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Renal cell carcinoma (RCC) is a malignant tumor that can metastasize easily. Hence, many patients have already developed metastasis when they are diagnosed. It is also one of the most common tumors that metastasize to the head and neck through extranodal disease. Herein, we reported a case of a 53-year-old man with cervical metastasis from bilateral RCC. Interestingly, whole exome sequencing (WES) and clonal evolution analysis revealed that bilateral renal tumor lesions and neck metastases (squamous cell carcinoma) share the same subclones and a large number of gene variants, while the pathological morphology is different (left nephrotic foci, a mixed pattern of mucinous tubular and spindle cell carcinoma (MTSCC) with papillary adenoma; right renal foci, papillary renal cell carcinoma (PRCC)). This was first reported in RCCs to the best of our knowledge. This case suggests that genotype analysis can be a powerful supplementary examination for clinical histopathological diagnosis. Gene detection has great significance for the accurate diagnosis and treatment of RCC metastasis or multiple lesions.

Bladder cancer (BCa) is a common malignant tumor of urinary system with few treatments, so more useful therapeutic targets are still needed. Antitumor effects of tristetraprolin (TTP) have been explored in many type tumors, but its roles in bladder cancer are still unknown until now. In this study, public expression profiles and tissue microarray analysis showed that TTP mRNA and protein levels decreased in BCa relative to the normal bladder tissue. To explore biological functions of TTP in BCa, 488 TTP target genes, which could be both suppressed and bound by TTP, were identified by comprehensively analyzing publicly available high‐throughput data obtained from Gene Expression Omnibus (GEO). Gene enrichment analysis showed that these genes were enriched in pathways such as cell cycle, epithelial to mesenchymal transition (EMT), and Wnt signaling. Clustering analysis and gene set variation analysis indicated that patients with high expression of TTP target genes had poorer prognosis and stronger tumor proliferation ability relative to the BCa patients with low expression of TTP target genes. In vitro experiments validated that TTP could suppress proliferation, migration, and invasiveness of BCa cells. And TTP could suppress mRNA expression of cyclin‐dependent kinase 1 (CDK1) in BCa cells by target its 3′ UTR. Then, we identified a new small double‐stranded RNA (dsRNA) named dsTTP‐973 which could increase TTP expression in BCa cells, in vivo and in vitro experiments revealed that dsTTP‐973 could suppress aggressiveness of BCa. In conclusion, TTP played a role of tumor suppressor gene in BCa like other tumors, and its dsRNA named dsTTP‐973 could induce TTP expression in BCa and suppress aggressiveness of BCa. With the help of materials science, dsTTP‐973 may become a potential treatment for BCa in the future., TTP overexpression can inhibit the malignant biological behaviors of BCa, and can be a potential therapeutic target for BCa. dsTTP‐973 which can induce TTP expression in BCa is able to suppress BCa progression, and dsTTP‐973 may be a potential treatment for BCa in the future.

Previous studies showed that miR-373 had the capacity to induce tumor suppressor gene E-cadherin expression in prostate cancer cells. However, whether miR-373 can activate the expression of E-cadherin in human bladder cancer (BCa) cells and inhibit cells remains to be elucidated. Here, we found that both miR-373 and E-cadherin were low expressed in BCa tissues and cell lines, and significantly correlated with tumor stage, grade, and lymph node metastasis. In addition, decreased E-cadherin expression or low expression of both miR-373 and E-cadherin is associated with poor overall survival in patients with BCa. Transfection of miR-373 into BCa cells readily activated E-cadherin expression by targeting promoter. Moreover, miR-373 exhibited robust capacity to inhibit cells proliferation, suppress migration and invasion by enhancing E-cadherin expression, and significantly suppress the growth of xenografts and metastasis in nude mice. Altogether, our findings indicate that miR-373 may as a tumor suppressor in BCa by activating E-cadherin expression.

Malignant pheochromocytoma (PHEO) is diagnosed only when metastasis has occurred, making it less likely for patients to obtain the benefits of traditional chemotherapy. Anti-oncogene TP53 mutation has been detected in PHEO and is possibly related to disease progression. However, whether the upregulation of wild-type TP53 has antitumoral effects on PHEO remains completely unknown. In the present study, we used RNA activation (RNAa) technique to upregulate the expression of wild-type TP53 by transfecting synthetic dsP53‑285 into PHEO cell line PC12. We found that the upregulation of wild-type p53 blocked the transition of PC12 cells from the G0/G1 to the S phase, with induction of apoptosis. Additionally, the above-mentioned findings were attested in vivo. Most importantly, dsP53-285-induced antitumoral effects were reversible following co-transfection with siRNA that targeted p53 mRNA. Collectively, our results revealed that the upregulation of p53 and possibly other anti-oncogenes may provide a potential effective therapeutic strategy for PHEO.

