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

Molecular Biology Reports - 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...     

FBXL6 depletion restrains clear cell renal cell carcinoma progression

BackgroundF-box proteins play important roles in cell cycle and tumorigenesis. However, its prognostic value and molecular function in clear cell renal cell carcinoma (ccRCC) remain unclear. In this study, we established a survival model to evaluate the prognosis of patients with ccRCC using the F-box gene signature and investigated the function of FBXL6 in ccRCC.MethodsComprehensive bioinformatics analyses were used to identify differentially expressed F-box and hub genes associated with ccRCC carcinogenesis. Based on the F-box gene signature, we constructed a risk model and nomogram to predict the overall survival (OS) of patients with ccRCC and assist clinicians in decision-making. Finally, we verified the function and underlying molecular mechanisms of FBXL6 in ccRCC using CCK-8 and EdU assays, flow cytometry, and subcutaneous xenografts.ResultsA risk model based on FBXO39, FBXL6, FBXO1, and FBXL16 was developed. In addition, we drew a nomogram based on the risk score and clinical features to assess the prognosis of patients with ccRCC. Subsequently, we identified FBXL6 as an independent prognostic marker that was highly expressed in ccRCC cell lines. In vivo and in vitro assays revealed that the depletion of FBXL6 inhibited cell proliferation and induced apoptosis. We also demonstrated that SP1 regulated the expression of FBXL6.ConclusionsFBXL6 was first identified as a diagnostic and prognostic marker in patients with ccRCC. Loss of FBXL6 attenuates proliferation and induces apoptosis in ccRCC cells. SP1 was also found to regulate the expression of FBXL6.     

Interplay between pVHL and mTORC1 pathways in clear-cell renal cell carcinoma

mTOR complex 1 (mTORC1) is implicated in cell growth control and is extensively regulated. We previously reported that in response to hypoxia, mTORC1 is inhibited by the protein regulated in development and DNA damage response 1 (REDD1). REDD1 is upregulated by hypoxia-inducible factor (HIF)-1, and forced REDD1 expression is sufficient to inhibit mTORC1. REDD1-induced mTORC1 inhibition is dependent on a protein complex formed by the tuberous sclerosis complex (TSC)1 and 2 (TSC2) proteins. In clear-cell renal cell carcinoma (ccRCC), the von Hippel-Lindau (VHL) gene is frequently inactivated leading to constitutive activation of HIF-2 and/or HIF-1, which may be expected to upregulate REDD1 and inhibit mTORC1. However, mTORC1 is frequently activated in ccRCC, and mTORC1 inhibitors are effective against this tumor type; a paradox herein examined. REDD1 was upregulated in VHL-deficient ccRCC by in silico microarray analyses, as well as by quantitative real-time PCR, Western blot, and immunohistochemistry. Vhl disruption in a mouse model was sufficient to induce Redd1. Using ccRCC-derived cell lines, we show that REDD1 upregulation in tumors is VHL dependent and that both HIF-1 and HIF-2 are, in a cell-type-dependent manner, recruited to, and essential for, REDD1 induction. Interestingly, whereas mTORC1 is responsive to REDD1 in some tumors, strategies have evolved in others, such as mutations disrupting TSC1, to subvert mTORC1 inhibition by REDD1. Sequencing analyses of 77 ccRCCs for mutations in TSC1, TSC2, and REDD1, using PTEN as a reference, implicate the TSC1 gene, and possibly REDD1, as tumor suppressors in sporadic ccRCC. Understanding how ccRCCs become refractory to REDD1-induced mTORC1 inhibition should shed light into the development of ccRCC and may aid in patient selection for molecular-targeted therapies.     

Familial non-syndromic clear cell renal cell carcinoma

The diagnosis of familial non-syndromic clear cell renal cell carcinoma is one of exclusion. In families presenting with clear cell RCC a germline VHL mutation and a constitutional translocation of chromosome 3 must be excluded before familial non-syndromic clear cell RCC can be diagnosed. Large familial non-syndromic clear cell RCC kindreds are uncommon and a predisposing gene has not been identified. However inheritance is autosomal dominant in most cases and age at onset is earlier than in sporadic cases. Recognition and appropriate screening of familial non-syndromic clear cell RCC cases will reduce morbidity and mortality. Large scale collaborative linkage studies may provide a basis for the identification of familial non-syndromic clear cell RCC susceptibility gene(s).     

