0.75 (95% confidence interval [CI], 0.68–0.80) and 0.75 (95% CI, 0.70–0.80), respectively. The pooled positive likelihood ratio was 3.03 (95% CI, 2.50–3.67); negative likelihood ratio was 0.33 (95% CI, 0.27–0.42); and diagnostic odds ratio was 9.07 (95% CI, 6.35–12.95). The pooled AUC was 0.81 (95% CI, 0.78–0.85). Subgroup analyses indicated that the multiple miRNAs assays and urine supernatant assays showed high accuracies in diagnosing BCa. Conclusion The miRNA assays may serve as potential noninvasive diagnostic tool for the detection of BCa. However, the clinical application of miRNA assays for BCa diagnosis still needs further validation by large prospective studies.

Perioperative treatments for resected upper tract urothelial carcinoma: a network meta-analysis

Background Perioperative treatments have been used to improve prognosis in patients with upper tract urothelial carcinoma (UTUC). However, optimal management remains unestablished. Methods We searched the Embase, Web of Science and Cochrane databases for studies published before June 20, 2015. All included studies were categorised into three groups on the basis of the outcome reported (overall survival (OS), disease-specific survival (DSS) and recurrence-free survival (RFS)). Relative hazard ratios (HRs) for death were calculated using random-effects Bayesian network meta-analysis methods. We also ranked the three different treatments in terms of three outcomes. Results A total of 31 trials with 8100 patients were included. Compared with the control, adjuvant chemotherapy (AC) could improve OS, DSS and RFS by 32% (HR 0.68, 95% CI 0.51-0.89), 29% (HR 0.71, 95% CI 0.54-0.89) and 51% (HR 0.49, 95% CI 0.23-0.85), respectively. We noted a marked prolongation of RFS in both intravesical chemotherapy (HR 0.32, 95% CI 0.09-0.69) as well as concurrent radiotherapy and intravesical chemotherapy (HR 0.32, 95% CI 0.03-0.97) than in the control. Neoadjuvant chemotherapy (NAC) showed a significant improvement in DSS relative to the control (HR 0.25, 95% CI 0.06-0.61) and a distinct advantage over AC (HR 0.36, 95% CI 0.08-0.90) or AR (HR 6.89, 95% CI 1.25-18.66). Conclusions Our results showed that AC; intravesical chemotherapy; and concurrent radiotherapy and intravesical chemotherapy could improve the prognosis of UTUC patients. NAC was found to be more favourable for UTUC than AC in terms of DSS.

Identification of circulating microRNA signatures for upper tract urothelial carcinoma detection

The involvement of circulating microRNAs (miRNAs) in cancer and their potential as diagnostic and prognostic biomarkers are becoming increasingly known. However, the significance of circulating miRNAs in upper tract urothelial carcinoma (UTUC) has remained to be investigated. The present study performed a genome wide serum miRNA analysis using a deep sequencing platform for initial screening. Subsequently, serum samples of 46 UTUC patients and 30 cancer free individuals with hematuria were subjected to a quantitative reverse transcription polymerase chain reaction analysis. The expression of thirteen miRNAs (miR-664a-3p, miR-423-5p, miR-431-5p, miR-191-5p, miR-92a-3p, miR-22-3p, miR-26a-5p, miR-33b-3p, miR-16-5p, let-7a 5p, let-7b-5p, let-7f-5p and let-7c) was significantly different in serum from UTUC patients compared with that in control samples. Receiver operator characteristic analysis showed that 10 miRNAs (miR-664a-3p, miR-431-5p, miR-423-5p, miR-191-5p, miR-33b-3p, miR-26a-5p, miR-22-3p, miR-16-5p, let-7b-5p and let-7c) had the potential to distinguish individuals with UTUC from the controls (areas under the curve >0.8). The present study provided the first evidence for the potential use of circulating miRNAs as biomarkers for UTUC diagnosis, which remains to be verified by further studies.

Meta-Analysis Reveals a Lack of Association between UGT2B17 Deletion Polymorphism and Tumor Susceptibility

Purpose UGT2B17 is a vital member of the UGT2 family and functions as a detoxification enzyme which catalyzes the glucuronidation of lipophilic compounds. Accumulating evidences implicates that it may contribute to the susceptibility of tumor risk. Identification of a UGT2B17 deletion polymorphism has attracted studies to evaluate the association between the UGT2B17 deletion polymorphism and tumor risk in diverse populations. However, the available results are conflicting. Methods A meta-analysis based on 14 studies from 10 publications including 5,732 cases and 5,112 controls was performed. Published literature from PubMed, EMBASE and Web of Science was pooled and the crude odds ratios (ORs) with 95% confidence intervals (CIs) were calculated to estimate the strength of the associations. Results Conclusively, our results indicate that individuals with a UGT2B17 deletion polymorphism were associated with tumor risks (OR = 1.29, 95% CI = 1.03–1.63, P<0.001) in a recessive model. However, after excluding two studies for their heterogeneity, the result then demonstrated that the UGT2B17 deletion polymorphism was not associated with tumor risks (OR = 1.118, 95%CI = 0.938–1.332, P>0.1). A subgroup analysis based on tumor type, sex or race did not show significant results. Conclusion These results suggest that the UGT2B17 deletion polymorphism is not associated with tumor risks.