Yonghao Zhan

Tetracycline-inducible shRNA targeting long non-coding RNA PVT1 inhibits cell growth and induces apoptosis in bladder cancer cells

Recent studies show that long non-coding RNAs (lncRNAs) may be significant functional regulators in tumor development, including bladder cancer. Here, we found that PVT1 was upregulated in bladder cancer tissues and cells. Further experiments revealed that PVT1 promoted cell proliferation and suppressed cell apoptosis. Furthermore we also used the emerging technology, synthetic biology, to create tetracycline-inducible small hairpin RNA (shRNA) vectors which silenced PVT1 in a dosage-dependent manner to inhibit the progression of bladder cancer. In conclusion, data suggest that PVT1 could be an oncogene and may be a therapeutic target in bladder cancer. Synthetic “tetracycline-on” switch system can be used to quantitatively control the expression of PVT1 in bladder cancer in response to different concentration of doxycycline to suppress the progression of bladder cancer.

Up-regulation of long non-coding RNA PANDAR is associated with poor prognosis and promotes tumorigenesis in bladder cancer.

BackgroundLong non-coding RNAs (lncRNAs) have emerged as biomarkers and important regulators of tumor development and progression. PANDAR (promoter of CDKN1A antisense DNA damage activated RNA) is a novel long non-coding RNA that acts as a potential biomarker and involves in development of multiple cancers. However, the clinical significance and molecular mechanism of PANDAR in bladder cancer is still unknown. In this study, we aimed to figure out the role of PANDAR in bladder cancer.MethodsThe relative expression level of lncRNA PANDAR was determined by Real-Time qPCR in a total of 55 patients with urothelial bladder cancer and in different bladder cancer cell lines. We inhibited PANDAR expression by transfecting PANDAR specific siRNA and enhanced PANDAR expression by transfecting a PANDAR expression vector (pcDNA3.1-PANDAR). Cell proliferation was determined by using both CCK-8 assay and Edu assay. Cell apoptosis was determined by using ELISA assay, Hoechst 33342 staining and Flow cytometry. Cell migration was determined by using transwell assay. All experimental data from three independent experiments were analyzed by χ2 test or Student’s t-test and results were expressed as meanu2009±u2009standard deviation.ResultsWe found that PANDAR was significantly up-regulated in bladder cancer tissues compared with paired-adjacent nontumorous tissues in a cohort of 55 bladder cancer patients. Moreover, increased PANDAR expression was positively correlated with higher histological grade (Pu2009<u20090.05) and advanced TNM stage (Pu2009<u20090.05). Further experiments demonstrated that inhibited cell proliferation/migration and induced apoptosis by silencing PANDAR were also observed in bladder cancer cells. Furthermore, over expression of PANDAR in bladder cancer cells promoted the proliferation/migration and suppressed apoptosis.ConclusionsThese findings demonstrate that PANDAR plays oncogenic roles in bladder cancer and PANDAR may serve as a potential prognostic biomarker and therapeutic target of bladder cancer.

Increased expression of SUMO1P3 predicts poor prognosis and promotes tumor growth and metastasis in bladder cancer

Bladder cancer is one of the most common malignancies worldwide. Long non-coding RNAs (lncRNAs) are a class of non-coding RNAs that play crucial roles in diverse biological processes. The pseudogene-expressed lncRNA is one major type of lncRNA family. Small ubiquitin-like modifier (SUMO) 1 pseudogene 3, (SUMO1P3) is a novel indentified lncRNA that was previously reported to be up-regulated in gastric cancer. However, we know nothing about the biological function and underlying mechanism of SUMO1P3 in tumor. Furthermore, the relationship between SUMO1P3 and bladder cancer is completely unknown. We hypothesized that SUMO1P3 also have roles in bladder cancer. In this study, we found that SUMO1P3 was significantly up-regulated in bladder cancer tissues compared with paired-adjacent nontumorous tissues in a cohort of 55 bladder cancer patients. Moreover, up-regulated SUMO1P3 expression was positively correlated with greater histological grade (P<0.05) and advanced TNM stage (P<0.05). Furthermore, we found cell proliferation / migration inhibition and apoptosis induction were also observed in SUMO1P3 siRNA-transfected bladder cancer cells. Our data suggest that SUMO1P3 plays oncogenic roles in bladder cancer and can be used as a potential prognostic and therapeutic target.

