cyclin-d1 and Carcinoma--Renal-Cell

Multiple primary malignant tumors (MPMTs), usually associated with worse malignant behavior and prognosis comparing to a single primary tumor, and have recently been found to have an increasing incidence globally. However, the pathogenesis of MPMTs remains to be clarified. Here, we report a unique case of the coexistence of malignant melanoma (MM), papillary thyroid carcinoma (PTC), and clear-cell renal cell carcinoma (ccRCC) along with our perceptions on its pathogenesis.. The case reported is of a 59-year-old male patient with unilateral nasal obstruction as well as a renal occupying lesion. Positron emission tomography-computed tomography (PET-CT) revealed a palpable mass of 32 × 30 mm on the posterior and left walls of the nasopharynx. In addition, an isodense nodule was observed in the right superior renal pole, approximately 25 mm in diameter, as well as a slightly hypodense shadow in the right leaf of the thyroid, approximately 13 mm in diameter. Nasal endoscopy and magnetic resonance imaging (MRI) confirmed the existence of a nasopharyngeal neoplasm. Afterward, biopsies of the nasopharyngeal neoplasm, thyroid gland and kidney were performed, and the patient was diagnosed with MM, PTC, and ccRCC according to the pathological and immunohistochemical results. Moreover, mutation of BRAF. This is the first reported case of a patient with the co-existence of MM, PTC and ccRCC undergoing chemotherapy with a favorable prognosis. Herein, we suggest that such a combination may be non-random, as for mutation of BRAF

Topics: Carcinoma; Carcinoma, Renal Cell; Cyclin D1; Humans; Kidney Neoplasms; Male; Melanoma, Cutaneous Malignant; Middle Aged; Mutation; Nasopharyngeal Neoplasms; Neoplasms, Multiple Primary; Positron Emission Tomography Computed Tomography; Proto-Oncogene Proteins B-raf; Thyroid Cancer, Papillary; Thyroid Neoplasms

Previous studies indicated that cyclin D1 shown the potential as a tumor biomarker. However, the prognostic value of cyclin D1 in renal cell carcinoma (RCC) remains controversial. This study investigated the correlation of cyclin D1 expression with the prognostic and clinicopathological features in RCC patients. We systematically searched the database of PubMed, Embase, Cochrane, and Web of Science updated on November 26, 2017. Eighteen studies with 2282 patients satisfied the inclusion criteria. Results demonstrated that cyclin D1 overexpression in RCC showed significant favorable prognostic impact on disease-free survival (DFS) (HR 0.57, 95% CI: 0.43-0.74) and disease-specific survival (DSS) (HR 0.59, 95% CI 0.41-0.85) without significant heterogeneity. In subgroup of clear cell RCC, the prognostic effect on DFS was robust and the pooled HR was 0.39 (95% CI: 0.27-0.57). However, no association between overall survival (OS) and cyclin D1 expression was observed. Stratified analysis in DFS studies by sample size, staining patterns race and metastasis status showed similar results. Otherwise, cyclin D1 overexpression predicted a reduced prevalence of high TNM stage (T3 + T4) (OR 0.63, 95% CI: 0.40-0.99), high-grade tumor (G3 + G4) (OR 0.51, 95% CI: 0.31-0.81) and large tumor size (OR 0.35, 95% CI: 0.19-0.62). Our meta-analysis indicated that cyclin D1 overexpression could predict the favorable prognosis in patients with RCC.

Topics: Biomarkers, Tumor; Carcinoma, Renal Cell; Cyclin D1; Humans; Kidney Neoplasms; Prognosis

Bardoxolone methyl, a novel synthetic triterpenoid and antioxidant inflammation modulator, potently induces Nrf2 and inhibits NF-κB and Janus-activated kinase/STAT signaling. This first-in-human phase I clinical trial aimed to determine the dose-limiting toxicities (DLT), maximum tolerated dose (MTD), and appropriate dose for phase II studies; characterize pharmacokinetic and pharmacodynamic parameters; and assess antitumor activity.. Bardoxolone methyl was administered orally once daily for 21 days of a 28-day cycle. An accelerated titration design was employed until a grade 2-related adverse event occurred. A standard 3 + 3 dose escalation was then employed until the MTD was reached. Single dose and steady-state plasma pharmacokinetics of the drug were characterized. Assessment of Nrf2 activation was examined in peripheral blood mononuclear cells (PBMC) by measuring NAD(P)H:quinone oxidoreductase (NQO1) mRNA levels. Immunohistochemical assessment of markers of inflammation, cell cycle, and apoptosis was carried out on tumor biopsies.. The DLTs were grade 3 reversible liver transaminase elevations. The MTD was established as 900 mg/d. A complete tumor response occurred in a mantle cell lymphoma patient, and a partial response was observed in an anaplastic thyroid carcinoma patient. NQO1 mRNA levels increased in PBMCs, and NF-κB and cyclin D1 levels decreased in tumor biopsies. Estimated glomerular filtration rate (eGFR) was also increased.. Bardoxolone methyl was well tolerated with an MTD of 900 mg/d. The increase in eGFR suggests that bardoxolone methyl might be beneficial in chronic kidney disease. Objective tumor responses and pharmacodynamic effects were observed, supporting continued development of other synthetic triterpenoids in cancer.

Topics: Adult; Aged; Aged, 80 and over; Antineoplastic Agents; Carcinoma, Renal Cell; Colorectal Neoplasms; Cyclin D1; Dose-Response Relationship, Drug; Drug Administration Schedule; Female; Humans; Janus Kinases; Kidney Neoplasms; Lymphoma; Male; Maximum Tolerated Dose; Melanoma; Middle Aged; NAD(P)H Dehydrogenase (Quinone); Neoplasms; NF-E2-Related Factor 2; NF-kappa B; Oleanolic Acid; RNA, Messenger; STAT Transcription Factors; Thyroid Neoplasms; Young Adult

Renal cell carcinoma (RCC) remains one of the most lethal urinary tumors in East Asia despite great advancements in treatment strategies in recent years. Ribosomal protein S20 (RPS20) is considered a new oncogene; however, little information is available on its expression, regulation and biological function in patients with RCC. In the present study, 43 pairs of human RCC and neighboring normal renal tissues were examined for protein expression and immunohistochemistry examination of RPS20. Lentiviral transduction was also employed to create RPS20 knockdown cell lines for downstream cellular experiments. MTT, flow cytometry, wound healing, colony formation and invasion assays were used to examine how RPS20 affected kidney renal clear cell carcinoma (KIRC) cell behavior. Western blotting was used to detect cycle‑related proteins (CDK4 and cyclin D1), Wnt‑related proteins (N‑cadherin and E‑cadherin) and signaling proteins [phosphorylated (p)‑AKT and p‑ERK]. The functions of RPS20

Topics: Cadherins; Carcinoma, Renal Cell; Cyclin D1; Humans; Kidney Neoplasms

To explore the role of miR-744-5p/CCND1 axis in clear-cell renal cell carcinoma (ccRCC).. We examined the expression levels of miR-744-5p in 65 pairs of ccRCC and adjacent tissue specimens and in 5 ccRCC cell lines and human renal tubular epithelial (HK2) cells using qRT-PCR. The ccRCC cell lines 786-O and OSRC2 were transfected with miR-744-5p mimic, CCND1 mimic, or their negative control mimics, and the changes in cell proliferation, migration, and invasion were evaluated with CCK-8, wound healing, and Transwell assays. The downstream target molecules of miR-744-5p were predicted by bioinformatics analysis, and the expression level of CCND1 in ccRCC cells was verified by qRT-PCR and Western blotting. The relationship between miR-744-5p and CCND1 was further validated by dual luciferase reporter assay, and the role of the miR-744-5p/CCND1 axis in ccRCC was explored by rescue experiments.. MiR-744-5p was significantly downregulated in ccRCC tissues and cell lines (all. MiR-744-5p inhibits the malignant phenotype of ccRCC cells by targeting CCND1, and the miR-744-5p/CCND1 axis may be a novel target for diagnosis and treatment of ccRCC.

Topics: Carcinoma, Renal Cell; Cell Line, Tumor; Cell Movement; Cell Proliferation; Cyclin D1; Humans; Kidney Neoplasms; MicroRNAs

Large-scale human genetic data

Topics: Alleles; Basic Helix-Loop-Helix Transcription Factors; Carcinogenesis; Carcinoma, Renal Cell; Cyclin D1; Gene Expression Regulation, Neoplastic; Humans; Kidney; Kidney Neoplasms; Mutation; PAX8 Transcription Factor; Proto-Oncogene Proteins c-myc; Signal Transduction; Von Hippel-Lindau Tumor Suppressor Protein

6-Gingerol, a major biochemical and pharmacological active ingredient of ginger, has shown anti-inflammatory and antitumor activities against various cancers. Searching for natural products with fewer side effects for developing adjunctive therapeutic options is necessary.. The effects of 6-gingerol on proliferation, colony formation, and cell cycle in RCC cells were detected by a 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay, colony formation assay, and propidium iodide (PI) staining, respectively. Western blotting, an immunofluorescence assay, and immunohistochemical staining were performed to assess the expression of relevant proteins. A subcutaneous tumor model was set up to investigate the 6-gingerol effects on tumor growth in vivo, and the pharmacokinetics of 6-gingerol in mice were detected by LC/MS assays.. 6-Gingerol treatment exerted time- and dose-dependent inhibition of the growth and colony formation of ACHN, 786-O, and 769-P cells, leading to a concomitant induction of cell-cycle G1-phase arrest and decrease in Ki-67 expression in the cell nucleus. Western-blotting results showed that 6-gingerol reduces phosphorylation of protein kinase B (AKT) Ser 473, cyclin-dependent kinases (CDK4), and cyclin D1 and, meanwhile, increases glycogen synthase kinase (GSK 3β) protein amount. Furthermore, the efficacy of 6-gingerol was demonstrated in an in vivo murine model of 786-O.. The above results indicate that 6-gingerol can induce cell-cycle arrest and cell-growth inhibition through the AKT-GSK 3β-cyclin D1 signaling pathway in vitro and in vivo, suggesting that 6-gingerol should be useful for renal-cell carcinoma treatment.

Topics: Animals; beta Catenin; Carcinoma, Renal Cell; Catechols; Cell Line, Tumor; Cell Proliferation; Cyclin D1; Fatty Alcohols; G1 Phase Cell Cycle Checkpoints; Glycogen Synthase Kinase 3 beta; Humans; Kidney Neoplasms; Mice; Mice, Inbred BALB C; Mice, Nude; Phosphorylation; Proto-Oncogene Proteins c-akt; Signal Transduction

Currently, there are limited effective treatment options for renal cell carcinoma (RCC), due to its poor responses to conventional therapies. Instead of using extrinsic anti-cancer drugs, cancer cell-intrinsic reactive oxygen species (ROS) can be a weapon of RCC treatment. In the present study, we found that the phytochemical thymoquinone (TQ), a bioactive natural product obtained from the black cumin seeds of Nigella sativa, generates intracellular ROS in human renal cancer Caki-1 cells. Treatment of Caki-1 cells with high concentration of TQ up-regulated pro-apoptotic p53 and Bax expression, while downregulated anti-apoptotic Bcl-2 and Bcl-xl expression. Simultaneously, TQ suppressed the pro-oncogenic JAK2/STAT3 pathway, resulting in decreased expression of Bcl-2, Bcl-xl, cyclin D1, cyclin D2, and survivin. Thus, TQ can integrate between apoptosis and the pro-survival JAK2/STAT3 pathway through the Bcl family members, collectively magnifying Caki-1 cell apoptosis. However, treatment with the ROS scavenger N-acetyl cysteine significantly blocked TQ-induced apoptosis as well as incorporated signaling pathways, supporting that its pro-oxidant property is crucial for Caki-1 cell apoptosis. Moreover, TQ reduced the tumor xenograft growth of Caki-1 cells in nude mice. Taken together, these data suggest that TQ is a prominent anti-cancer drug to treat human RCC by enhancing apoptosis through its pro-oxidant nature.

Topics: Animals; Antineoplastic Agents; Apoptosis; bcl-X Protein; Benzoquinones; Carcinoma, Renal Cell; Cell Line, Tumor; Cell Proliferation; Cuminum; Cyclin D1; Cyclin D2; Humans; Janus Kinase 2; Kidney Neoplasms; Male; Mice; Mice, Nude; Phytochemicals; Proto-Oncogene Proteins c-bcl-2; Reactive Oxygen Species; Seeds; Signal Transduction; STAT3 Transcription Factor; Survivin; Xenograft Model Antitumor Assays

Kidney renal clear cell carcinoma (KIRC) is one of the most common lethal cancers in the human urogenital system. As members of the Homeobox (HOX) family, Homeobox-A (HOXA) cluster genes have been reported to be involved in the development of many cancer types. However, the expression and clinical significance of HOXA genes in KIRC remain largely unknown.. In this study, we comprehensively analyzed the mRNA expression and prognostic values of HOXA genes in KIRC using The Cancer Genome Atlas (TCGA) analysis databases online. Colony formation assay, flow cytometry and Western blot were used to detect cell proliferation, apoptosis, cell cycle, and protein level of the indicated gene.. We found that the HOXA genes were differentially expressed in KIRC tissues when compared with normal tissues. The expression of HOXA4 and HOXA13 were significantly up-regulated, while HOXA7 and HOXA11 were down-regulated in KIRC. High mRNA levels of HOXA2, HOXA3 and HOXA13, and low level of HOXA7 predicted poor overall survival (OS) of KIRC patients. High mRNA level of HOXA13 further indicated a poor disease-free survival (DFS) of KIRC patients. Functionally, knockdown of HOXA13 significantly suppressed cell proliferation of KIRC in vitro, increased the protein level of p53 and decreased the protein level of cyclin D1 in KIRC cells. Over-expression of HOXA13 had the opposite effects on KIRC cells.. Collectively, our findings suggest that HOXA13 functions as a novel oncogene in KIRC and may be a potential biomarker for this malignancy.