OTUD7B stabilizes estrogen receptor α and promotes breast cancer cell proliferation

Breast cancer is the most common malignancy in women worldwide. Estrogen receptor α (ERα) is expressed in ∼70% of breast cancer cases and promotes estrogen-dependent cancer progression. In the present study, we identified OTU domain-containing 7B (OTUD7B), a deubiquitylase belonging to A20 subgroup of ovarian tumor protein superfamily, as a bona fide deubiquitylase of ERα in breast cancer. OTUD7B expression was found to be positively correlated with ERα in breast cancer and associated with poor prognosis. OTUD7B could interact with, deubiquitylate, and stabilize ERα in a deubiquitylation activity-dependent manner. Depletion of OTUD7B decreased ERα protein level, the expression of ERα target genes, and the activity of estrogen response element in breast cancer cells. In addition, OTUD7B depletion significantly decreased ERα-positive breast cancer cell proliferation and migration. Finally, overexpression of ERα could rescue the suppressive effect induced by OTUD7B depletion, suggesting that the ERα status was essential to the function of OTUD7B in breast carcinogenesis. In conclusion, our study revealed an interesting post-translational mechanism between ERα and OTUD7B in ERα-positive breast cancer. Targeting the OTUD7B–ERα complex may prove to be a potential approach to treat patients with ERα-positive breast cancer.Subject terms: Breast cancer, Cell growth     

Orai1 and STIM1 are critical for cell migration and proliferation of clear cell renal cell carcinoma

The intracellular Ca²⁺ regulation has been implicated in tumorigenesis and tumor progression. Notably, store-operated Ca²⁺ entry (SOCE) is a major Ca²⁺ entry mechanism in non-excitable cells, being involved in cell proliferation and migration in several types of cancer. However, the expression and biological role of SOCE have not been investigated in clear cell renal cell carcinoma (ccRCC). Here, we demonstrate that Orai1 and STIM1, not Orai3, are crucial components of SOCE in the progression of ccRCC. The expression levels of Orai1 in tumor tissues were significantly higher than those in the adjacent normal parenchymal tissues. In addition, native SOCE was blunted by inhibiting SOCE or by silencing Orai1 and STIM1. Pharmacological blockade or knockdown of Orai1 or STIM1 also significantly inhibited RCC cell migration and proliferative capability. Taken together, Orai1 is highly expressed in ccRCC tissues illuminating that Orai1-mediated SOCE may play an important role in ccRCC development. Indeed, Orai1 and STIM1 constitute a native SOCE pathway in ccRCC by promoting cell proliferation and migration.     

KIF20B promotes the progression of clear cell renal cell carcinoma by stimulating cell proliferation

Renal cell carcinoma (RCC) is a common urinary system cancer with high morbidity and mortality rate. Clear cell renal cell carcinoma (ccRCC) is a highly aggressive and common type of RCC. More and effective therapeutic targets are badly needed for the treatment of ccRCC. Kinesin family protein (KIF)20B, also named M-phase phosphoprotein 1, was reported as a microtubule-associated, plus-end-directed kinesin. KIF20B was involved in multiple cellular processes such as cytokinesis. Multiple studies indicated the oncogenic role for KIF20B in several types of tumors, including breast cancer and bladder cancer. However, the possible role of KIF20B in the progression of renal carcinoma is still unknown. Herein, our study demonstrated that KIF20B was relatively highly expressed in ccRCC tissues. In addition, KIF20B was inversely related to the clinical features including tumor size and T stage. We further found that inhibition of the KIF20B expression by a specific short hairpin RNA obviously reduces proliferation of ccRCC cells both in vitro and in vivo. Our study reveals the involvement of KIF20B in ccRCC progression. Generally, KIF20B is a promising novel therapeutic for the treatment of clear cell RCC.     

Calcium-activated nucleotidase 1 silencing inhibits proliferation,migration, and invasion in human clear cell renal cell carcinoma

Calcium-activated nucleotidase 1 (CANT1, belongs to the apyrase family, is widely expressed in various organs. However, the biological function of CANT1 remains poorly explored. In this study, we aimed to investigate the expression profile and functions of CANT1 in clear cell renal cell carcinoma (ccRCC). Our data show that the protein level of CANT1 was significantly higher in tumor tissues than in adjacent normal tissues. CANT1 silencing suppressed cell proliferation, migration, and invasion obviously in 769-P and 786-O cells, arrested cell cycle in S phase and promoted apoptosis in 769-P cells. In conclusion, the present study shows the different expression mode of CANT1 in human ccRCC tumor tissue and adjacent normal tissue, denotes the function of CANT1 in ccRCC cells and provides potential molecular mechanisms and pathways of CANT1 antitumor function in ccRCC.     

