Upregulation of homeobox D10 expression suppresses invasion and migration of clear cell renal cell carcinoma through targeting of E-cadherin.

BACKGROUND: Clear cell renal cell carcinoma (CCRCC) is one of the most common types of renal cell carcinoma. Accumulating evidence indicates that homeobox D10 (HOXD10) acts as a tumor suppressor or oncogene in various carcinomas. However, the regulation and potential mechanisms of HOXD10 in CCRCC remain largely unknown. PURPOSE: To explore the effect and potential mechanism of HOXD10 on the invasion and migration of CCRCC cells. METHODS: The expression of HOXD10, E-cadherin and other epithelial mesenchymal transition (EMT)-related proteins was assessed by reverse transcription-quantitative real-time PCR (qRT-PCR) and Western blots. A series of functional assays were performed in RCC cell lines to explore the function of HOXD10 in CCRCC progression. Bioinformatics analysis, ChIP assays, and dual luciferase reporter assays were utilized to identify the interaction between HOXD10 and E-cadherin. RESULTS: Low expression of HOXD10 and E-cadherin was observed in CCRCC tissues and ACHN and 786-O cells. Downregulation of HOXD10 expression was correlated with the TNM stage of CCRCC patients. Functional experiments demonstrated that malignant biological ability was significantly inhibited by HOXD10 overexpression in RCC cells. Moreover, E-cadherin was a potential target gene of HOXD10, as evidenced by a series of assays. In addition, overexpression of HOXD10 inhibited the progression of CCRCC by regulating the expression of E-cadherin, vimentin, and ß-catenin in vitro. CONCLUSION: HOXD10 acts as a tumor suppressor and suppresses invasion and migration of CCRCC cells by regulating E-cadherin and EMT processes. Thus, targeting HOXD10 may be a therapeutic strategy for CCRCC treatment.

KLK4 Silencing Inhibits the Growth of Chromophobe Renal Cell Carcinoma through ERK/AKT Signaling Pathway.

INTRODUCTION: Renal cell carcinoma (RCC) generally has a poor prognosis because of late diagnosis and metastasis. Despite its abundance in RCC cells, the functions of kallikrein-related peptidase 4 (KLK4) in RCC cells remain unknown. The results of this investigation were examined to discover if KLK4 gene silencing influences the development of RCC cells. METHODS: The mRNA levels of KLK4 and the relationship between KLK4 and tumor stage in patients with RCC were analyzed from the GEPIA database. Real-time PCR and Western blotting were used to measure the mRNA and protein levels of KLK4. Cell Counting Kit 8 (CCK-8), colony formation, wound healing, and Transwell assays were used to examine the proliferation, invasion, and migration of RCC cells after KLK4 suppression. Finally, xenograft experiments in a mouse model helped understand the in vivo effects of KLK4 knockdown. RESULTS: Our research found that KLK4 expression was upregulated in the kidney chromophobe (KICH) specimens and cell lines. Moreover, inhibiting KLK4 growth led to a slowdown in RCC cell proliferation and colony formation. Additionally, KLK4 knockdown inhibited migration, invasion, and epithelial-mesenchymal transition (EMT) of RCC cells. AKT and ERK phosphorylation were enhanced with KLK4 silencing. In the nude mouse xenograft cancer model, KLK4 silencing also prevented the expression of Ki-67, CD105, and the growth of tumors. CONCLUSION: KLK4 accelerated KICH progression via the ERK/AKT signaling pathway, providing a novel regulatory mechanism for KICH pathogenesis.

RUNX2 Mediates Renal Cell Carcinoma Invasion through Calpain2.