Topics: Biomarkers, Tumor; Carcinoma, Renal Cell; Cell Line, Tumor; Cell Proliferation; Cyclin D1; Databases, Genetic; Down-Regulation; Gene Knockdown Techniques; Homeodomain Proteins; Humans; Kidney Neoplasms; Multigene Family; Oncogenes; RNA, Messenger; Transcription Factors; Transcription, Genetic; Tumor Suppressor Protein p53; Up-Regulation

The molecular heterogeneity of renal cell carcinoma (RCC) complicates the therapeutic interventions for advanced metastatic disease and thus its management remains a significant challenge. This study investigates the role of the lncRNA CDKN2B-AS1 and miR-141-3p interactions in the progression and metastasis of kidney cancer. Human renal cancer cell lines (ACHN and Caki1), normal RPTEC cells, tissue cohorts, and a series of in vitro assays and in vivo mouse model were used for this study. An overexpression of CDKN2B-AS1 was observed in RCC compared to normal samples in TCGA and our in-house SFVAMC tissue cohorts. Reciprocally, we observed reduced expression of miR-141 in RCC compared to normal in the same cohorts. CDKN2B-AS1 shares regulatory miR-141 binding sites with CCND1 and CCND2 genes. Direct interactions of CDKN2B-AS1/miR-141/Cyclin D1-D2 were confirmed by RNA immunoprecipitation and luciferase reporter assays indicating that CDKN2B-AS1/miR-141/Cyclin D1-D2 acts as a ceRNA network in RCC. Functionally, attenuation of CDKN2B-AS1 and/or overexpression of miR-141 inhibited proliferation, clonogenicity, migration/invasion, induced apoptosis in vitro and suppressed tumor growth in xenograft mouse model. Further, overexpression of CDKN2B-AS1 is positively correlated with poor overall survival of RCC patients. Expression of miR-141 also robustly discriminated malignant from non-malignant tissues and its inhibition in normal RPTEC cells induced pro-cancerous characteristics. CDKN2B-AS1 attenuation or miR-141 overexpression decreased CCND1/CCND2 expression, resulting in reduced RAC1/pPXN that are involved in migration, invasion and epithelial-mesenchymal transition. This study, for the first time, deciphered the role of CDKN2B-AS1/miR-141/Cyclin D axis in RCC and highlights this network as a promising therapeutic target for the regulation of EMT driven metastasis in RCC.

Topics: Animals; Apoptosis; Carcinogenesis; Carcinoma, Renal Cell; Cell Line, Tumor; Cell Movement; Cell Proliferation; Cyclin D; Cyclin D1; Cyclin D2; Cyclin-Dependent Kinase Inhibitor p15; Disease Progression; Epithelial-Mesenchymal Transition; Female; Gene Expression Regulation, Neoplastic; Humans; Male; Mice; Mice, Nude; MicroRNAs; Neoplasm Metastasis; RNA, Long Noncoding; Xenograft Model Antitumor Assays

Topics: Aged; Carcinoma, Renal Cell; Cell Line, Tumor; Cell Proliferation; Cyclin D1; Disease Progression; Female; Gene Expression Regulation, Neoplastic; Humans; Kidney Neoplasms; Lymphatic Metastasis; Male; MicroRNAs; Middle Aged; Phenotype; RNA, Long Noncoding; Survival Rate

The purpose of this study was to investigate the effect of microRNA-206 on the malignant progression of renal clear cell carcinoma (RCC). In addition, whether microRNA-206 could regulate ZEB2 expression and the underlying mechanisms was also explored.. Quantitative Real Time-Polymerase Chain Reaction (qRT-PCR) was performed to examine microRNA-206 level in 46 tumor tissue specimens and adjacent ones of RCC patients. Also, the relationship between microRNA-206 expression and clinical indicators of RCC was analyzed. The negative control (NC) and microRNA-206 mimics were transfected into RCC cell lines, and the transfection efficiency was verified by qRT-PCR. The effects of microRNA-206 on the proliferation and apoptosis of RCC cells were analyzed by cell counting kit-8 (CCK-8), clone formation, and flow cytometry assays. Finally, the regulation of microRNA-206 on the downstream gene ZEB2 was indicated by Western Blot and cell recovery experiments.. qRT-PCR results showed that the expression level of microRNA-206 in tumor tissue samples of RCC patients was remarkably lower than that in adjacent normal tissues, and the difference was statistically significant. Meanwhile, compared with patients with high expression of microRNA-206, the pathological stage of patients with low expression of microRNA-206 was higher, and the overall survival rate was lower. In the RCC cell lines (Caki-1 and Caki-2), the cell proliferation ability of the microRNA-206 overexpression group was remarkably weakened, while the cell apoptosis rate was oppositely enhanced when compared with the NC group. In addition, this study demonstrated that ZEB2 expression was remarkably increased in RCC cells as well as tissues and was negatively correlated with microRNA-206 expression. At the same time, microRNA-206 mimics was found remarkably reduced in the expression of proteins in ZEB2-related signaling pathway, including ZEB2, β-catenin, cyclinD1, c-Myc, MMP-2, and MMP-9. In the cell reverse experiment, the overexpression of ZEB2 was found to be able to counteract the impact of microRNA-206 mimics on RCC cell proliferation and apoptosis and thus, participated in the malignant progression of RCC.. This study revealed that microRNA-206 was remarkably associated with the pathological stage and poor prognosis of RCC patients. In addition, microRNA-206 might inhibit the malignant progression of RCC by regulating the targeted ZEB2.

Topics: Aged; Apoptosis; beta Catenin; Carcinoma, Renal Cell; Case-Control Studies; Cell Line, Tumor; Cell Proliferation; Cyclin D1; Disease Progression; Female; Gene Expression Regulation; Gene Expression Regulation, Neoplastic; Humans; Kidney Neoplasms; Male; Matrix Metalloproteinase 2; Matrix Metalloproteinase 9; MicroRNAs; Middle Aged; Proto-Oncogene Proteins c-myc; Survival Rate; Transfection; Zinc Finger E-box Binding Homeobox 2

Clear cell renal cell carcinoma (ccRCC) is one of the most common cancers worldwide. Despite intense efforts to elucidate its pathogenesis, the molecular mechanisms and genetic characteristics of this cancer remain unknown. In this study, three expression profile data sets (GSE15641, GSE16441 and GSE66270) were integrated to identify candidate genes that could elucidate functional pathways in ccRCC. Expression data from 63 ccRCC tumors and 54 normal samples were pooled and analyzed. The GSE profiles shared 379 differentially expressed genes (DEGs), including 249 upregulated genes, and 130 downregulated genes. A protein-protein interaction network (PPI) was constructed and analyzed using STRING and Cytoscape. Functional and signaling pathways of the shared DEGs with significant p values were identified. Kaplan-Meier plots of integrated expression scores were used to analyze survival outcomes. These suggested that

Topics: Biomarkers, Tumor; Carcinoma, Renal Cell; Cyclin D1; Disease Progression; Gene Expression Profiling; Humans; Kidney Neoplasms; Platelet Endothelial Cell Adhesion Molecule-1; Prognosis; Transcriptome

The prognostic and therapeutic values of fibronectin have been reported in patients with renal cell carcinoma (RCC). However, the underlying mechanisms of malignancy in RCC are not completely understood. We found that silencing of fibronectin expression attenuated human RCC 786-O and Caki-1 cell growth and migration. Silencing of potential fibronectin receptor integrin α5 and integrin β1 decreased 786-O cell ability in movement and chemotactic migration. Biochemical examination revealed a reduction of cyclin D1 and vimentin expression, transforming growth factor-β1 (TGF-β1) production, as well as Src and Smad phosphorylation in fibronectin-silenced 786-O and Caki-1 cells. Pharmacological inhibition of Src decreased 786-O cell growth and migration accompanied by a reduction of cyclin D1, fibronectin, vimentin, and TGF-β1 expression, as well as Src and Smad phosphorylation. In 786-O cells, higher activities in cell growth and migration than in Caki-1 cells were noted, along with elevated fibronectin and TGF-β1 expression. The additions of exogenous fibronectin and TGF-β1 promoted Caki-1 cell growth and migration, and increased cyclin D1, fibronectin, vimentin, and TGF-β1 expression, as well as Src and Smad phosphorylation. These findings highlight the role of fibronectin in RCC cell growth and migration involving Src and TGF-β1 signaling.

Topics: Carcinoma, Renal Cell; Cell Line, Tumor; Cell Movement; Cell Proliferation; Cyclin D1; Fibronectins; Humans; Integrin alpha5; Integrin beta1; Kidney Neoplasms; Smad Proteins; src-Family Kinases; Vimentin

Cyclin-D1 (CCND1) belongs to the highly conserved cyclin family whose members are characterized by abundant expression during the cell cycle. As an oncogene, high level of CCND1 was observed and related to poor prognosis and tumor recurrence in many cancers. In this study, we focused on the role of CCND1 in the clinical outcome of clear cell renal cell carcinoma (ccRCC). Gene Expression Omnibus database, The Cancer Genome Atlas database, and immunohistochemical staining were used. The mRNA and protein levels of CCND1 were significantly enhanced in ccRCC tumor tissues. However, the low level of CCND1, but not high level of CCND1, was related to poor prognosis and tumor recurrence in ccRCC. Further analysis showed that CCND1 mRNA level decreased with increasing ccRCC tumor grades and the rate of recurrence in ccRCC patients. In a nomogram model, the CCND1 mRNA level was shown to help predict ccRCC patient recurrence. CCND1 is a strong determinant for prediction of recurrence. The patients with high CCND1 level appear to have a more favorable prognosis together with more frequent low-grade tumors and low rate of recurrence. This is the first study to investigate the prognostic roles of CCND1 in ccRCC and discovered that CCND1 had an unconventional positive impact on the clinical outcome of ccRCC patients.

Topics: Biomarkers, Tumor; Carcinoma, Renal Cell; Cyclin D1; Down-Regulation; Female; Gene Expression Regulation, Neoplastic; Humans; Kidney Neoplasms; Male; Neoplasm Grading; Neoplasm Recurrence, Local; Nomograms; Prognosis

Cytosine arabinoside (Ara-c) is a pyrimidine anti-metabolite that is capable of interfering with cellular proliferation by inhibiting DNA synthesis. Each inhibitor of cyclin-dependent kinase 4 (INK4) family member has the ability to bind to cyclin-dependent kinase 4 (CDK4) and inhibit the formation of the cell cycle-dependent CDK4/cyclin D1 complex, subsequently leading to cell cycle arrest in the G1/S phase. In this study, the expression of INK4 family genes in kidney cancer and the impact of these genes on patient prognosis were examined. Additionally, the effects of INK4 family genes and Ara-c on cell proliferation and tumor formation and development were examined. Finally, a potential association between Ara-c-induced cell cycle arrest and INK4-associated gene expression was evaluated. An upregulation of INK4 family genes was found to be positively correlated with the prognosis of patients with kidney cancer. Both the INK4 family genes and Ara-c were shown to induce cell cycle arrest and inhibit tumor formation and development. Moreover, Ara-c-induced cell cycle arrest was found to be associated with an Ara-c-induced upregulation of INK4 family gene expression, which ultimately inhibited the formation of the CDK4/cyclin D1 complex. These findings suggested that an upregulation of INK4 family genes has a positive effect on kidney cancer prognosis and can inhibit the formation and development of tumors. Moreover, Ara-c was shown to promote the upregulation of INK4 family genes, at the same time, Ara-c could directly regulate the cell cycle-dependent genes

Topics: Animals; Carcinoma, Renal Cell; Cell Line, Tumor; Cell Proliferation; Cyclin D1; Cyclin-Dependent Kinase 4; Cyclin-Dependent Kinase Inhibitor Proteins; Cytarabine; G1 Phase Cell Cycle Checkpoints; Gene Expression Regulation, Neoplastic; Heterografts; Humans; Kidney Neoplasms; Mice; Mice, Nude; Prognosis; S Phase Cell Cycle Checkpoints; Transfection; Up-Regulation

The third subunit of the COP9 signalosome (COPS3) is associated with cell proliferation and tumorigenesis process in cancer. The present study showed that the expression level of COPS3 was upregulated in malignant cell lines and COPS3 overexpression was related with clinical stage, T stage, historical grade. Kaplan-Meier survival curves showed that COPS3 may function as a prognostic factor for overall survival. CCK-8 and colony formation assays revealed that knockdown of COPS3 in ACHN and 786-O significantly impacted proliferation in vitro. In addition, flow cytometry showed that inhibition of COPS3 induced G0/G1 arrest and promoted apoptosis. COPS3 may promote kidney cancer progression by altering Phospho-AKT(Thr308), Cyclin D1 and Caspase-3 expression. Collectively, Our findings suggest that COPS3 may be a new potential target of ccRCC.