VHL inactivation in sporadic clear cell renal carcinoma

Clear cell renal carcinoma (CCRC) accounts for 75% of all renal cancer cases. The majority of CCRCs displays inactivation of the VHL suppressor gene as a result of mutations, allelic deletions, and/or methylation. Data on the effect of VHL inactivation on the prognosis in CCRC are discrepant. Comprehensive molecular genetic analysis of VHL was performed for 64 CCRCs: mutations were identified by single strand conformation polymorphism analysis and subsequent sequencing, a loss of heterozygosity was studied with two STR markers, and methylation was assessed by methylation-sensitive PCR. In total, 17 somatic mutations, including 12 new ones, were found in VHL. Allelic deletions of VHL were detected in 31.6% of cases; methylation was observed in 7.8% of cases. In total, VHL was inactivated in 46.9% of CCRC cases and in 51.7% of patients with CCRC stage I. The frequencies of mutations, loss of heterozygosity, and methylation did not correlate with clinical features of CCRC or pathological characteristics of the tumor. Studies of the molecular genetics alterations of VHL are thought to facilitate the identification of diagnostic and prognostic markers of renal cancer, e.g., the selection of an optimal panel of methylated suppressor genes.     

Pathology and the evolutionary dynamics of clear cell renal cell carcinoma

Cancer is an ecosystem whose intrinsic mechanisms do not show up under the microscope of pathologists. However, the information provided by pathologists is absolutely necessary for the correct implementation of personalized treatments. This short paper seeks to analyze this apparent paradox, i.e. static snapshots for making crucial decisions in essentially dynamic diseases, taking clear cell renal cell carcinoma as a paradigmatic example of tumor variability. We seek to call the attention of pathologists and other cancer-related medical specialists to extend knowledge of the evolutionary features of the disease to help obtain a better understanding of why cancer behaves as it does.     

Abnormal cholesterol metabolism in renal clear cell carcinoma

The clear cell form of renal cell carcinoma is known to derive its histologic appearance from accumulations of glycogen and lipid. We have found that the most consistently stored lipid form is cholesteryl ester. Clear cell cancer tissue contained 8-fold more total cholesterol and 35-fold more esterified cholesterol than found in normal kidney. Cholesteryl ester appeared to be formed intracellularly since it was not membrane-bound and since oleate was the predominant form, as opposed to linoleate in lipoprotein cholesteryl esters. The cholesterol in clear cell tumors did not appear to be a result of excessive synthesis from acetate since HMG-CoA reductase (EC 1.1.1.34) activity was lower in cancer tissue than in normal kidney (2.9 +/- 0.8 vs. 7.2 +/- 1.2 pmol/mg of protein per min). In contrast, intracellular activity of fatty acyl-coenzyme A:cholesterol acyl transferase (ACAT, EC 2.3.1.26) was higher in tumor tissue than in normal kidney (2405 +/- 546 vs. 1326 +/- 301 pmol/mg of protein per 20 min) while cytosolic cholesteryl ester hydrolase activity appeared normal. Cholesteryl ester storage in clear cell renal cancer may be a result of a primary abnormality in ACAT activity or it may be a result of reduced release of free cholesterol (relative to cell content) with a secondary elevation in ACAT activity.     

FKBP51 promotes invasion and migration by increasing the autophagic degradation of TIMP3 in clear cell renal cell carcinoma

The occurrence of metastasis is a serious risk for renal cell carcinoma (RCC) patients. In order to develop novel therapeutic approaches to control the progression of metastatic RCC, it is of urgent need to understand the molecular mechanisms underlying RCC metastasis and identify prognostic markers of metastatic risk. Matrix metalloproteinases (MMPs) and tissue inhibitors of metalloproteinases (TIMPs) have been known to be closely associated with extracellular matrix (ECM) turnover, which plays a highly active role in tumor metastasis. Recent studies have shown that immunophilin FK-506-binding protein 51 (FKBP51) may be important for the regulation of ECM function, and exert effects on the invasion and migration of tumor cells. However, the mechanisms underlying these activities remain unclear. The present study detected the role of FKBP51 in clear cell renal cell carcinoma (ccRCC), the most common subtype of RCC, and found that FKBP51 significantly promotes ccRCC invasion and migration by binding with the TIMP3, connecting TIMP3 with Beclin1 complex and increasing autophagic degradation of TIMP3. Given the important roles that TIMPs/MMPs play in ECM regulation and remodeling, our findings will provide new perspective for future investigation of the regulation of metastasis of kidney cancer and other types of cancer.Subject terms: Renal cell carcinoma, Extracellular matrix     

Integrated glycoproteomic characterization of clear cell renal cell carcinoma

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Semaphorin 6D as an independent predictor for better prognosis in clear cell renal cell carcinoma