Immune checkpoint inhibitors (ICIs) are novel treatments that significantly improve the survival time of MIBC patients, but immunotherapeutic responses are different among MIBC patients. Therefore, it is urgent to find predictive biomarkers that can accurately identify MIBC patients who are sensitive to ICIs. In this study, we computed the relative abundances of 24 immune cells based on the expression profiles of MIBC patients using single‐sample gene set enrichment analysis (ssGSEA). Unsupervised clustering analysis of the 24 immune cells was performed to classify MIBC patients into different immune‐infiltrating groups. Genome (gene mutation and copy number variation), transcriptome (mRNA, lncRNA, and miRNA), and functional enrichment were found to be heterogeneous among different immune‐infiltrating groups. We identified 282 differentially expressed genes (DEGs) associated with immune infiltration by comparing the expression profiles of patients with different immune infiltration profiles, and 20 core prognostic DEGs were identified by univariate Cox regression analysis. An immune‐relevant gene signature (TIM signature) consisting of nine key prognostic DEGs (CCDC80, CD3D, CIITA, FN1, GBP4, GNLY, SPINK1, UBD, and VIM) was constructed using least absolute shrinkage and selection operator (LASSO) Cox regression analysis. Receiver operating characteristic (ROC) curves and subgroup analysis confirmed that the TIM signature was an ideal biomarker for predicting the prognosis of MIBC patients. Its value in predicting immunotherapeutic responses was also validated in The Cancer Genome Atlas (TCGA) cohort (AUC = 0.69, 95% CI = 0.63‐0.74) and the IMvigor210 cohort (AUC = 0.64, 95% = 0.55‐0.74). The TIM signature demonstrates a powerful ability to distinguish MIBC patients with different prognoses and immunotherapeutic responses, but more prospective studies are needed to assess its reliability in the future., Immune checkpoint inhibitors (ICIs) are novel treatments for MIBC and significantly improve survival time of MIBC patients, but ICIs are only effective for minority of MIBC patients. We first constructed a tumor immune infiltration–associated gene signature (TIM signature) for MIBC patients to predict prognosis and immunotherapeutic response by comprehensively comparing expression profiles of MIBC patients with different immune infiltrations in this study. This signature might be useful for future clinical immunotherapy of MIBC patients.

Accumulating data suggest that micro RNAs (miRNAs) or double-stranded RNAs (dsRNAs) can activate gene expression by targeting promoters. The cyclin-dependent kinase inhibitor p21 (CIP1/WAF1) (p21) has also been shown to suppress epithelial-mesenchymal transition (EMT) which plays a crucial role in the early stage of tumor metastases and invasiveness. In a previous study, we have reported that miR-370-5p is low-expressed in bladder cancer (BCa) tissues and cell lines. Here, we identified that miR-370-5p and sequence homology dsRNA (dsP21-555) fully complementary to promoter hold the potent abilities to induce p21 expression. Moreover, transfection of miR-370-5p or dsP21-555 into BCa cells remarkably inverts EMT-associated genes (increases epithelial cell makers E-cadherin and β-catenin, and decreases mesenchymal cell markers ZEB1 and Vimentin) expression mainly via regulating p21 expression. Besides, through manipulating p21, both the candidates can retard BCa cell migration and invasion. In summary, our results provide evidence that both endogenous and exogenous small RNAs may function to induce p21 expression by interacting with the similar promoter region and impede BCa metastasis.

Current studies indicate that microRNAs (miRNAs) are widely decreased in various tumors and function as tumor suppressors by inhibiting cancer cell proliferation, survival, invasion, and migration. The potential application of using miRNAs to predict therapeutic responses to multiple types of cancer treatment holds high promise. In current study, we demonstrate that miR-3619-5p is downregulated in bladder cancer (BCa) tissues and cells. Exogenous overexpression of miR-3619-5p in BCa cells inhibits proliferation, migration, and invasion. Moreover, a nude mouse xenograft model shows that miR-3619-5p inhibits BCa cell growth. We also demonstrate that miR-3619-5p leads to the activation of p21 by targeting its promoter in BCa cells. Enforced miR-3619-5p expression consistently leads to the downregulation of β-catenin and cyclin-dependent kinase 2 (CDK2) through predicted binding sites within the β-catenin and CDK2 3′-untranslated regions (UTRs), respectively. Moreover, β-catenin and CDK2 knockdown is able to mimic BCa cells growth and metastasis effects induced by overexpressing miR-3619-5p. We further confirm that miR-3619-5p inhibits Wnt-β-catenin signal pathway and EMT progression in BCa cells. We also found that miR-3619-5p-induced growth arrest and metastasis inhibition are p21-dependent in BCa cells. Taken together, these results confirm that miR-3619-5p plays a tumor suppressive role in BCa by interfering with cell growth and metastasis and may serve as a potential therapeutic target in BCa treatment.