Topics: Adult; Aged; Aged, 80 and over; Carcinoma, Renal Cell; Caspase 3; Cell Line, Tumor; Cell Proliferation; COP9 Signalosome Complex; Cyclin D1; Disease Progression; Female; Gene Expression Regulation, Neoplastic; Humans; Kidney Neoplasms; Male; Middle Aged; Phosphorylation; Proto-Oncogene Proteins; Proto-Oncogene Proteins c-akt; Signal Transduction; Tissue Array Analysis; Up-Regulation

The chronic inflammatory microenvironment within or surrounding the primary renal cell carcinoma (RCC) site promotes oncogenic transformation as well as contributes to the development of metastasis. G3BP stress granule assembly factor 1 (G3BP1) was found to be involved in the regulation of multiple cellular functions. However, its functions in RCC have not been previously explored. Here, we first showed that the expression of G3BP1 is elevated in human RCC and correlates with RCC progression. In cultured RCC cells, knockdown of G3BP1 results in inhibition of tumor cell proliferation, migration, and invasion, consistently with the alteration of epithelial-mesenchymal transition (EMT) and cell proliferative markers, including Cadherins, Vimentin, Snail, Slug, c-Myc, and cyclin D1. Remarkably, knockdown of G3BP1 dramatically impaired the signaling connection of pro-inflammatory cytokine IL-6 stimulation and downstream STAT3 activation in RCC, thus eventually contributing to the disruption of IL-6-elicited RCC migration and metastasis. In addition, in vivo orthotopic tumor xenografts results confirmed that knockdown of G3BP1 suppressed RCC tumor growth and metastasis in mice. Collectively, our findings support the notion that G3BP1 promotes tumor progression and metastasis through IL-6/G3BP1/STAT3 signaling axis in RCC.

Topics: Aged; Animals; Cadherins; Carcinoma, Renal Cell; Cell Line, Tumor; Cell Movement; Cell Proliferation; Cyclin D1; DNA Helicases; Female; Gene Expression Regulation, Neoplastic; Humans; Interleukin-6; Kidney Neoplasms; Liver Neoplasms; Lung Neoplasms; Male; Mice; Middle Aged; Neoplasm Grading; Nephrectomy; Poly-ADP-Ribose Binding Proteins; RNA Helicases; RNA Recognition Motif Proteins; RNA, Small Interfering; Signal Transduction; Snail Family Transcription Factors; STAT3 Transcription Factor; Tumor Burden; Vimentin; Xenograft Model Antitumor Assays

The role of long non-coding RNA in Renal cell carcinoma (RCC) tumorigenesis and progression remains largely unknown. Here, we found that LINC01510 functions as a tumor suppressor in RCC tumorigenesis. We screened TCGA database and then found that LINC01510 is significantly down-regulated in malignant RCC tissues, and the lower expression of LINC01510 predicts poor prognosis. Moreover, the down-regulated LINC01510 was further confirmed in our fresh tissues and cell lines. Biological functions assays shown that Ectopic expression of LINC01510 not only inhibits RCC cell proliferation both in vitro and in vivo, but also impairs cell invasion ability. Moreover, we found overexpression of LINC01510 inhibits the expression of CCND1 and CCNE1, as well as MMPs (MMP2, MMP7 and MMP9), and thus affecting RCC cell cycle and invasion. Meanwhile, Western blot assays revealed that the expression of β-catenin is regulated by LINC01510; overexpression of β-catenin could partly rescue the cell viability and invasion ability caused by ectopic expression of LINC01510. Taken together, we found that LINC01510 regulates cell proliferation and invasion by modulating Wnt/β-catenin signaling in RCC.

Topics: Animals; Carcinoma, Renal Cell; Cell Line, Tumor; Cell Movement; Cell Proliferation; Cyclin D1; Down-Regulation; Gene Expression Regulation, Neoplastic; Genes, Tumor Suppressor; Humans; Kaplan-Meier Estimate; Kidney Neoplasms; Matrix Metalloproteinases; Mice, Inbred BALB C; Mice, Nude; Neoplasm Invasiveness; Prognosis; RNA, Long Noncoding; Transplantation, Heterologous; Wnt Signaling Pathway

BACKGROUND The aims of this study were to investigate the expression of methylenetetrahydrofolate dehydrogenase 1 (MTHFD1) in human tissue containing clear cell renal cell carcinoma (CCRCC) compared with normal renal tissue, and the effects of upregulating the expression of MTHFD1 in the human CCRCC cell line, Caki-1. MATERIAL AND METHODS Tumor and adjacent normal renal tissue were obtained from 44 patients who underwent radical nephrectomy for CCRCC. Caki-1 human CCRCC cells were divided into the control group, the empty vector (EV) group, and the plasmid-treated group that overexpressed MTHFD1. MTHFD1 mRNA and protein levels were measured by quantitative real-time polymerase chain reaction (qRT-PCR) and Western blot, respectively. The cell counting kit-8 (CCK-8) assay measured cell viability. Flow cytometry evaluated apoptosis and the cell cycle. Western blot measured the protein levels of MTHFD1, Bax, Bcl-2, Akt, p53, and cyclin D1, and qRT-PCR determined the gene expression profiles. RESULTS MTHFD1 mRNA and protein levels in CCRCC tumor tissues were significantly lower compared with adjacent normal renal tissue. MTHFD1 over-expression in Caki-1 cells inhibited cell proliferation, arrested cells in the G1 phase, increased cell apoptosis, and upregulated gene and protein expression of Bax/Bcl-2 and p53 and inhibited p-Akt, and cyclin D1. CONCLUSIONS MTHFD1 was underexpressed in CCRCC tissue when compared with normal renal tissue. MTHFD1 transfection of human CCRCC Caki-1 cells in vitro inhibited cell proliferation and promoted apoptosis, associated with reduced expression of cyclin D1, reduced Akt phosphorylation, and increased expression of Bax/Bcl-2 and p53.

Topics: Apoptosis; bcl-2-Associated X Protein; Carcinoma, Renal Cell; Cell Cycle; Cell Line, Tumor; Cell Movement; Cell Proliferation; Cyclin D1; Humans; Kidney Neoplasms; Methylenetetrahydrofolate Dehydrogenase (NADP); Minor Histocompatibility Antigens; Phosphorylation; Proto-Oncogene Proteins c-akt; Proto-Oncogene Proteins c-bcl-2; RNA, Messenger; Transcriptome; Tumor Suppressor Protein p53

Isoliquiritigenin (ISL) is a flavonoid with chalcone structure that has been noted in licorice and shallot, which are generally used in traditional Chinese medicine. ISL has demonstrated various pharmacological effects including antioxidant, anti-inflammatory and antitumor activity. However, the molecular mechanisms underlying the anticancer effects of ISL remain poorly understood. The present study revealed that ISL significantly decreased viability and induced apoptosis in human renal carcinoma Caki cells. The ISL-induced apoptosis was associated with the cleavage of caspase-9, -7 and -3, and that of PARP. Moreover, ISL increased the expression of pro-apoptotic protein Bax and diminished the expression of anti-apoptotic protein Bcl-2, and Bcl-xl, thereby increasing cytochrome c release. Treatment of cells with ISL also induced the expression of p53 through downregulation of murine double minute 2 (Mdm2). Furthermore, ISL generated reactive oxygen species (ROS), and pretreatment with ROS scavenger N-acetyl cysteine (NAC) and NADPH oxidase inhibitor diphenyleneiodonium abrogated the ISL-induced apoptosis. One of the key oncogenic signaling pathways is mediated through signal transducer and activator of transcription 3 (STAT3), which promotes abnormal cell proliferation. Incubation of cells with ISL markedly diminished phosphorylation and DNA binding activity of STAT3, and reduced expression of STAT3 responsive gene products, such as cyclin D1 and D2. ISL also attenuated constitutive phosphorylation of upstream kinase, Janus-activated kinase 2 (Jak2). Pretreatment with NAC abrogated the inhibitory effect of ISL on activation of STAT3 and blocked the cleavage of caspase-9, -7 and -3, and that of PARP in Caki cells. Taken together, the present study provides the first report that ISL induces apoptosis in Caki cells via generation of ROS, which causes induction of p53 and inhibition of the STAT3 signaling pathway.

Topics: Acetylcysteine; Antineoplastic Agents; Apoptosis; Carcinoma, Renal Cell; Caspase 3; Caspase 7; Caspase 9; Cell Line, Tumor; Cell Proliferation; Cell Survival; Chalcones; Cyclin D1; Cyclin D2; Down-Regulation; Humans; Janus Kinase 2; Kidney Neoplasms; NADPH Oxidases; Onium Compounds; Phosphorylation; Poly(ADP-ribose) Polymerases; Proto-Oncogene Proteins c-mdm2; Reactive Oxygen Species; Signal Transduction; STAT3 Transcription Factor; Tumor Suppressor Protein p53

Un-physiological activation of hypoxia inducible factor (HIF) is an early event in most renal cell cancers (RCC) following inactivation of the von Hippel-Lindau tumor suppressor. Despite intense study, how this impinges on cancer development is incompletely understood. To test for the impact of genetic signals on this pathway, we aligned human RCC-susceptibility polymorphisms with genome-wide assays of HIF-binding and observed highly significant overlap. Allele-specific assays of HIF binding, chromatin conformation and gene expression together with eQTL analyses in human tumors were applied to mechanistic analysis of one such overlapping site at chromosome 12p12.1. This defined a novel stage-specific mechanism in which the risk polymorphism, rs12814794, directly creates a new HIF-binding site that mediates HIF-1α isoform specific upregulation of its target BHLHE41. The alignment of multiple sites in the HIF cis-acting apparatus with RCC-susceptibility polymorphisms strongly supports a causal model in which minor variation in this pathway exerts significant effects on RCC development.

Topics: Alleles; Basic Helix-Loop-Helix Transcription Factors; Carcinoma, Renal Cell; Cell Line, Tumor; Chromatin Immunoprecipitation; Chromosomes, Human, Pair 12; Cyclin D1; Gene Expression Regulation, Neoplastic; Genetic Predisposition to Disease; Genome-Wide Association Study; HeLa Cells; Hep G2 Cells; High-Throughput Nucleotide Sequencing; Humans; Hypoxia-Inducible Factor 1, alpha Subunit; MCF-7 Cells; Polymorphism, Single Nucleotide; Quantitative Trait Loci; RNA, Small Interfering; Up-Regulation

Sunitinib is a tyrosine kinase inhibitor (TKI) targeting tumour angiogenesis in patients with advanced renal cell carcinoma (RCC). Currently no universally agreed model exists correlating the expression of angiogenesis markers with the success of treatment.. We retrospectively analysed archival tissue for 59 RCC patients treated with sunitinib. The expression of angiogenesis markers VEGF-A, VEGFR, PDGFββ, PDGFR, CCND1 and CA9 was assessed by immunohistochemistry (IHC) and correlated with overall survival (OS) and progression-free survival (PFS).. The median OS and median PFS of the whole group of patients was 24.6 months (17.3-34.2) and 19.5 months (11-27) respectively. VEGFA was positive in 29% of tumors, whereas VEGFR was expressed in only 12% of tumours. PDGFββ and its receptor were detected in a minority of cases. CCND1 and CA9 were positive in 44% and 60% of cases.. The OS and PFS achieved by our patients reflected previous observations seen with sunitinib, but no correlation was found between expression of angiogenesis markers and clinical outcome.

Topics: Adult; Aged; Aged, 80 and over; Angiogenesis Inhibitors; Antigens, Neoplasm; Becaplermin; Biomarkers, Tumor; Carbonic Anhydrase IX; Carcinoma, Renal Cell; Cyclin D1; Disease-Free Survival; Female; Humans; Indoles; Kaplan-Meier Estimate; Kidney Neoplasms; Male; Middle Aged; Neovascularization, Pathologic; Proto-Oncogene Proteins c-sis; Pyrroles; Receptors, Platelet-Derived Growth Factor; Receptors, Vascular Endothelial Growth Factor; Sunitinib; Treatment Outcome; Vascular Endothelial Growth Factor A

In this study we tried to systematically investigate the tumor suppressing microRNAs in ccRCC.. The MTS cell viability and colony formation assay were used to systematically detect the tumor suppressing ability of down-regulated miRNAs in ccRCC. Then miR-206 expression was detected by RT-qPCR and in situ hybridization in ccRCC cell lines and clinical samples. Oligonucleotides were used to overexpress or down-regulate miR-206. MTS cell viability, EdU cell proliferation, colony formation assay, flow cytometry, Xenograft subcutaneously and orthotopic implantations were done to examine tumor suppressing effects of miR-206 in vitro and in vivo. Luciferase assay was performed to verify the precise target of miR-206.. We reviewed and experimentally analyzed the currently available miRNA expression profiles data of ccRCC and identified miR-206 as one of the most critical tumor-suppressing microRNAs in ccRCC. In addition, miR-206 inhibited ccRCC cell proliferation through inducing cell cycle arrest by directly targeting cell cycle related gene CDK4, CDK9 and CCND1.. All these results suggested that miR-206 functioned as a novel cell cycle regulator and tumor suppressor in ccRCC and could be considered as a potential target for ccRCC therapy.