IntroductionClear cell renal cell carcinoma (ccRCC) is the most common type of RCC and is associated with poor survival. However, the mechanisms underlying its development have not been thoroughly investigated. Semaphorin 6D (SEMA6D) is differentially expressed in various cancers, including lung adenocarcinoma and colorectal cancer. However, the role and mechanism of SEMA6D in ccRCC remain unexplored.Materials and methodsWe obtained 25 pairs of ccRCC tissue samples and 57 urine samples from patients with ccRCC and 52 urine samples from healthy volunteers. We performed RNA sequencing and compared the results with data from The Cancer Genome Atlas database to identify our gene of interest, SEMA6D. To verify the differential expression of SEMA6D, we used real-time quantitative polymerase chain reaction, immunohistochemistry, and enzyme-linked immunosorbent assay. Finally, we conducted in vitro proliferation, migration and invasion experiments.ResultsSEMA6D expression was significantly lower in ccRCC tissue compared to that in normal tissue. Comparative analysis of our results with data from online databases revealed that the expression level of SEMA6D in ccRCC tissue correlated with the clinical stage and pathological grade of ccRCC. Furthermore, higher SEMA6D expression was associated with improved quality of life of patients with ccRCC. In addition, the diagnostic value of SEMA6D was confirmed using data from two Gene Expression Omnibus ccRCC databases. The results showed that SEMA6D can be used as a predictor for ccRCC diagnosis, with an area under the curve of 0.9642.ConclusionSEMA6D may serve as a diagnostic and prognostic biomarker for ccRCC.     

MicroRNA-187, down-regulated in clear cell renal cell carcinoma and associated with lower survival,inhibits cell growth and migration though targeting B7-H3

Aberrantly expressed microRNAs (miRNAs) are frequently associated with the aggressive malignant behavior of human cancers, including clear cell renal cell carcinoma (ccRCC). Based on the preliminary deep sequencing data, we hypothesized that miR-187 may play an important role in ccRCC development. In this study, we found that miR-187 was down-regulated in both tumor tissue and plasma of ccRCC patients. Lower miR-187 expression levels were associated with higher tumor grade and stage. All patients with high miR-187 expression survived 5 years, while with low miR-187 expression, only 42% survived. Suppressed in vitro proliferation, inhibited in vivo tumor growth, and decreased motility were observed in cells treated with the miR-187 expression vector. Further studies showed that B7 homolog 3 (B7-H3) is a direct target of miR-187. Over-expression of miR-187 decreased B7-H3 mRNA level and repressed B7-H3-3′-UTR reporter activity. Knockdown of B7-H3 using siRNA resulted in similar phenotype changes as that observed for overexpression of miR-187. Our data suggest that miR-187 is emerging as a novel player in the disease state of ccRCC. miR-187 plays a tumor suppressor role in ccRCC.     

Single-cell analysis reveals metastatic cell heterogeneity in clear cell renal cell carcinoma

Renal cell carcinoma (RCC) is one of the leading causes of cancer-related death worldwide. Tumour metastasis and heterogeneity lead to poor survival outcomes and drug resistance in patients with metastatic RCC (mRCC). In this study, we aimed to assess intratumoural heterogeneity (ITH) in mRCC cells by performing a combined analysis of bulk data and single-cell RNA-sequencing data, and develop novel biomarkers for prognosis prediction on the basis of the potential molecular mechanisms underlying tumorigenesis. Eligible single-cell cohorts related to mRCC were acquired using the Gene Expression Omnibus (GEO) dataset to identify potential mRCC subpopulations. We then performed gene set variation analysis to understand the differential function in primary RCC and mRCC samples. Subsequently, we applied weighted correlation network analysis to identify coexpressing gene modules that were related to the external trait of metastasis. Protein-protein interactions were used to screen hub subpopulation-difference (sub-dif) markers (ACTG1, IL6, CASP3, ACTB and RAP1B) that might be involved in the regulation of RCC metastasis and progression. Cox regression analysis revealed that ACTG1 was a protective factor (HR < 1), whereas the other four genes (IL6, CASP3, ACTB and RAP1B) were risk factors (HR > 1). Kaplan-Meier survival analysis suggested the potential prognostic value of these sub-dif markers. The expression of sub-dif markers in mRCC was further evaluated in clinical samples by immunohistochemistry (IHC). Additionally, the genetic features of sub-dif marker expression patterns, such as genetic variation profiles, correlations with tumour-infiltrating lymphocytes (TILs), and targeted signalling pathway activities, were assessed in bulk RNA-seq datasets. In conclusion, we established novel subpopulation markers as key prognostic factors affecting EMT-related signalling pathway activation in mRCC, which could facilitate the implementation of a treatment for mRCC patients.