Long non-coding RNA HNF1A-AS1 promotes proliferation and suppresses apoptosis of bladder cancer cells through upregulating Bcl-2

Emerging evidences have indicated that long non-coding RNAs (lncRNAs) are pivotal regulators of tumor development and progression. HNF1A-AS1 (HNF1A antisense RNA 1, C12 or f27) is a novel long non-coding RNA that acts as a potential biomarker and is involved in development and progression of several cancers. Nevertheless, we know nothing about the clinical significance and molecular mechanism of HNF1A-AS1 in bladder cancer. In this study, we found that HNF1A-AS1 is significantly up-regulated in bladder cancer. Further experiments had demonstrated that silencing HNF1A-AS1 in bladder cancer cells could inhibit the proliferation and induce apoptosis. Mechanistically, we found down-regulated of HNF1A-AS1 increased the expression of miR-30b-5p and subsequently inhibited the expression of Bcl-2, in a ceRNA-dependent way. Moreover, knockdown of miR-30b-5p reversed cell proliferation inhibition and cell apoptosis induced by silencing HNF1A-AS1. In conclusions, we demonstrated that HNF1A-AS1 plays an important regulatory role in bladder cancer and shed new light on lncRNA-directed diagnostic and therapeutics in bladder cancer.

Increased expression of ZEB1-AS1 correlates with higher histopathological grade and promotes tumorigenesis in bladder cancer

Bladder cancer is one of the most common urinary cancers worldwide. Emerging studies indicated that long non-coding RNAs (lncRNAs) play crucial roles in cancer biology. In this study, we found that a novel lncRNA Zinc finger E-box-binding homeebox1 (ZEB1) antisense RNA (ZEB1-AS1) was overexpressed in bladder cancer tissues compared to paired noncancerous tissues. Moreover, the expression of ZEB1-AS1 was positive correlated with higher histological grade and TNM stage in bladder cancer. Furthermore, Loss-of-function experiments showed that down-regulation of ZEB1-AS1 not only can suppress cell growth but also can inhibit migration and induce apoptosis in bladder cancer cell lines 5637 and SW780. In conclusion, these findings indicated that ZEB1-AS1 plays regulatory roles in bladder cancer and it may become a novel molecular biomarker of prognosis and therapy in bladder cancer.

Increased expression of long non-coding RNA CCEPR is associated with poor prognosis and promotes tumorigenesis in urothelial bladder carcinoma

Recent emerging evidences have showed that long non-coding RNAs play important regulatory roles in diverse biological processes of tumor development and progression. CCEPR (cervical carcinoma expressed PCNA regulatory lncRNA) is a novel identified lncRNA that acts as a potential biomarker and involves in development and progression of cervical carcinoma. Nevertheless, we know nothing about the clinical significance and molecular mechanism of CCEPR in bladder cancer. In this study, we found that CCEPR was significantly up-regulated in bladder cancer. Furthermore, up-regulated CCEPR expression was positively correlated with advanced TNM stage and higher histological grade. Moreover, further experiments demonstrated that CCEPR promotes cell proliferation and suppresses cell apoptosis in bladder cancer. Mechanistically, we found CCEPR upregulates the expression of PCNA in mRNA and protein level to promote cancer growth. In conclusions, these findings demonstrated that CCEPR plays an important regulatory role in bladder cancer and may serve as a potential diagnostic biomarker and therapeutic target.

Inhibiting cell migration and cell invasion by silencing the transcription factor ETS-1 in human bladder cancer.

As one of the members of the ETS gene family, the transcription factor v-ets avian erythroblastosis virus E26 oncogene homolog 1 (ETS-1) plays key role in the regulation of physiological processes in normal cells and tumors. In this study, we aimed to investigate the relationship between the transcription factor ETS-1 and malignant phenotypes of bladder cancer. We demonstrated that ETS-1 was up-regulated in human bladder cancer tissue compared to paired normal bladder tissue. In order to evaluate the functional role of ETS-1 in human bladder cancer, vectors expressing ETS-1 shRNA and ETS-1 protein were constructed in vitro and transfected into the human bladder cancer T24 and 5637 cells. Our results showed that the transcription factor ETS-1 could promote cell migration and cell invasion in human bladder cancer, without affecting cell proliferation and apoptosis. In conclusion, ETS-1 plays oncogenic roles through inducing cell migration and invasion in human bladder cancer, and it can be used as a therapeutic target for treating human bladder cancer.

Vitamin C increases 5-hydroxymethylcytosine level and inhibits the growth of bladder cancer

Background5-Hydroxymethylcytosine (5hmC) is converted from 5-methylcytosine (5mC) by a group of enzymes termed ten-eleven translocation (TET) family dioxygenases. The loss of 5hmC has been identified as a hallmark of most types of cancer and is related to tumorigenesis and progression. However, the role of 5hmC in bladder cancer is seldom investigated. Vitamin C was recently reported to induce the generation of 5hmC by acting as a cofactor for TET dioxygenases. In this study, we explored the role of 5hmC in bladder cancer and the therapeutic efficacy of vitamin C in increasing the 5hmC pattern.Results5hmC was decreased in bladder cancer samples and was related to patient overall survival. Genome-wide mapping of 5hmC in tumor tissues and vitamin C-treated bladder cancer cells revealed that 5hmC loss was enriched in cancer-related genes and that vitamin C treatment increased 5hmC levels correspondingly. Vitamin C treatment shifted the transcriptome and inhibited the malignant phenotypes associated with bladder cancer cells in both in vitro cell lines and in vivo xenografts.ConclusionsThis study provided mechanistic insights regarding the 5hmC loss in bladder cancer and a rationale for exploring the therapeutic use of vitamin C as a potential epigenetic treatment for bladder cancer.