Topics: Animals; Carcinoma, Renal Cell; Cell Cycle Proteins; Cell Growth Processes; Cell Line, Tumor; Cyclin D1; Cyclin-Dependent Kinase 4; Cyclin-Dependent Kinase 9; G1 Phase Cell Cycle Checkpoints; Humans; Kidney Neoplasms; Male; Mice; Mice, Inbred BALB C; MicroRNAs; Resting Phase, Cell Cycle

Galectins are thought to be prognosticators for survival in renal cell cancer. However, the biological activity of galectin-3 (Gal-3) in renal carcinoma cells is still debated. In this study, immunohistochemical staining confirmed a high expression of Gal-3 in tumor tissue from renal cell carcinoma. Critically, Gal-3 expression was related to tumor cell differentiation. Consistent with Gal-3 expression in renal cell cancer, strong expression of Gal-3 was also observed in several renal tumor cell lines but not in normal renal cells. A Gal-3 high-expression cell line Caki-1 was chosen to study the biological activity of Gal-3. Using short hairpin RNA method, Gal-3 expression in Caki-1 cells was knocked down. We evidenced that Gal-3 knockdown inhibited cell proliferation and invasion, induced Caspase-3-dependent apoptosis and arrested cell cycle at G1 phase. Mechanically, Cyclin D1 expression decreased, but p27 increased after Gal-3 knockdown. Taken together, these results suggest that Gal-3 is related to the development of renal cell cancer and could serve as a target to therapy renal cell cancer.

Topics: Apoptosis; Carcinoma, Renal Cell; Caspase 3; Cell Line, Tumor; Cell Movement; Cell Proliferation; Cyclin D1; Cyclin-Dependent Kinase Inhibitor p27; G1 Phase Cell Cycle Checkpoints; Galectin 3; Gene Expression Regulation, Neoplastic; Gene Knockdown Techniques; Humans; Immunohistochemistry

The objective of the study was to investigate the impact of BTG2 on growth, migration and invasion of human clear cell renal cell carcinoma (ccRCC) cells. Endogenous expression of BTG2 was evaluated in the ccRCC cell lines (Caki-1, 786-O and Caki-2) and noncancerous human renal proximal tubular cell lines (HKC, HK-2 and RPTEC). BTG2 expression was decreased in the ccRCC cells compared with the noncancerous cells (P < 0.01). Then Caki-1 and 786-O cells described as suitable transfection hosts were used in transfection to carry out biological function studies. The three experimental groups were as follows: BTG2-ORF (transfected with BTG2-ORF plasmid), blank-Vector (transfected with pCMV6-Entry), and Cell-alone group (no DNA transfected in). BTG2 expression in the BTG2-ORF groups was significantly higher than that in the controls (P < 0.01). Cell growth was remarkably reduced and the number of migrating or invading cells was reduced in the BTG2-ORF groups compared with the controls (P < 0.01). Furthermore, Matrix Metalloproteinase-9 (MMP-9), Cyclin D1 and Cyclin E expression were reduced in the BTG2-ORF groups compared with the controls. Here, we have provided data for attenuated BTG2 expression in the ccRCC cells. Overexpressed BTG2 could inhibit cell proliferation, migration and invasion of human ccRCC, and the suppressive effects might be due to down-regulation of MMP-9, Cyclin D1 and Cyclin E expression.

Topics: Carcinoma, Renal Cell; Cell Growth Processes; Cell Line, Tumor; Cell Movement; Cyclin D1; Cyclin E; G1 Phase Cell Cycle Checkpoints; Humans; Immediate-Early Proteins; Kidney Neoplasms; Matrix Metalloproteinase 9; Neoplasm Invasiveness; Resting Phase, Cell Cycle; Transfection; Tumor Suppressor Proteins

To investigate the antitumor effect of lycorine on renal cell carcinoma ACHN cells and explore the possible mechanism.. We used flow cytometry to examine the effect of lycorine on ACHN cell cycle and apoptosis. The cell proliferation, migration and invasion were assessed with MTS assay, wound healing assay, and Transwell assay, respectively. Colony forming assay was performed, and the mRNA and protein levels of Bax, Bcl-2, survivin, caspase-3, cyclin D1 and CDK4 were measured with qRT-PCR and Western blotting.. Lycorine obviously inhibited the proliferation of ACHN cells with an IC(50) of 24.34 µmol/L. Lycorine also induced apoptosis of ACHN cells, caused cell cycle arrest at G(0)/G(1) phase, and suppressed the colony forming ability of the cells in a dose-dependent manner. The migration and invasion of ACHN cells were significantly inhibited by 5 µmol/L lycorine. Lycorine up-regulated the mRNA levels of CDK4, Bax, caspase-3 while down-regulated the levels of survivin, Bcl-2 and Cyclin D1; the protein levels of CDK4 and Bax were increased and cyclin D1, Bcl-2 and surviving expressions were decreased, but caspase-3 expression showed no significant changes following the treatment.. Lycorine has obvious antitumor effect against ACHN cells, suggesting its value as a new therapeutic agent for renal cell carcinoma.

Topics: Amaryllidaceae Alkaloids; Antineoplastic Agents, Phytogenic; Apoptosis; bcl-2-Associated X Protein; Carcinoma, Renal Cell; Caspase 3; Cell Cycle Checkpoints; Cell Line, Tumor; Cell Proliferation; Cyclin D1; Cyclin-Dependent Kinase 4; Humans; Inhibitor of Apoptosis Proteins; Phenanthridines; Proto-Oncogene Proteins c-bcl-2; Survivin

Renal cell carcinoma (RCC), also called kidney cancer or renal adenocarcinoma, is highly resistant to current treatments. It has been previously reported that a Kunitz-type inhibitor domain-containing protein, isolated from the salivary glands of the Amblyomma cajennense tick, triggers apoptosis in murine renal adenocarcinoma cells (Renca) by inhibiting the proteasome and endoplasmic reticulum stress. Of note, Amblyomin-X is the corresponding recombinant protein identified in the cDNA library from A. cajennense salivary glands. Herein, using orthotopic kidney tumors in mice, we demonstrate that Amblyomin-X is able to drastically reduce the incidence of lung metastases by inducing cell cycle arrest and apoptosis. The in vitro assays show that Amblyomin-X is capable of reducing the proliferation rate of Renca cells, promoting cell cycle arrest, and down-regulating the expression of crucial proteins (cyclin D1, Ki67 and Pgp) involved in the aggressiveness and resistance of RCC. Regarding non-tumor cells (NIH3T3), Amblyomin-X produced minor effects in the cyclin D1 levels. Interestingly, observing the image assays, the fluorescence-labelled Amblyomin-X was indeed detected in the tumor stroma whereas in healthy animals it was rapidly metabolized and excreted. Taken the findings together, Amblyomin-X can be considered as a potential anti-RCC drug candidate.

Topics: Animals; Apoptosis; Arthropod Proteins; ATP Binding Cassette Transporter, Subfamily B, Member 1; Carcinoma, Renal Cell; Cell Cycle Checkpoints; Cell Line, Tumor; Cyclin D1; Down-Regulation; Endoplasmic Reticulum Stress; Humans; Ki-67 Antigen; Kidney; Kidney Neoplasms; Male; Mice; Mice, Inbred BALB C; NIH 3T3 Cells; Proteasome Endopeptidase Complex; Proteasome Inhibitors; Proto-Oncogene Proteins c-bcl-2; Recombinant Proteins; Salivary Proteins and Peptides; Toxicity Tests; Xenograft Model Antitumor Assays

Renal cell carcinoma (RCC) is a seventh ranked malignancy with poor prognosis. RCC is lethal at metastatic stage as it does not respond to conventional systemic treatments, and there is an urgent need to find out promising novel biomarkers for effective treatment. The goal of this study was to evaluate the biomarkers that can be potential therapeutic target and predict effective inhibitors to treat the metastatic stage of RCC.. We conducted transcriptomic profiling to identify differentially expressed genes associated with RCC. Molecular pathway analysis was done to identify the canonical pathways and their role in RCC. Tissue microarrays (TMA) based immunohistochemical stains were used to validate the protein expression of cyclinD1 (CCND1) and were scored semi-quantitatively from 0 to 3+ on the basis of absence or presence of staining intensity in the tumor cell. Statistical analysis determined the association of CCND1 expression with RCC. Molecular docking analyses were performed to check the potential of two natural inhibitors, rutin and curcumin to bind CCND1.. We detected 1490 significantly expressed genes (1034, upregulated and 456, downregulated) in RCC using cutoff fold change 2 and p value < 0.05. Hes-related family bHLH transcription factor with YRPW motif 1 (HEY1), neuropilin 2 (NRP2), lymphoid enhancer-binding factor 1 (LEF1), and histone cluster 1 H3h (HIST1H3H) were most upregulated while aldolase B, fructose-bisphosphate (ALDOB), solute carrier family 12 (SLC12A1), calbindin 1 (CALB1) were the most down regulated genes in our dataset. Functional analysis revealed Wnt/β-catenin signaling as the significantly activated canonical pathway (z score = 2.53) involving cyclin D1 (CCND1). CCND1 was overexpressed in transcriptomic studies (FC = 2.26, p value = 0.0047) and TMA results also showed the positive expression of CCND1 in 53 % (73/139) of RCC cases. The ligands - rutin and curcumin bounded with CCND1 with good affinity.. CCND1 was one of the important upregulated gene identified in microarray and validated by TMA. Docking study showed that CCND1 may act as a potential therapeutic target and its inhibition could focus on the migratory, invasive, and metastatic potential of RCC. Further in vivo and in vitro molecular studies are needed to investigate the therapeutic target potential of CCND1 for RCC treatment.

Topics: Biomarkers, Tumor; Carcinoma, Renal Cell; Cluster Analysis; Cyclin D1; Gene Expression Profiling; Humans; Kidney Neoplasms; Molecular Docking Simulation; Saudi Arabia; Tissue Array Analysis

Collagen triple helix repeat containing-1 (CTHRC1), a secreted glycoprotein, is frequently upregulated in human cancers. However, the functional role of CTHRC1 in renal cell carcinoma (RCC) remains unclear. Thus, the aim of this study was to explore the role of CTHRC1 in RCC. Our results demonstrated that CTHRC1 was upregulated in RCC tissues and cell lines. Knockdown of CTHRC1 significantly inhibits the proliferation in RCCs. Furthermore, knockdown of CTHRC1 significantly inhibited the epithelial-to-mesenchymal transition (EMT) process in RCCs, as well as suppressed RCC cell migration and invasion. Mechanistically, knockdown of CTHRC1 inhibited the expression of β-catenin, c-Myc, and cyclin D1 in RCC cells. In conclusion, the results of the present study indicated that CTHRC1 downregulation inhibited proliferation, migration, EMT, and β-catenin expression in RCC cells. Therefore, CTHRC1 may be a potential therapeutic target for the treatment of RCC.

Topics: beta Catenin; Carcinoma, Renal Cell; Cell Line; Cell Line, Tumor; Cell Movement; Cell Proliferation; Cyclin D1; Down-Regulation; Epithelial-Mesenchymal Transition; Extracellular Matrix Proteins; Humans; Kidney Neoplasms; Neoplasm Invasiveness; Proto-Oncogene Proteins c-myc

Here we found that dual mTORC1/2 inhibitor AZD-2014 significantly inhibited RCC cell survival and growth, with higher efficiency than conventional mTORC1 inhibitors rapamycin and RAD001. RCC cell apoptosis was also induced by AZD-2014. AZD-2014 disrupted mTORC1/2 assembly and activation, while downregulating HIF-1α/2α and cyclin D1 expressions in RCC cells. Meanwhile, AZD-2014 activated autophagy, detected by p62 degradation, Beclin-1/ATG-5 upregulation and light LC3B-I/-II conversion. Autophagy inhibition by pharmacologic or siRNA-based means increased AZD-2014 activity in vitro, causing substantial RCC cell apoptosis. In vivo, AZD-2014 was more efficient than RAD001 in inhibiting 786-0 xenografts and downregulating HIF-1α/2α or p-AKT (Ser-473). Finally, AZD-2014's activity in vivo was further enhanced by co-administration of the autophagy inhibitor 3-methyaldenine. We provide evidence for clinical trials of using AZD-2014 in RCC treatment.