Low expression levels of E‑cadherin are correlated with poor prognosis in patients with bladder cancer (BCa). A small activating RNA (saRNA) targeting a specific promoter region can activate gene expression. In the present study, two small double-stranded RNAs (dsRNAs) targeting the promoter region of human E‑cadherin were designed and synthesized, and the regulatory role of saRNAs in E‑cadherin expression was investigated. The results of reverse transcription-quantitative polymerase chain reaction and western blotting demonstrated that transfection of dsEcad‑346 into the BCa cell lines T24 and 5637 significantly activated E‑cadherin expression. Furthermore, transfection of dsEcad‑346 and miR‑373 induced cell cycle arrest in G0/G1 phase, promoted apoptosis and significantly inhibited migration and invasion of BCa cells. Results of immunofluorescence and western blotting indicated that β-catenin was redistributed from the nucleus to the cell membrane following transfection of dsEcad‑346 and miR‑373. Additionally, the expression of β-catenin/T-cell factor complex (TCF) target genes (c‑MYC, matrix metallopeptidase 2, cyclin D1) was suppressed following transfection of BCa cells with saRNA. Silencing of E‑cadherin expression blocked the inhibitory effect of dsEcad‑346 and miR‑373 on BCa cells. In conclusion, a novel designed dsEcad‑346 can activate the expression of E‑cadherin in BCa cells. saRNA-mediated activation of E‑cadherin expression inhibited the growth and metastasis of BCa cells by promoting the redistribution of β-catenin from nucleus to cell membrane and inhibiting the β-catenin/TCF target genes.

The mechanism of dsRNA-induced gene activation (RNAa) is being gradually unveiled. The plentiful evidence that it existed in mammalian species other than human demonstrated that dsRNA-mediated RNAa is a conservative phenomenon. Simultaneously, accumulating evidence suggested that microRNAs could activate gene expression by targeting promoter. Nevertheless, it is ambiguous whether microRNA-induced gene activation in different human cells is a common phenomenon. The study we performed verified that miR-1236-3p (miR-1236) and miR-370-5p can activate p21 expression in bladder cancer (BCa) T24, EJ cells, and non-small-cell lung carcinoma A549 cells, while in hepatocellular HepG2 cells both microRNAs cannot effectively induce the expression of P21 WAF1/CIP1 (p21). In pancreatic cancer PANC-1 cells, only miR-370-5p had the potent abilities to induce p21 expression rather than miR-1236-3p. Unlike microRNA-mediated RNA activation, we can observe that dsP21-322 significantly activated p21 in above cells. Besides, we demonstrated that miR-1236 and miR-370 inhibited cyclin D1-CDK4/CDK6 pathway while upregulated E-cadherin expression by upregulation of p21 . Overexpression of these two microRNAs in A549 induced cell-cycle arrest and cell senescence, delayed cell proliferation and colony formation, and inhibited migration and invasion. In conclusion, microRNA-mediated RNAa depends on the cell context, and miR-1236 and miR-370 can inhibit non-small-cell lung carcinoma cell growth by upregulating p21 expression in vitro.

Recent studies have shown that miRNAs have potent abilities to activate gene expression by targeting promoter elements, a phenomenon known as RNA activation (RNAa). In the present study, we identified a new endogenous miR-3619-5p which was decreased in prostate cancer tissues and cells compared to corresponding normal controls. Moreover, overexpression of miR-3619-5p readily induced CDKN1A gene expression by directly targeting the putative site in the promoter. Besides, miR-3619-5p possessed considerable capacity to inhibit prostate cancer DU145 and PC3 cell growth, and downregulate several CDKN1A downstream genes, such as cyclin D1, CDK4 and CDK6. Notably, this antitumor function of miR-3619-5p was mainly achieved by stimulating CDKN1A expression.

Qiangqiang Ge,1,* Chenghe Wang,2,* Yajun Ruan,1,* Zhong Chen,1 Jihong Liu,1 Zhangqun Ye1 1Department of Urology, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, Hubei, 2Department of Urology, Shanghai Jiao Tong University Affiliated Sixth People’s Hospital, Shanghai, People’s Republic of China *These authors contributed equally tothis work Abstract: Previous research has reported that a particular double-stranded RNA, named dsP53-285, has the capacity to induce expression of the tumor suppressor gene TP53 in chimpanzee cells by targeting its promoter. Usually, it is the wild-type p53 protein, rather than mutants, which exhibits potent cancer-inhibiting effects. In addition, nonhuman primates, such as chimpanzees, share almost identical genome sequences with humans. This prompted us to speculate whether dsP53-285 can trigger wild-type p53 protein expression in human prostate cancer (PCa) cells and consequently suppress cell growth. The human PCa cell lines LNCaP and DU145 were transfected with dsP53-285 for 72hours. Compared with the dsControl and mock transfection groups, expression of both p53 messenger RNA and p53 protein was significantly enhanced after dsP53-285 transfection, and this enhancement was followed by upregulation of p21, which indirectly indicated that dsP53-285 induced wild-type p53 expression. Moreover, overexpression of wild-type p53 mediated by dsP53-285 downregulated the expression of Cyclin D1 and cyclin-dependent kinase 4/6, thereby inducing PCa cell cycle arrest in G0/G1 phase and then inhibiting cell proliferation and clonogenicity. More importantly, dsP53-285 suppressed PCa cells mainly by modulating wild-type p53 expression. In conclusion, our study provides evidence that dsP53-285 can significantly stimulate wild-type p53 expression in the human PCa cell lines LNCaP and DU145 and can exert potent antitumor effects. Keywords: p53, small activating RNA, prostate cancer