Runt-related transcription factor 2 (RUNX2), a specific transcription factor of osteocytes, has been confirmed to be involved in the malignant biological behavior of various tumor cells, including renal cell carcinoma. However, the mechanism of action of RUNX2 in renal cell carcinoma cells is not yet fully understood. In this study, RUNX2-negative A498 cells and strongly positive ACHN cells were selected as the study subjects. An invasion chamber assay was used to detect the invasive ability of the cells. The expression of each protein was detected by Western blotting or immunofluorescence assays. The invasive ability of A498 cells was enhanced after the expression of RUNX2 protein was upregulated, whereas ACHN cells decreased after the expression of RUNX2 protein was silenced. The expression of calcium-activated neutral protease 2 (Calpain2) and fibronectin (FN) proteins was upregulated in A498 cells overexpressing RUNX2 protein, whereas it was downregulated after the downregulation of RUNX2 protein expression in ACHN cells. It was found that Calpain2 small interfering RNA (siRNA) or calpain inhibitor calpeptin could inhibit the expression of FN in ACHN and A498 cells overexpressing RUNX2. Calpain2 siRNA or calpeptin inhibited the invasion of A498 cells overexpressing RUNX2. Similarly, in ACHN cells, Calpain2 siRNA or calpeptin inhibited cell invasion. RUNX2 upregulates FN protein expression via Calpain2, thereby mediating renal cell carcinoma invasion.

Expression and Prognostic Significance of Ferroptosis-related Proteins SLC7A11 and GPX4 in Renal Cell Carcinoma.

BACKGROUND: The ferroptosis inhibitory gene solute carrier family 7 member 11 (SLC7A11) and glutathione peroxidase 4 (GPX4) inhibit ferroptosis in carcinoma cells. However, whether SLC7A11 and GPX4 serve as an oncogene in renal cell carcinoma (RCC) remains unclear. METHODS: Immunohistochemistry (IHC) assays were performed to assess the expression of SLC7A11 and GPX4 in human RCC tissues. Clinical-pathological analysis was performed to explore the correlation between SLC7A11 and GPX4 expression. Kaplan-Meier survival analysis was performed to characterise the associations between protein expression and patient progressionfree survival (PFS). RESULTS: The upregulation of SLC7A11 and GPX4 was detected by IHC in RCC tissues compared with that in normal renal tissues. Meanwhile, the expression level of SLC7A11 and GPX4 was correlated with tumour diameter and distant metastasis (P<0.05). Kaplan-Meier survival analysis indicated that patients with high SLC7A11 and GPX4 expression levels exhibited worse PFS than those with low SLC7A11 and GPX4 expression levels (P<0.05). CONCLUSION: The upregulation of SLC7A11 and GPX4 expression was associated with poor prognosis in patients with RCC. SLC7A11 and GPX4 may serve as diagnostic and prognostic biomarkers for patients with RCC.

KLHL21/CYLD signaling confers aggressiveness in bladder cancer through inactivating NF-κB signaling.

Bladder carcinoma (BC) is one of the most commonly diagnosed malignant cancers worldwide. Kelch-like protein 21 (KLHL21) has been shown to be involved in a number of human tumors. The study aimed to investigate the effects and mechanism of KLHL21 on BC progression. We found that KLHL21 expression was significantly decreased in human BC tissues and cell lines compared with the paired normal samples, and patients with lower KLHL21 expression exhibited poorer overall survival. In vitro studies then showed that KLHL21 over-expression significantly reduced the proliferation, migration and invasion in BC cells, while KLHL21 knockdown markedly accelerated the proliferative, migratory and invasive properties of BC cells. Animal studies confirmed that KLHL21 exhibited anti-tumor function in the xenograft mouse models, as indicated by the reduced tumor growth rates, and mice with KLHL21 knockdown showed the opposite tumor growth profile. Additionally, we found that KLHL21 negatively mediated the nuclear factor-κB (NF-κB) signaling activation, as well as its down-streaming molecules involved in the biological regulation of cell survival, death and migratory processes. Mechanistically, cylindromatosis (CYLD) expression levels were significantly up-regulated in BC cells over-expressing KLHL21, but were down-regulated upon KLHL21 knockdown. We further uncovered that KLHL21 directly interacted with CYLD in BC cells. Of note, we found that KLHL21 mainly in cytoplasm could restrain CYLD degradation by prohibiting its ubiquitination in BC cells. More importantly, our in vitro experiments displayed that KLHL21-inhibited progression and NF-κB/p65 activation in BC cells were completely abolished by CYLD deletion, revealing that CYLD expression was required for KLHL21 to perform its anti-tumor function in BC. Collectively, all these findings uncovered that KLHL21/CYLD axis may be a promising therapeutic target for BC treatment.