Topics: Animals; Apoptosis; Autophagy; Basic Helix-Loop-Helix Transcription Factors; Benzamides; Blotting, Western; Carcinoma, Renal Cell; Cell Line, Tumor; Cyclin D1; Drug Screening Assays, Antitumor; Female; Humans; Hypoxia-Inducible Factor 1, alpha Subunit; Kidney Neoplasms; Mechanistic Target of Rapamycin Complex 1; Mechanistic Target of Rapamycin Complex 2; Mice, Nude; Morpholines; Multiprotein Complexes; Protein Kinase Inhibitors; Proto-Oncogene Proteins c-akt; Pyrimidines; RNA Interference; TOR Serine-Threonine Kinases; Xenograft Model Antitumor Assays

The objective of the study was to investigate the impact of Numb on cell growth, cell migration, and invasion in human clear cell renal cell carcinoma (ccRCC). Endogenous expression of Numb was evaluated in the ccRCC cell lines (786-O, Caki-1, and Caki-2) and control reference human renal proximal tubular epithelial cells. Numb expression was decreased in the ccRCC cells compared with the control cells (P < 0.01). Then, 786-O and Caki-1 cells described as suitable transfection hosts were used in transfection to carry out biological function studies. The three experimental groups were as follows: Numb-ORF (transfected with Numb-ORF plasmid), blank-vector (transfected with pCMV6-entry), and cell-alone group (no DNA). Numb expression in the Numb-ORF groups was significantly higher than that in the controls (P < 0.01). Cell growth was remarkably reduced (P < 0.01), and the number of migrating or invading cells was reduced (P < 0.01) in the Numb-ORF groups compared with controls. Furthermore, the ratio of G0/G1 phase in the Numb-ORF group of 786-O cells was increased, and the S phase fraction and proliferation index was decreased (P < 0.01). Cyclin D1 and MMP-9 expression was reduced in the Numb-ORF groups compared with controls. Here, we have provided data for attenuated Numb expression in the ccRCC cells. Overexpression of Numb could induce G0/G1 phase arrest and inhibit cell proliferation, migration, and invasion. The suppressive effects might be due to downregulation of cyclin D1 or MMP-9 expression. Taken together, our data suggest that Numb may possibly function as a tumor suppressor involved in the carcinogenesis of ccRCC.

Topics: Apoptosis; Carcinoma, Renal Cell; Cell Line, Tumor; Cell Movement; Cell Proliferation; Cyclin D1; G1 Phase; Gene Expression Regulation, Neoplastic; Humans; Matrix Metalloproteinase 9; Membrane Proteins; Neoplasm Invasiveness; Nerve Tissue Proteins

The distinction between chromophobe renal cell carcinoma (ChRCC), clear cell renal cell carcinoma (CRCC) and renal oncocytoma may cause a diagnostic dilemma. The usefulness of DOG1, cyclin D1, CK7, CD117 and vimentin in the differential diagnosis of these renal epithelial tumors was investigated. DOG1 was positive in ChRCC (32 of 32, 100%) and in renal oncocytoma (21 of 21, 100%). In contrast, DOG1 was absent in all CRCC (0 of 30). Cyclin D1 was positive in renal oncocytomas (17 of 21, 81%) but negative in the ChRCC (0/23) and CRCC (0 of 30). CK7 was positive in ChRCC (30 of 32, 94%), but was negative in oncocytoma (only scattered single positive cells), and was only focal positive in two cases of CRCC. CD117 was expressed in 88% of ChRCC (28 of 32), 86% of renal oncocytoma (18 of 21), and was negative in all CRCC (0 of 30). Twenty-six of the 30 cases of CRCC were positive (87%) for vimentin with prominent membrane staining patterns. All 23 chromophobe carcinomas were negative for vimentin and 15 of 21 oncocytomas demonstrated focal vimentin positivity, but less than 10%. The above results demonstrate that: (1) DOG1 was very sensitive and specific marker for distinguish ChRCC from CRCC; (2) Cyclin D1 was a useful marker to discriminate between ChRCC and renal oncocytoma; (3) CK7 and CD117 were useful markers to distinguish ChRCC from renal oncocytoma and CRCC. (4) Vimentin was helpful for distinguishing clear cell RCC from chromophobe and oncocytoma (87% of clear cell RCC positive, negative in chromophobe, only focally positive in oncocytoma). (5) CK8/18, CK19, CD10, β-catenin and E-cadherin could not be used to distinguish ChRCC from renal oncocytoma and CRCC.

Topics: Adenoma, Oxyphilic; Anoctamin-1; Biomarkers, Tumor; Carcinoma, Renal Cell; Chloride Channels; Cyclin D1; Diagnosis, Differential; Humans; Immunohistochemistry; Keratin-7; Kidney Neoplasms; Neoplasm Proteins; Proto-Oncogene Proteins c-kit; Vimentin

Nuclear auto-antigenic sperm protein (NASP), initially described as a highly auto-immunogenic testis and sperm-specific protein, is a histone chaperone that is proved to present in all dividing cells. NASP has two splice variants: testicular NASP (tNASP) and somatic form of NASP (sNASP). Only cancer, germ, transformed, and embryonic cells have a high level of expression of the tNASP. Up to now, little has been known about tNASP in renal cell carcinoma (RCC). In the present study, the molecular mechanism of tNASP in RCC was explored. The expression level of tNASP in 16 paired human RCC specimens was determined. Downregulation of tNASP by small interfering RNA (siRNA) was transfected in RCC cell lines. The effect of downregulation of tNASP by siRNA on cell colony formation and proliferation was examined by colony formation assay and CCK-8 assay, cell cycle was analyzed by flow cytometry, and the expression of cyclin D1 and P21 were detected by Western blotting. ERK/MAPK signaling was also analyzed. tNASP has a relative high expression level in human RCC tissues. Via upregulation of P21 and downregulation of cyclinD1, silence of tNASP can inhibit cell proliferation, which induces cell cycle arrest. Furthermore, ERK signaling pathway is confirmed to mediate the regulation of cell cycle-related proteins caused by silence of tNASP. Our research demonstrates that knockdown of tNASP effectively inhibits the proliferation and causes G1 phase arrest through ERK/MAPK signal pathway.

Topics: Autoantigens; Carcinoma, Renal Cell; Cell Cycle Checkpoints; Cell Cycle Proteins; Cell Line, Tumor; Cell Proliferation; Cyclin D1; G1 Phase; Gene Expression Regulation, Neoplastic; Humans; MAP Kinase Signaling System; Nuclear Proteins; p21-Activated Kinases; RNA, Small Interfering

Mutational inactivation of the VHL tumor suppressor plays key roles in the development of renal cell carcinoma (RCC), and mutated VHL-mediated VEGF induction has become the main target for the current RCC therapy. Here we identified a signal pathway of VEGF induction by androgen receptor (AR)/miRNA-145 as a new target to suppress RCC progression. Mechanism dissection revealed that AR might function through binding to the androgen receptor element (ARE) located on the promoter region of miRNA-145 to suppress p53's ability to induce expression of miRNA-145 that normally suppresses expression of HIF2α/VEGF/MMP9/CCND1. Suppressing AR with AR-shRNA or introducing exogenous miRNA-145 mimic can attenuate RCC progression independent of VHL status. MiR-145 mimic in preclinical RCC orthotopic xenograft mouse model revealed its efficacy in suppression of RCC progression. These results together identified signals by AR-suppressed miRNA-145 as a key player in the RCC progression via regulating HIF2α/VEGF/MMP9/CCND1 expression levels. Blockade of the newly identified signal by AR inhibition or miRNA-145 mimics has promising therapeutic benefit to suppress RCC progression.

Topics: Animals; Basic Helix-Loop-Helix Transcription Factors; Binding Sites; Carcinoma, Renal Cell; Cell Line, Tumor; Cell Movement; Cell Proliferation; Cyclin D1; Disease Progression; Down-Regulation; Gene Expression Regulation, Neoplastic; Humans; Kidney Neoplasms; Male; Matrix Metalloproteinase 9; Mice, Nude; MicroRNAs; Mutation; Neoplasm Invasiveness; Promoter Regions, Genetic; Receptors, Androgen; RNA Interference; RNAi Therapeutics; Signal Transduction; Time Factors; Transfection; Vascular Endothelial Growth Factor A; Von Hippel-Lindau Tumor Suppressor Protein; Xenograft Model Antitumor Assays

Oncocytomas are predominantly benign neoplasms possessing pathogenic mitochondrial mutations and accumulation of respiration-defective mitochondria, characteristics of unknown significance. Using exome and transcriptome sequencing, we identified two main subtypes of renal oncocytoma. Type 1 is diploid with CCND1 rearrangements, whereas type 2 is aneuploid with recurrent loss of chromosome 1, X or Y, and/or 14 and 21, which may proceed to more aggressive eosinophilic chromophobe renal cell carcinoma (ChRCC). Oncocytomas activate 5' adenosine monophosphate-activated protein kinase (AMPK) and Tp53 (p53) and display disruption of Golgi and autophagy/lysosome trafficking, events attributed to defective mitochondrial function. This suggests that the genetic defects in mitochondria activate a metabolic checkpoint, producing autophagy impairment and mitochondrial accumulation that limit tumor progression, revealing a novel tumor-suppressive mechanism for mitochondrial inhibition with metformin. Alleviation of this metabolic checkpoint in type 2 by p53 mutations may allow progression to eosinophilic ChRCC, indicating that they represent higher risk.

Topics: Adenoma, Oxyphilic; AMP-Activated Protein Kinases; Autophagy; Carcinoma, Renal Cell; Cathepsins; Cell Transformation, Neoplastic; Cyclin D1; DNA Copy Number Variations; Female; Golgi Apparatus; Humans; Karyotype; Kidney; Kidney Neoplasms; Lysosomal Membrane Proteins; Lysosomes; Male; Metformin; Mitochondria; Sequence Analysis, RNA; Transcriptome; Tumor Suppressor Protein p53

Renal tumours have recently been described in association with mutations in the gene encoding the B subunit of succinate dehydrogenase, a mitochondrial Krebs cycle and electron transport chain enzyme (SDHB-associated renal cell carcinomas). The aim of this study was to investigate the roles of different signalling pathways in the pathogenesis of these tumours.. We used immunohistochemistry and antibodies against phospho-specific epitopes to examine the activity of three potential signalling pathways in tumour cells of three genetically confirmed cases of SDHB-associated renal cell carcinomas. We found no evidence supporting a role for either the mTOR [p-mTOR (Ser2448), p-S6 riboprotein (Ser235/236)] or hypoxia-inducible (carbonic anhydrase 9 and EGFR) pathways. However, there was immunohistochemical reactivity for phosphorylated AMP-dependent kinase (p-AMPK Thr172) and glycogen synthase kinase 3 (GSK3) phosphorylation (p-GSK3 Ser12), and nuclear expression of cyclin D1.. We suggest that these tumours may arise through a mechanism involving ATP depletion, activation of AMPK, and induction of cyclin D1, and that this may be a unique pathway of tumour development that has the potential for therapeutic intervention in these rare tumours.

Topics: Adult; Aged; AMP-Activated Protein Kinases; Carcinoma, Renal Cell; Cyclin D1; Female; Germ-Line Mutation; Glycogen Synthase Kinase 3; Humans; Immunohistochemistry; Kidney Neoplasms; Middle Aged; Phosphorylation; Signal Transduction; Succinate Dehydrogenase; TOR Serine-Threonine Kinases

Renal cell carcinoma (RCC) is a family of distinct tumors, and a variety of molecules have been evaluated as prognostic markers for RCC. Cyclin D1, a cell cycle regulator, is overexpressed in several primary tumors.. To evaluate cyclin D1 expression as a prognostic marker in RCC.. In total, 109 tumor specimens from patients with RCC were obtained from 2005 to 2010 at Hospital das Clínicas--Ribeirão Preto School of Medicine--USP, Brazil, and submitted to immunohistochemical analysis along with seven normal kidney tissue samples.. All of the normal kidney samples lacked cyclin D1 immunohistochemical staining. In addition, there was lower protein expression in the papillary and chromophobe RCC samples. Patients with cyclin D1(low) tumors (≤ 30 % positive cells) showed worse clinical outcome (p = 0.03), lower survival without metastasis and/or death by RCC (p = 0.03), high nuclear grade (p = 0.001), larger tumor size (p = 0.01), presence of symptoms at diagnosis (p = 0.04), necrosis (p = 0.004) and sarcomatoid morphology (p = 0.04). After multivariate analysis, cyclin D1 was not an independent significant factor for worse outcome; however, it improved the accuracy of the adopted prognostic system. The analysis performed for clear cell RCC alone showed similar statistical significance to that of the total cases.. Cyclin D1 protein was overexpressed in RCC. The types of RCC appear to exhibit different immunohistochemical staining patterns for cyclin D1; high protein expression was related to good clinical outcome and to most known favorable prognostic factors. Further investigations are necessary to reveal which mechanisms lead to cyclin D1 accumulation in neoplastic cells.