Background microRNA’s function to silence gene expression by targeting 3′UTR regions has been widely studied. And microRNAs, similar to small double-stranded RNAs, have also been implicated to gene activation triggered by promoter-targeted, which is known as RNA activation (RNAa). p21 WAF1/CIP1 (p21), as a key negative regulator of cell proliferation, is frequently down-regulated in various cancers, making it an ideal target for RNAa-based activation to restore its tumor suppressor function in renal cell carcinoma (RCC). Methods Real-time polymerase chain reaction was used to identify the expression of miR-1236 and p21 in RCC cell lines and clinical specimens. Protein expression and signaling pathway modulation were validated through Western blot analysis, whereas p21, direct target of miR-1236, was validated by using chromatin immunoprecipitation assay. The Kaplan-Meier method and the log-rank test were used to calculate overall survival. The CellTiter 96 ® AQ ueous One Solution Cell Proliferation Assay, colony formation assay, and 5-ethynyl-2′-deoxyuridine assays were used to evaluate the effect of miR-1236 on RCC cell proliferation. Results In this study, we found that miR-1236 and p21 were both significantly down-regulated in RCC tissues and cell lines. Combined low expression of both miR-1236 and p21 emerged as an independent prognostic factor for poor clinical survival in 45 patients with RCC. Chromatin immunoprecipitation assay in the human RCC cell lines 786-O and ACHN showed that miR-1236 directly targeted the p21 promoter and that its activation may provide a plausible link between the positive correlation of miR-1236 and p21 in RCC specimens. Functional experiments showed that increased miR-1236 expression inhibited cell proliferation, and decreased CDK4/6 and cyclin D1 expression. Furthermore, knockdown of p21 using small interfering RNA reversed the antiproliferation function of miR-1236, whereas silencing the p21 expression attenuated the function of miR-1236 in RCC cells. Conclusion Our findings provide insight into the specific RNAa of miR-1236–targeted p21 promoter activation in suppressing RCC cell proliferation. Considering the poor prognostic outcomes associated with reduced miR-1236 and p21 expression, our data imply that these factors likely play important antitumor effects in RCC progression.

Background Previous study showed that dsP53-285 has the capacity to induce tumor suppressor gene p53 expression by targeting promoter in non-human primates’ cells. And it is well known that TP53 gene is frequently mutant or inactivated in human bladder cancer. Hereby, whether this small RNA can activate the expression of wild-type p53 and inhibit human bladder cancer cells remains to be elucidated. Methods Oligonucleotide and lentivirus were used to overexpress dsP53-285 and dsControl. Real-time PCR and western blot were used to detect genes’ mRNA and protein expression, respectively. Cell proliferation assay, colony formation, flow cytometry, transwell assay and wound healing assay were performed to determine the effects on bladder cancer cells proliferation and migration/invasion in vitro. Animal models were carried out to analyze the effects on cells growth and metastasis in vivo. Results Transfection of dsP53-285 into human bladder cancer cell lines T24 and EJ readily activate wild-type p53 expression by targeting promoter. Moreover, dsP53-285 exhibited robust capacity to inhibit cells proliferation and colony formation, induce cells G0/G1 arrest, suppress migration and invasion. Besides, the Cyclin-CDK genes (Cyclin D1 and CDK4/6) were down-regulated and the EMT-associated genes (E-cadherin, β-catenin, ZEB1 and Vimentin) were also expressed inversely after dsP53-285 treatment. In addition, dsP53-285 could also significantly suppress the growth of bladder cancer xenografts and metastasis in nude mice. Most importantly, the anti-tumor effects mediated by dsP53-285 were mainly achieved by manipulating wild-type p53 expression. Conclusion Our findings indicate that the dsP53-285 can upregulate wild-type p53 expression in human bladder cancer cells through RNA activation, and suppresses cells proliferation and metastasis in vitro and in vivo. Electronic supplementary material The online version of this article (doi:10.1186/s13046-016-0329-8) contains supplementary material, which is available to authorized users.