Long Noncoding RNA MMP2-AS1 Contributes to Progression of Renal Cell Carcinoma by Modulating miR-34c-5p/MMP2 Axis.

Renal cell carcinoma (RCC) serves as a prevalent malignancy of urinary system and presents severe mortality and increasing incidence. Long noncoding RNAs (lncRNAs) have demonstrated critical roles in RCC development. Here, we were interested in the function of MMP2-AS1 during RCC progression. We observed that MP2-AS1 localized in both nucleus and cytoplasm of RCC cells using fluorescent in situ hybridization (FISH). The cell viability, proliferation, invasion, and migration of RCC cells were reduced by the depletion of MMP2-AS1. The MMP2-AS1 depletion-inhibited viability, proliferation, migration, and invasion of RCC cells were rescued by the overexpression of MMP2 in vitro. Consistently, the tumor growth of RCC cells was repressed by the depletion of MMP2-AS1 in the nude mice, while the overexpression of MMP2 could reverse this effect in vivo. Mechanically, we predicted the potential interaction of miR-34c-5p with both MMP2-AS1 and MMP2. The treatment of miR-34c-5p mimic reduced the luciferase activity of MMP2-AS1 and MMP2 3'UTR. The depletion of MMP2-AS1 enhanced miR-34c-5p expression and the expression of MMP2 was inhibited by miR-34c-5p in RCC cells. The protein levels of MMP2 were downregulated by MMP2-AS1 knockdown, while the inhibitor of miR-34c-5p rescued the expression of MMP2 in the cells. The treatment of miR-34c-5p mimic attenuated the cell viability, proliferation, invasion, and migration of RCC cells, in which MMP2 overexpression restored the phenotypes. MMP2-AS1 depletion-attenuated viability, proliferation, migration, and invasion of RCC cells were reversed by miR-34c-5p inhibitor. We concluded that MMP2-AS1 contributed to progression of renal cell carcinoma by modulating miR-34c-5p/MMP2 axis.

CircSMARCA5 Facilitates the Progression of Prostate Cancer Through miR-432/PDCD10 Axis.

Background: Circular RNAs (circRNAs) have been reported to be implicated in the pathogenesis of prostate cancer (PCa). Herein, the authors explore the role and molecular mechanism of circRNA SWI/SNF-related, matrix-associated, actin-dependent regulator of chromatin, subfamily a, member 5 (circSMARCA5) in PCa. Materials and Methods: The levels of circSMARCA5, SMARCA5, miR-432, and programmed cell death 10 (PDCD10) were determined by quantitative real-time polymerase chain reaction (qRT-PCR). The circular structure and stability of circSMARCA5 were validated by qRT-PCR using Oligo dT primer, transcriptional inhibitor actinomycin D, or RNase R treatment, respectively. Cell proliferation, migration, invasion, epithelial/mesenchymal transition (EMT), and glycolysis were detected by 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT), transwell migration and invasion assays, Western blot assay, and Glucose or Lactate Detection Kit, respectively. The target relationship between miR-432 and circSMARCA5 or PDCD10 was validated by dual-luciferase reporter assay and RNA immunoprecipitation (RIP) assay. Western blot was performed to detect the protein expression of PDCD10 in PCa cells. Results: CircSMARCA5 was aberrantly upregulated, and was a circular and stable RNA in PCa cells. CircSMARCA5 accelerated the proliferation, metastasis, and glycolysis of PCa cells. MiR-432 was a direct target of circSMARCA5, and circSMARCA5 accelerated the development of PCa through miR-432 in PCa cells. PDCD10 was a direct target of miR-432, and PDCD10 addition reversed the inhibitory effects of miR-432 accumulation on the proliferation, metastasis, and glycolysis of PCa cells. CircSMARCA5 upregulated the expression of PDCD10 through sponging miR-432 in PCa cells. Conclusion: CircSMARCA5 deteriorated PCa through the miR-432/PDCD10 axis. CircSMARCA5/miR-432/PDCD10 axis might be an underlying therapeutic target for PCa treatment.