Topics: Adolescent; Adult; Aged; Aged, 80 and over; Biomarkers, Tumor; Carcinoma, Renal Cell; Child; Cyclin D1; Female; Humans; Kidney; Kidney Neoplasms; Male; Middle Aged; Neoplasm Grading; Neoplasm Metastasis; Neoplasm Staging; Predictive Value of Tests; Prognosis; Survival Rate; Tumor Burden; Young Adult

Several entities have been individualized recently within the family of renal neoplasms with papillary features. Clear cell papillary renal cell carcinoma (CCPRCC) was first described in patients with end-stage renal disease, but is also observed in patients with normal renal function. The objective of this study was to document the clinicopathological and immunohistochemical characteristics of CCPRCC, with a special emphasis on cyclin D1 expression.. The patients were 25 men and 17 women, mean age 60.7 years. Seventeen patients had a chronic renal disease. All tumours were stage pT1, with a mean diameter of 2 cm. Six tumours were multifocal. Tumours cells were mainly cuboidal, with clear cytoplasm and low-grade nuclei apically aligned. In all cases, Fuhrman nuclear grade was one or two. No necrosis or vascular invasion was seen. During follow-up (10-72 months), no metastasis or death related to the disease was observed. Immunohistochemistry showed strong and diffuse cytokeratin 7 immunoreactivity in all cases, but no labelling for AMACR or TFE3. There was diffuse nuclear cyclin D1 immunoreactivity in 83% of cases.. CCPRCC is now a well-characterized entity. This tumour is an indolent and very low-grade neoplasm. Here we report the first study, to our knowledge, demonstrating the overexpression of cyclin D1 immunostaining by this tumour.

Topics: Adult; Aged; Biomarkers, Tumor; Carcinoma, Renal Cell; Cyclin D1; Female; Gene Expression Regulation, Neoplastic; Humans; Immunohistochemistry; In Situ Hybridization, Fluorescence; Kidney Neoplasms; Male; Middle Aged; Neoplasm Grading

The aim of this study was to investigate the effects of the dickkopf homolog 4 (DKK4)/Wnt/β-catenin signaling pathway on tumorigenesis and metastasis in clear cell renal cell carcinoma (ccRCC), as well as to elucidate the underlying mechanisms. We examined the expression of DKK4 in 30 cases of ccRCC and matched adjacent normal tissues, and investigated its correlation with clinicopathological characteristics. Stable DKK4-transfected cells were established, and DKK4 functional analyses were performed, including a T-cell factor/lymphoid enhancer factor (TCF/LEF) reporter assay, and experiments on cell viability, apoptosis, invasive capability and tumor growth in vivo. Finally, western blot analysis was performed to detect Von Hippel-Lindau (VHL) expression in 50 clinical specimens. The expression levels of the DKK4, β-catenin and β-catenin downstream target genes, cyclin D1 and c-myc, were determined in the these specimens, as well as in RCC4(-), T3-14(+) cell lines by qRT-PCR and western blot analysis. The same tests were also performed in human embryonic kidney (HEK)293 cells which were transfected with the pCDH-DKK4 plasmid. After 6 weeks the tumor weight significantly increased in the mice transfected with the tumor cells. DKK4 mRNA and protein expression levels were significantly upregulated (p<0.001). DKK4 was distinctly overexpressed (68.0%) in all patient tissues. VHL(-) samples accounted for 60.0% of all samples, while DKK4 expression was significantly upregulated in 50% of these samples, indicating a correlation with VHL(-) expression (r=0.403, p<0.05). We also observed reduced expression levels of cyclin D1, c-myc and β-catenin (to a greater extent) in the VHL(-), RCC4(-) and T3-14(+) cells, as well as in the stably transfected HEK293 cells. DKK4 may be an oncogene, and its upregulated expression may be involved in the pathogenesis of ccRCC as a downstream gene of VHL. By activating other pathways apart from the Wnt/β-catenin pathway, DKK4 may play an important role in ccRCC tumorigenesis and metastasis.

Topics: Animals; Carcinoma, Renal Cell; Cell Line, Tumor; Cyclin D1; Gene Expression Regulation, Neoplastic; Humans; Intercellular Signaling Peptides and Proteins; Kidney Neoplasms; Mice; Signal Transduction; Von Hippel-Lindau Tumor Suppressor Protein

Phosphodiesterase type 5 (PDE5) plays a key role in regulating the intracellular cyclic GMP (cGMP) concentration, which influences anti-proliferative and pro-apoptotic mechanisms in multiple carcinomas. PDE5 inhibitors, such as exisulind and its analogs have anticancer activities. In this study, we found that suppressing PDE5 gene expression by PDE5 siRNA inhibited cell proliferation and induced apoptosis in OS-RC-2 human renal cell carcinoma cells. These effects were enhanced by 8-Br-cGMP, a cell membrane permeable cGMP derivative, and were inhibited by KT5823, a protein kinase G (PKG) inhibitor, indicating that PKG was activated by intracellular cyclic GMP. In addition, there was a reduction in both the mRNA and protein expression of cyclin D1, while p21 protein expression was increased; the reduction in cyclin D1 expression was blocked by the proteasome inhibitor, MG132, or c-Jun N-terminal kinase (JNK) inhibitor; both β-catenin and JNK were phosphorylated by activated PKG. Furthermore, p21 protein expression was decreased in Sp1 siRNA transfected-cells treated with 8-Br-cGMP, indicating that p21 may be partly controlled by the PKG activation through Sp1. Furthermore, we found that PKG Iβ was responsible for the anticancer activities. Our findings indicate that the downregulation of PKG-activated genes, such as cyclin D1 partly accounts for the pro-apoptotic effects in PDE5 siRNA-transfected OS-RC-2 cells.

Topics: Antineoplastic Agents; Apoptosis; beta Catenin; Carbazoles; Carcinoma, Renal Cell; Cell Line, Tumor; Cell Proliferation; Cell Survival; Cyclic GMP; Cyclic Nucleotide Phosphodiesterases, Type 5; Cyclin D1; Cyclin-Dependent Kinase Inhibitor p21; Gene Expression Regulation; Humans; JNK Mitogen-Activated Protein Kinases; Phosphodiesterase 5 Inhibitors; Phosphorylation; Signal Transduction; Sp1 Transcription Factor; Sulindac

Renal cell carcinoma (RCC) is a complex with diverse biological characteristics and distinct molecular signature. New target therapies to molecules that drive RCC initiation and progression have achieved promising responses in some patients, but the total effective rate is still far from satisfaction. Dachshund (DACH1) network is a key signaling pathway for kidney development and has recently been identified as a tumor suppressor in several cancer types. However, its role in renal cell carcinoma has not been fully investigated.. Immunohistochemical staining for DACH1, PCNA and cyclin D1 was performed on human renal tissue microarrays and correlation with clinic-pathological characteristics was analyzed. In vitro proliferation, apoptosis and in vivo tumor growth were evaluated on human renal cancer cell lines with decitabine treatment or ectopic expression of DACH1. Downstream targets and potential molecular mechanism were investigated through western blot, immunoprecipitation and reporter gene assays.. Expression of DACH1 was significantly decreased in human renal carcinoma tissue. DACH1 protein abundance was inversely correlated with the expression of PCNA and cyclin D1, tumor grade, and TNM stage. Restoration of DACH1 function in renal clear cell cancer cells inhibited in vitro cellular proliferation, S phase progression, clone formation, and in vivo tumor growth. In mechanism, DACH1 repressed cyclin D1 transcription through association with AP-1 protein.. Our results indicated that DACH1 was a novel molecular marker of RCC and it attributed to the malignant behavior of renal cancer cells. Re-activation of DACH1 may represent a potential therapeutic strategy.

Topics: Animals; Carcinoma, Renal Cell; Cell Growth Processes; Cell Proliferation; Cyclin D1; Eye Proteins; Heterografts; Humans; Immunohistochemistry; Kidney Neoplasms; Male; Mice; Mice, Nude; Proliferating Cell Nuclear Antigen; Signal Transduction; Transcription Factors

Renal cell carcinoma (RCC) is a malignant disease insensitive to conventional treatments such as radiochemotherapy and immunotherapy. Search for new approaches to induce cancer cell apoptosis will improve the management of RCC. Here, we reported that zerumbone, a monosesquiterpine, shows anticancer effects on human RCC cells via induction of apoptosis in vitro. Human renal clear cell carcinoma 786-0 and 769-P cell lines were used as the model system. Exposure of RCC cells to zerumbone resulted in cell viability inhibition, accompanied by DNA fragmentation and increased apoptotic index. Mechanically, treatment of RCC cells with zerumbone activated caspase-3 and caspase-9, and finally led to cleavage of PARR In addition, downregulation of Gli-1 and Bcl-2, which were closely related to the chemoresistance of RCC, was observed in zerumbone-treated RCC cells. Taken together, our study provided the first evidence that zerumbone imparted strong inhibitory and apoptotic effects on human RCC cells. The zerumbone-induced apoptosis might be related to the activation of the caspase cascade and deregulation of the Gli-1/Bcl-2 pathway. Our results suggest that zerumbone merit further investigation as an apoptosis inducer as well as a novel RCC chemotherapeutic agent in the clinical setting.

Topics: Apoptosis; Blotting, Western; Carcinoma, Renal Cell; Caspases; Cell Line, Tumor; Cell Survival; Cyclin D1; DNA Fragmentation; Enzyme Activation; Flow Cytometry; Humans; Oncogene Proteins; Sesquiterpenes; Signal Transduction; Trans-Activators; Zinc Finger Protein GLI1

Renal cell carcinoma (RCC) is the most common type of kidney cancer, and represents the third most common urological malignancy. Despite the advent of targeted therapies for RCC and the improvement of the lifespan of patients, its cost-effectiveness restricted the therapeutic efficacy. In a recent report, we showed that synthetic phosphoethanolamine (Pho-s) has a broad antitumor activity on a variety of tumor cells and showed potent inhibitor effects on tumor progress in vivo.. We show that murine renal carcinoma (Renca) is more sensitive to Pho-s when compared to normal immortalized rat proximal tubule cells (IRPTC) and human umbilical vein endothelial cells (HUVEC). In vitro anti-angiogenic activity assays show that Pho-s inhibits endothelial cell proliferation, migration and tube formation. In addition, Pho-s has anti-proliferative effects on HUVEC by inducing a cell cycle arrest at the G2/M phase. It causes a decrease in cyclin D1 mRNA, VEGFR1 gene transcription and VEGFR1 receptor expression. Pho-s also induces nuclear fragmentation and affects the organization of the cytoskeleton through the disruption of actin filaments. Additionally, Pho-s induces apoptosis through the mitochondrial pathway. The putative therapeutic potential of Pho-s was validated in a renal carcinoma model, on which our remarkable in vivo results show that Pho-s potentially inhibits lung metastasis in nude mice, with a superior efficacy when compared to Sunitinib.. Taken together, our findings provide evidence that Pho-s is a compound that potently inhibits lung metastasis, suggesting that it is a promising novel candidate drug for future developments.

Topics: Angiogenesis Inhibitors; Animals; Antineoplastic Agents; Apoptosis; Carcinoma, Renal Cell; Caspase 3; Cell Cycle Checkpoints; Cell Line, Tumor; Cell Movement; Cell Proliferation; Cyclin D1; Cytoskeleton; Ethanolamines; Gene Expression Regulation, Neoplastic; Human Umbilical Vein Endothelial Cells; Humans; Indoles; Kidney Tubules, Proximal; Lung Neoplasms; Male; Mice; Mice, Inbred BALB C; Mitochondria; Neoplasm Metastasis; Neovascularization, Pathologic; Oxidation-Reduction; Pyrroles; Rats; Sunitinib; Transcription, Genetic; Vascular Endothelial Growth Factor Receptor-1; Wound Healing

Krüppel-like factor 4 (KLF4) is a transcription factor with diverse functions in various cancer types; however, the function of KLF4 in clear cell renal cell carcinoma (ccRCC) carcinogenesis remains unknown. In this study, we initially examined KLF4 expression by using a cohort of surgically removed ccRCC specimens and cell lines. Results indicated that the transcription and translation of KLF4 were lower in ccRCC tissues than in patient-matched normal tissues. Furthermore, the KLF4 expression was significantly downregulated in the five ccRCC cell lines at protein and mRNA levels compared with that in normal renal proximal tubular epithelial cell lines (HKC). KLF4 downregulation was significantly correlated with tumor stage and tumor diameter. Promoter hypermethylation may contribute to its low expression. In addition, in vitro studies indicated that the KLF4 overexpression significantly inhibited proliferation in human ccRCC cell lines 786-O and ACHN. Moreover, the KLF4 overexpression arrested the cell cycle progress at the G1/S phase transition by upregulating p21 (WAF1/CIP1) expression and downregulating cyclin D1 expression, KLF4 knockdown in HKC cells did the opposite. In vivo studies confirmed the anti-proliferative effect of KLF4. Our results suggested that KLF4 had an important function in suppressing the growth of ccRCC.