Predictive value of gene methylation for second recurrence following surgical treatment of first bladder recurrence of a primary upper‑tract urothelial carcinoma

The clinical relevance of aberrant DNA promoter methylation is being increasingly recognized in urothelial carcinoma. The present study was conducted to explore the methylation status of patients with upper-tract urothelial carcinoma (UTUC) who experienced bladder recurrence, and to evaluate the predictive value of gene methylation for second bladder recurrence and tumor progression. A total of 85 patients with primary UTUC, who experienced bladder recurrence after radical nephroureterectomy, were enrolled between January 2001 and December 2013. Using methylation-sensitive polymerase chain reaction, the promoter methylation statuses of 10 genes were analyzed in the bladder tumor specimens. Among the patient group, 32 patients experienced second bladder recurrence, and bladder progression was detected in 16. With the exception of BRCA1, the methylation rate of the majority of genes tended to gradually increase to varying extents with the number of recurrences; a smaller proportion of primary tumors exhibited gene methylation when compared with the first recurrent tumors and second recurrent tumors. Univariate and multivariate Cox regression analyses revealed that unmethylated GDF15 [hazard ratio (HR)=0.36; 95% confidence interval (CI), 0.14-0.92] and methylated VIM (HR=2.91; 95% CI, 1.11-7.61) in the first recurrent bladder tumor, as well as male gender (HR=2.28; 95% CI, 1.06-4.87), first recurrence interval <8 months (HR=2.34; 95% CI, 1.15-4.78) and primary UTUC tumor size ≥5 cm (HR=3.48; 95% CI, 1.43-8.45) were independent risk factors for a second bladder recurrence after surgery for the first bladder recurrence; the Harrells concordance index (c-index) for the related nomogram was 0.71 (95% CI: 0.61-0.81). Furthermore, methylated CDH1 (HR=2.91; 95% CI, 1.08-7.77) and VIM (HR=4.91; 95% CI, 1.11-21.7) in the first recurrent bladder tumor, male gender (HR=3.6; 95% CI, 1.1-11.73), and primary tumor stage T2-T4 (HR=4.57; 95% CI, 1.22-17.13), multifocality (HR=3.64; 95% CI, 1.19-11.16) and size ≥5 cm (HR=3.1; 95% CI, 1.91-10.54) for the primary UTUC were considered to be predictors of tumor progression; the c-index for the nomogram was 0.88 (95% CI, 0.69-0.92). The present findings demonstrated that promoter methylation of cancer-related genes was frequently observed in patients with urothelial carcinoma, and that the gene methylation rate of certain genes tended to gradually increase with the number of bladder recurrences. This may be used as a predictive factor for a second bladder recurrence and tumor progression after the surgical treatment of the first bladder recurrence.

Using microRNAs as Novel Predictors of Urologic Cancer Survival: An Integrated Analysis

Background MicroRNAs(miRNAs) are involved in the formation, maintenance, and metastasis of urologic cancer. Here, we aim to gather and evaluate all of the evidence regarding the potential role of miRNAs as novel predictors of urologic cancer survival. Methods A systematic review was performed to identify and score all of the published studies that evaluated the prognostic effects of miRNAs in kidney (KCa), bladder (BCa) or prostate cancer (PCa). Where appropriate, the summary effects of miRNAs on urologic cancer were meta-analysed. The reliability of those results was then further validated by an integrated analysis of the TCGA cohort and miRNA panel. Results Of 151 datasets, 80 miRNAs were enrolled in this systematic review. A meta-analysis of the prognostic qualities of each miRNA identified an objective association between miRNA and prognosis. miR-21 was identified as an unfavourable miRNA with the overall survival (HR:2.699, 1.76–4.14, Pu202f<u202f0.001) across various prognostic events. Our further meta-analyses, integrating a parallel TCGA analysis, confirmed these partial previous results and further revealed different summary effects, such as the moderate effect of miR-21 in BCa. The refined miRNA panel (KCa-6: miR-27b, −942, −497, −144, −141 and -27a) was more capable of predicting the overall survival than was any single miRNAs included in it (HR: 3.214, 1.971–5.240, Pu202f<u202f0.01). Conclusions A miRNA panel may be able to determine the prognosis of urologic tumour more effectively and compensate for the unreliability of individual miRNA in estimating prognosis. More large-scale studies are therefore required to evaluate the unbiased prognostic value of miRNAs in urologic cancer effectively.