Clinical analysis of everolimus in the treatment of metastatic renal cell carcinoma.

BACKGROUND: Renal cell carcinoma (RCC) is the most common type of kidney cancer, and accounts for approximately 3% of all malignancies. Metastatic RCC (mRCC) is not sensitive to traditional radiotherapy and chemotherapy, therefore targeted therapy has become an important treatment option. In this study, the second-line targeted drug everolimus (Afinitor), a mammalian target of rapamycin (mTOR) inhibitor, was investigated for its clinical efficacy and adverse events in mRCC after failure of first-line targeted therapy, such as sorafenib, sunitinib or pazopanib. METHODS: A total of 21 patients with mRCC who had been treated with surgery or other therapies such as tyrosine kinase inhibitors (TKIs) were given oral everolimus (10 mg/day) until disease progression. Clinical efficacy was evaluated using the Response Evaluation Criteria in Solid Tumors (RECIST) 2 months after therapy, including complete response (CR), partial response (PR), stable disease (SD), and progressive disease (PD). The adverse events were observed, and timely treatment was provided. RESULTS: Everolimus extended progression-free survival (PFS) in mRCC patients from 4 to 8 months (median 6.3 months). There were 3 patients with PR, 12 with SD, and 6 with PD, and the disease control rate (DCR) was 15/21 (71.4%). Common adverse events included stomatitis, rash, and pneumonitis. CONCLUSIONS: This study provides further support that everolimus is still an important option in mRCC treatment after failure of first-line targeted therapy. However, clinical studies are still needed to further improve its therapeutic efficacy.

Long noncoding RNA DANCR contributes to docetaxel resistance in prostate cancer through targeting the miR-34a-5p/JAG1 pathway.

Background: Chemotherapy is one of the available options for prostate cancer (PC). However, the acquisition of chemoresistance has become a major cause of chemotherapy failure. The long noncoding RNA DANCR is demonstrated to serve as an oncogene in various human cancers, including PC. However, the potential role of DANCR in docetaxel (DTX) resistance of PC and its underlying mechanism remains unclear. Methods: The abundance of DANCR, miR-34a-5p, and JAG1 mRNA was examined by quantitative reverse transcription PCR. The Cell Counting Kit-8 (CCK8) was used to determine the 50% inhibitory concentration value. Cell viability was evaluated by CCK8 and colony-formation assays. Transwells were utilized to analyze cell migration and invasion ability. The protein levels of LRP, P-gp, MRP1, and JAG1 were measured by Western blot assay. The target relationship between DANCR and miR-34a-5p, as well as miR-34a-5p and JAG1, was demonstrated by dual-luciferase, RNA immunoprecipitation, and RNA pull-down analysis. Tumor xenograft was undertaken to confirm the effect of DANCR on DTX resistance in PC. Results: DANCR and JAG1 were significantly upregulated, but miR-34a-5p was downregulated in DTX-resistant PC. Silencing of DANCR improved the DTX efficacy in DTX-resistant PC cells. DANCR served as a competing endogenous RNA of miR-34a-5p, leading to the derepression of miR-34a-5p target JAG1, which eventually triggered the resistance to DTX in DTX-tolerated PC. Conclusion: The DANCR/miR-34a-5p axis enhanced DTX resistance of PC via targeting JAG1, providing a novel insight to improve chemotherapy for PC.