Topics: Adult; Aged; Animals; Carcinoma, Renal Cell; Cell Cycle Checkpoints; Cell Line, Tumor; Cell Proliferation; Cell Transformation, Neoplastic; CpG Islands; Cyclin D1; Cyclin-Dependent Kinase Inhibitor p21; DNA Methylation; Female; G1 Phase Cell Cycle Checkpoints; Gene Silencing; Heterografts; Humans; Kidney Neoplasms; Kruppel-Like Factor 4; Kruppel-Like Transcription Factors; Male; Mice; Middle Aged; Neoplasm Grading; Neoplasm Staging; Promoter Regions, Genetic; S Phase Cell Cycle Checkpoints; Tumor Burden

Acyldepsipeptides are a group of potent antibiotics discovered in the secondary metabolites of Streptomyces species. However, besides the function of antibiotics, no other activities have been reported about these important compounds so far. In the course of searching the natural products as chemotherapeutic agents for renal cell carcinoma, we found that ADEP1, a major metabolic component of Streptomyces hawaiiensis NRRL 15010, could effectively inhibit the growth of 786-O, 769-P, and ACHN renal carcinoma cells in MTT assay. Flow cytometric analysis demonstrated that ADEP1 could block the cell cycle arrested at G1 phase. Moreover, it was found that ADEP1 down-regulated the expressions of cyclin D1, CDK4 and PCNA and inhibited activity of MAPK-ERK pathway by detection of decreased expression of phosphorylated ERK1/2 and c-Fos in 786-O and 769-P cells by Western blotting. To our knowledge, this is the first report concerning to the antitumor activities of acyldepsipeptides. Based on these results, ADEP1 may become a promising lead compound to be developed a novel chemotherapeutic agent for treatment of renal carcinoma.

Topics: Antibiotics, Antineoplastic; Carcinoma, Renal Cell; Cyclin D1; Depsipeptides; G1 Phase; G1 Phase Cell Cycle Checkpoints; Humans; Kidney Neoplasms; MAP Kinase Signaling System; Peptides, Cyclic

Renal cell carcinoma (RCC) is the sixth most common cancer in the US. While RCC is highly metastatic, there are few therapeutics options available for patients with metastatic RCC, and progression-free survival of patients even with the newest targeted therapeutics is only up to two years. Thus, novel therapeutic targets for this disease are desperately needed. Based on our previous metabolomics studies showing alteration of peroxisome proliferator-activated receptor α (PPARα) related events in both RCC patient and xenograft mice materials, this pathway was further examined in the current study in the setting of RCC. PPARα is a nuclear receptor protein that functions as a transcription factor for genes including those encoding enzymes involved in energy metabolism; while PPARα has been reported to regulate tumor growth in several cancers, it has not been evaluated in RCC. A specific PPARα antagonist, GW6471, induced both apoptosis and cell cycle arrest at G0/G1 in VHL(+) and VHL(-) RCC cell lines (786-O and Caki-1) associated with attenuation of the cell cycle regulatory proteins c-Myc, Cyclin D1, and CDK4; this data was confirmed as specific to PPARα antagonism by siRNA methods. Interestingly, when glycolysis was blocked by several methods, the cytotoxicity of GW6471 was synergistically increased, suggesting a switch to fatty acid oxidation from glycolysis and providing an entirely novel therapeutic approach for RCC.

Topics: Apoptosis; Carcinoma, Renal Cell; Cell Cycle Checkpoints; Cell Line; Cell Line, Tumor; Cell Survival; Cyclin D1; Cyclin-Dependent Kinase 4; Deoxyglucose; G1 Phase; Glucose; Glycolysis; Humans; Immunoblotting; Immunohistochemistry; Kidney Neoplasms; Oxazoles; PPAR alpha; Proto-Oncogene Proteins c-myc; Resting Phase, Cell Cycle; RNA Interference; Tyrosine

MicroRNAs regulate post-transcriptomic landscape in many tumors including renal cell carcinoma. We have recently shown significantly increased expression of miR-21 in renal tumors and that this miRNA contributes to the proliferation of renal cancer cells in culture. However, the mechanism by which miR-21 regulates renal cancer cell proliferation is poorly understood. Addiction to constitutive NFκB activity is hallmark of many cancers including renal cancer. Using miR-21 Sponge in renal cancer cells to block endogenous function of miR-21, we show inhibition of phosphorylation of p65 subunit of NFκB, IKKβ and IκB, which results in attenuation of NFκB transcriptional activity. Subtle reduction in the tumor suppressor PTEN has been linked to various malignancies. We showed previously that miR-21 targeted PTEN in renal cancer cells. Inhibition of PTEN by siRNAs restored miR-21 Sponge-induced suppression of phosphorylation of p65, IKKβ, IκB and NFκB transcriptional activity along with reversal of miR-21 Sponge-reduced phosphorylation of Akt. Expression of constitutively active Akt protected against miR-21 Sponge- and PTEN-mediated decrease in p65/IKKβ/IκB phosphorylation and NFκB transcriptional activity. Furthermore, IKKβ and p65 were required for miR-21-induced renal cancer cell proliferation. Interestingly, miR-21 controlled the expression of cyclin D1 through NFκB-dependent transcription. Finally, we demonstrate that miR-21-regulated renal cancer cell proliferation is mediated by cyclin D1 and CDK4. Together, our results establish a molecular order of a phosphatase-kinase couple involving PTEN/Akt/IKKβ and NFκB-dependent cyclin D1 expression for renal carcinoma cell proliferation by increased miR-21 levels.

Topics: Carcinoma, Renal Cell; Cell Line, Tumor; Cell Proliferation; Cyclin D1; Gene Expression Regulation, Neoplastic; Humans; Kidney; Kidney Neoplasms; MicroRNAs; NF-kappa B; PTEN Phosphohydrolase; Signal Transduction; Transcriptional Activation

Somatic mutations or loss of expression of tumor suppressor VHL happen in the vast majority of clear cell Renal Cell Carcinoma, and it's causal for kidney cancer development. Without VHL, constitutively active transcription factor HIF is strongly oncogenic and is essential for tumor growth. However, the contribution of individual HIF-responsive genes to tumor growth is not well understood. In this study we examined the contribution of important HIF-responsive genes such as VEGF, CCND1, ANGPTL4, EGLN3, ENO2, GLUT1 and IGFBP3 to tumor growth in a xenograft model using immune-compromised nude mice. We found that the suppression of VEGF or CCND1 impaired tumor growth, suggesting that they are tumor-promoting genes. We further discovered that the lack of ANGPTL4, EGLN3 or ENO2 expression did not change tumor growth. Surprisingly, depletion of GLUT1 or IGFBP3 significantly increased tumor growth, suggesting that they have tumor-inhibitory functions. Depletion of IGFBP3 did not lead to obvious activation of IGFIR. Unexpectedly, the depletion of IGFIR protein led to significant increase of IGFBP3 at both the protein and mRNA levels. Concomitantly, the tumor growth was greatly impaired, suggesting that IGFBP3 might suppress tumor growth in an IGFIR-independent manner. In summary, although the overall transcriptional activity of HIF is strongly tumor-promoting, the expression of each individual HIF-responsive gene could either enhance, reduce or do nothing to the kidney cancer tumor growth.

Topics: Angiopoietin-Like Protein 4; Angiopoietins; Animals; Carcinoma, Renal Cell; Cell Line, Tumor; Cyclin D1; Disease Models, Animal; Gene Expression Regulation, Neoplastic; Gene Knockout Techniques; Glucose Transporter Type 1; Humans; Hypoxia-Inducible Factor 1; Hypoxia-Inducible Factor-Proline Dioxygenases; Insulin-Like Growth Factor Binding Protein 3; Mice; Phosphopyruvate Hydratase; Receptor, IGF Type 1; Transcriptional Activation; Tumor Burden; Vascular Endothelial Growth Factor A; Von Hippel-Lindau Tumor Suppressor Protein; Xenograft Model Antitumor Assays

Renal cell carcinoma (RCC) is a heterogeneous group of kidney cancers with clear cell RCC (ccRCC) as the major subgroup. To expand the number of clinically relevant tumor-associated antigens (TAA) that can be targeted by immunotherapy, we analyzed samples from 23 patients with primary ccRCC for the expression and immunogenicity of various TAAs. We found high-frequency expression of MAGE-A9 and NY-ESO-1 in 36% and 55% of samples, respectively, and overexpression of PRAME, RAGE-1, CA-IX, Cyclin D1, ADFP, C-MET, and RGS-5 in many of the tumor samples. We analyzed the blood of patients with HLA-A2(+) ccRCC for the presence of CD8(+) T cells specific for TAA-derived HLA-A2-restricted peptides and found spontaneous responses to cyclin D1 in 5 of 6 patients with Cyclin D1-positive tumors. Cyclin D1-specific CD8(+) T cells secreted TNF-α, IFN-γ, and interleukin-2 (IL-2), and degranulated, indicating the presence of polyfunctional tumor-specific CD8(+) T cells in the blood of these patients with ccRCC. The high frequency (43%) of Cyclin D1 overexpression and the presence of functional cyclin D1-specific T cells in 83% of these patients with ccRCC suggest that cyclin D1 may be a target for immunotherapeutic strategies.

Topics: Adult; Aged; Aged, 80 and over; Antigens, Neoplasm; Carcinoma, Renal Cell; CD8-Positive T-Lymphocytes; Cyclin D1; Female; HLA-A2 Antigen; Humans; Immunotherapy; Kidney Neoplasms; Male; Middle Aged; Reverse Transcriptase Polymerase Chain Reaction

A recent genome-wide association study of renal cell carcinoma (RCC) in European population has identified genetic variants in the regions of 2p21 (rs7579899), 11q13.3 (rs7105934) and 12q24.31 (rs4765623) conferred susceptibility to RCC. In our study, we assessed whether these polymorphisms are also associated with RCC risk in a Chinese population. We genotyped these polymorphisms using TaqMan method and assessed their associations with RCC risk in a case-control study of 710 patients with histologically confirmed RCC and 760 cancer-free controls. Normal renal tissues adjacent to tumors were used to evaluate the functional consequences of these polymorphisms. We found that rs7105934 was significantly associated with reduced RCC risk [adjusted odds ratio (OR) = 0.67, 95% confidence intervals (CIs) = 0.47-0.95, GA+AA versus GG], particularly among subgroups of normal-weight individuals (OR = 0.51, 95%CI = 0.29-0.88), never-smokers (OR = 0.53, 95%CI = 0.33-0.85) and non-drinkers (OR = 0.57, 95%CI = 0.370.87). Furthermore, the rs7105934 GA genotype was associated with lower levels of CCND1 mRNA compared with GG genotype, although this association was only marginally significant (P = 0.055). No significant association between rs7579899 or rs7105934 and RCC risk was observed. Our results suggest that rs7105934 on 11q13.3 may confer susceptibility to RCC in our population. Large population-based prospective and functional studies are required to validate the associations between these loci and RCC risk.

Topics: Asian People; Basic Helix-Loop-Helix Transcription Factors; Carcinoma, Renal Cell; Case-Control Studies; Chromosomes, Human, Pair 11; Cyclin D1; Female; Genetic Association Studies; Genetic Predisposition to Disease; Humans; Kidney Neoplasms; Male; Middle Aged; Polymorphism, Single Nucleotide; Proto-Oncogene Proteins; Risk Factors; Scavenger Receptors, Class B; Sequence Analysis, DNA; Transcription, Genetic

Widespread functions of the c-myc pathway play a crucial role in renal cell carcinoma (RCC) carcinogenesis. Thus, we evaluated the connection between proto-oncogenic c-myc and anti-neoplastic hsa-let-7a (let-7a) in RCC cell lines. The levels of c-myc and let-7a in 3 RCC cell lines (769P, Caki-1 and 786O) were measured after transfecting the cells with let-7a mimics or a negative control. The change in c-myc protein level was confirmed by Western blot. The anti-neoplastic function of let-7a was evaluated using cell counting kit-8 (CCK-8) for proliferation analysis and cell flow cytometry for cell cycle analysis. The changes of downstream targets of c-myc were measured using reverse transcription quantitative real-time PCR (qRT-PCR). Our results suggest for the first time that let-7a acts as a tumor suppressor in RCC cell lines by down-regulating c-myc and c-myc target genes such as proliferating cell nuclear antigen (PCNA), cyclin D1 (CCND1) and the miR17-92 cluster, which is accompanied by proliferation inhibition and cell cycle arrest.

Topics: Carcinoma, Renal Cell; Cell Cycle; Cell Line, Tumor; Cyclin D1; Down-Regulation; Gene Expression Regulation, Neoplastic; Genes, Tumor Suppressor; Humans; Kidney Neoplasms; MicroRNAs; Proliferating Cell Nuclear Antigen; Proto-Oncogene Proteins c-myc; RNA, Long Noncoding

Hypoxia-inducible factors, HIF-1α and HIF-2α, are expressed in the majority of clear-cell renal cell carcinoma (CC-RCC). In vitro, HIFα isoforms regulate a differential set of genes, and their effects in vivo within CC-RCC tumours may affect outcome. The role of angiogenesis and HIFα transcriptional products, including those involved in cell metabolism and morphological dedifferentiation have not been extensively investigated and might have relevance to the development of antiangiogenic or anti-HIFα trials in primary CC-RCC, either before or after radical nephrectomy. We analysed 168 consecutive clear-cell renal tumours from 1983 to 1999 within tissue microarrays and assessed expression of HIF-1α and HIF-2α together with the protein expression of seven of their target genes (BNIP3, CA9, Cyclin D1, GLUT-1, LDH5, Oct-4 and VEGF). The expression of these factors was compared with patient overall survival and CD31 angiogenesis. We found that HIFα antigenicity deteriorated with the age of the paraffin block (P < 0.0001) and in tumours from 1983 to 1992 was deemed not to be reliable. Similar findings were found in aged archival osteosarcoma samples. This might have important implications for retrospective biomarker studies that rely on archival tissue material. HIF-1α(HIGH)/HIF-2α(LOW) tumours had a worse overall survival compared with HIF-1α(LOW)/HIF-2α(LOW) tumours (P = 0.04). Surprisingly, on multivariate analysis, high levels of CD31(+) angiogenesis was shown to be an independent prognostic marker of increased overall survival (P = 0.003). We propose that better differentiation of vascular endothelium may be a reflection of a greater production of vessel stabilization factors versus pro-angiogenic factors, and therefore a less aggressive phenotype.

Topics: Adult; Aged; Aged, 80 and over; Angiogenesis Inhibitors; Basic Helix-Loop-Helix Transcription Factors; Biomarkers, Tumor; Carcinoma, Renal Cell; Cyclin D1; Humans; Hypoxia-Inducible Factor 1, alpha Subunit; Kidney Neoplasms; Middle Aged; Platelet Endothelial Cell Adhesion Molecule-1; Prognosis

Renal cell carcinoma (RCC) is a malignancy with poor prognosis. WNT/β-catenin signaling dysregulation, especially β-catenin overactivation and WNT antagonist silencing, is associated with RCC carcinogenesis and progression. However, the role of WNT ligands in RCC has not yet been determined. We screened 19 WNT ligands from normal kidney and RCC cell lines and tissues and found that WNT10A was significantly increased in RCC cell lines and tissues as compared to that in normal controls. The clinical significance of increase in WNT10A was evaluated by performing an immunohistochemical association study in a 19-year follow-up cohort comprising 284 RCC and 267 benign renal disease (BRD) patients. The results of this study showed that WNT10A was dramatically upregulated in RCC tissues as compared to that in BRD tissues. This result suggests that WNT10A, nuclear β-catenin, and nuclear cyclin D1 act as independent risk factors for RCC carcinogenesis and progression, with accumulative risk effects. Molecular validation of cell line models with gain- or loss-of-function designs showed that forced WNT10A expression induced RCC cell proliferation and aggressiveness, including higher chemoresistance, cell migration, invasiveness, and cell transformation, due to the activation of β-catenin-dependent signaling. Conversely, WNT10A siRNA knockdown decreased cell proliferation and aggressiveness of RCC cells. In conclusion, we showed that WNT10A acts as an autocrine oncogene both in RCC carcinogenesis and progression by activating WNT/β-catenin signaling.

Topics: Adult; beta Catenin; Carcinoma, Renal Cell; Cell Line, Tumor; Cell Nucleus; Cell Proliferation; Cell Transformation, Neoplastic; Cyclin D1; Disease Progression; Female; Follow-Up Studies; Gene Expression Regulation, Neoplastic; Humans; Kidney; Kidney Neoplasms; Male; Middle Aged; Renal Insufficiency; Risk Factors; RNA, Small Interfering; Signal Transduction; Wnt Proteins

Hepatocyte cell adhesion molecule (hepaCAM) encodes a generally inactive phosphorylated glycoprotein which mediates cancer cell proliferation, migration, and differentiation. We have reported that hepaCAM is down-regulated in renal cell carcinoma (RCC) and takes responsibility of cell growth inhibition. However, the precise mechanisms of hepaCAM inhibits cell growth is still unknown. In this study, we demonstrated that re-expression of hepaCAM can cause an accumulation in G0/G1 phase in 786-0 cells. This reaction was accompanied by a substantial reduction of c-Myc expression through using an ectopic hepaCAM expression system. Furthermore, we found a comparable decrease in proliferation and G0/G1 accumulation of 786-0 and RC-2 cells after treatment with a small molecule c-Myc inhibitor, 10058-F4. This indicated that the down regulation of c-Myc was an essential process in controlling growth inhibitory actions of hepaCAM. Nevertheless, re-expression of hepaCAM results in apparent reduction of c-Myc protein with no corresponding reduction of c-Myc mRNA. This suggests that this reaction might take place at a post-transcriptional level rather than transcriptional one. Consistent with these findings, hepaCAM decreased c-Myc stability by increasing the proportion of c-Myc phosphorylation on T58 which can be abrogated by a proteasomal inhibitor (MG132). Thus, our research implies that the decrease in c-Myc protein expression, resulting from ectopic expression of hepaCAM, may contribute to the inhibition of proliferation in these cells.

Topics: Blotting, Western; Carcinoma, Renal Cell; Cell Cycle; Cell Cycle Proteins; Cell Line, Tumor; Cell Proliferation; Cyclin D1; G1 Phase; Humans; In Vitro Techniques; Protein Stability; Proteins; Proto-Oncogene Proteins c-myc; Real-Time Polymerase Chain Reaction; Reverse Transcriptase Polymerase Chain Reaction; Thiazoles

In this report, we review our series of patients with pT3a clear cell renal carcinoma (CCRC) and comment on their outcome.. We have reviewed 260 cases of CCRC operated in the Móstoles General Hospital, Madrid, between 2000 and 2004. We have found 30 cases with pT3a tumors. Eleven of them were invading the perinephric fat, nine were invading the renal sinus fat and ten were pT3a locally but showed metastasis at the moment of diagnosis (cM1, TNM stage IV). We have analyzed the prognostic influence of histopathological parameters (vascular invasion, size, Fuhrman grade) and also immunohistochemical ones (p53, cyclin D1, proliferation index with Ki67, bcl-2 and vascular density with CD34).. Only six of 10 patients with perinephric fat involvement died of disease compared with all the patients with sinus fat involvement, suggesting a worse prognosis for the latter. However, this difference did not reach statistical significance, probably due to the small number of cases. Of all the clinical, histological and immunohistochemical factors analyzed, only cyclin D1 was a strong indicator of worse prognosis in pT3a CCRC (p=0.02). We could not show any statistically significant relation between vascular density and prognosis. Vascular invasion was the only histological parameter that showed a trend toward significance (p=0.09).. Sinus fat involvement might be underestimated in some series. A protocol for nephrectomy specimen handling could improve the detection rate of sinus fat involvement and allow the performance of randomized prospective studies to determine whether these tumors behave similarly.

Topics: Antigens, CD34; Carcinoma, Renal Cell; Cyclin D1; Humans; Immunohistochemistry; Kaplan-Meier Estimate; Kidney Neoplasms; Lipid Metabolism; Neoplasm Invasiveness; Neoplasm Staging; Nephrons; Tumor Suppressor Protein p53

The loss of tuberin, the tuberous sclerosis-2 (Tsc-2) gene product, is associated with cytoplasmic mislocalization of p27 in uterine leiomyomas derived from Eker rats (Tsc-2(EK/+)) and in human metastatic renal cell carcinoma tissue. Signaling associated with cytoplasmic mislocalization of p27 in renal cancer is relatively unknown. Renal tumors derived from 2,3,5-tris-(glutathion-S-yl)hydroquinone (TGHQ)-treated Tsc-2(EK/+) rats, and null for tuberin, display elevated nuclear and cytosolic p27, with parallel increases in cytosolic cyclin D1 levels. Similar changes are observed in TGHQ-transformed renal epithelial cells derived from Tsc-2(EK/+) rats (QTRRE cells), which, in addition to the cytoplasmic mislocalization of p27 and cyclin D1, exhibit high ERK, B-Raf, and Raf-1 kinase activity. Renal tumor xenografts, derived from subcutaneous injection of QTRRE cells into nude mice, also display increases in cytosolic mislocalization of p27 and cyclin D1. Dibutyryl cAMP and/or phosphodiesterase inhibitors (PIs; pentoxifylline or theophylline) increase Rap1B activation, B-Raf kinase activity, and cytosolic p27/cyclin D1 protein levels in QTRRE cells. Inhibition of Raf kinases with either sorafenib or B-Raf small interfering RNA (siRNA) caused a mitogen-activated protein kinase-mediated downregulation of p27. Moreover, decreases in cyclin D1 were also associated with p27 siRNA knockdown in QTRRE cells. Finally, theophylline-mediated increases in p27 and cyclin D1 were attenuated by sorafenib, which modulated Raf/MEK/ERK signaling. Collectively, these data suggest that the cAMP/Rap1B/B-Raf pathway modulates the expression of p27 and the cytoplasmic mislocalization of p27-cyclin D1 in tuberous sclerosis gene-regulated-renal cancer. Therefore, the loss of tuberin and engagement of the cAMP pathway may independently direct p27-cyclin D1 cytosolic stabilization during renal tumor formation.

Topics: Animals; Benzenesulfonates; Bucladesine; Carcinoma, Renal Cell; Cell Line; Cyclin D1; Cyclin-Dependent Kinase Inhibitor p27; Cytosol; Down-Regulation; Female; Gene Expression Regulation, Neoplastic; Glutathione; Humans; Hydroquinones; Kidney Neoplasms; Male; Mice; Mice, Nude; Mitogen-Activated Protein Kinases; Niacinamide; Pentoxifylline; Phenylurea Compounds; Phosphodiesterase Inhibitors; Proto-Oncogene Proteins B-raf; Proto-Oncogene Proteins c-raf; Pyridines; Rats; RNA, Small Interfering; Signal Transduction; Sorafenib; Theophylline; Tuberous Sclerosis; Tuberous Sclerosis Complex 2 Protein; Tumor Suppressor Proteins

YueF is a novel putative tumor suppressor gene that can inhibit proliferation and induce apoptosis in hepatoma cells, but its role in renal cell carcinoma (RCC) remains unclear. Here, we examined the expression of the YueF gene in RCC tissues and the effect of YueF on cell proliferation in RCC 786-0 cells. The results showed that YueF was expressed at high levels in normal kidney tissues and cell lines but was reduced or absent in RCC tissues and 786-0 cells. Lentivirus-mediated YueF overexpression in RCC 786-0 cells caused cell-cycle arrest in the G1 phase and dramatically reduced proliferation in culture. YueF overexpression resulted in increased protein levels of p53 and p21(WAF1/Cip1), whereas the protein levels of cyclin D1 and pRb were decreased. The proliferation defects caused by YueF overexpression could be partially rescued by the expression of p21 siRNA. These findings suggest a critical role for p21 in the YueF-induced growth inhibition of 786-0 cells and provide novel insights into the mechanism underlying the tumor-suppressive action of YueF.

Topics: Carcinoma, Renal Cell; Cell Proliferation; Cyclin D1; Cyclin-Dependent Kinase Inhibitor p21; Down-Regulation; Female; G1 Phase Cell Cycle Checkpoints; Humans; Male; Retinoblastoma Protein; RNA Interference; RNA, Small Interfering; Tumor Cells, Cultured; Tumor Suppressor Protein p53; Tumor Suppressor Proteins; Up-Regulation

The mammalian target of rapamycin (mTOR) and mitogen-activated protein kinase signaling cascades have been implicated in a number of human cancers. The tumor suppressor gene tuberous sclerosis-2 (Tsc-2) functions as a negative regulator of mTOR. Critical proteins in both pathways are activated following treatment of Eker rats (Tsc-2(EK/+)) with the nephrocarcinogen 2,3,5-tris-(glutathion-S-yl)hydroquinone (TGHQ), which also results in loss of the wild-type allele of Tsc-2 in renal preneoplastic lesions and tumors. Western blot analysis of kidney tumors formed following treatment of Tsc-2(EK/+) rats with TGHQ for 8 months revealed increases in B-Raf, Raf-1, pERK, cyclin D1, 4EBP1, and p-4EBP1-Ser65, -Thr70, and -Thr37/46 expression. Similar changes are observed following TGHQ-mediated transformation of primary renal epithelial cells derived from Tsc-2(EK/+) rats (quinol-thioether rat renal epithelial [QTRRE] cells) that are also null for tuberin. These cells exhibit high ERK, B-Raf, and Raf-1 kinase activity and increased expression of all p-4EBP1s and cyclin D1. Treatment of the QTRRE cells with the Raf kinase inhibitor, sorafenib, or the MEK1/2 kinase inhibitor, PD 98059, produced a significant decrease in the protein expression of all p-4EBP1s and cyclin D1. Following siRNA knockdown of Raf-1, Western blot analysis revealed a significant decrease in Raf-1, cyclin D1, and all p-4EBP1 forms noted above. In contrast, siRNA knockdown of B-Raf resulted in a nominal change in these proteins. The data indicate that Raf-1/MEK/ERK participates in crosstalk with 4EBP1, which represents a novel pathway interaction leading to increased protein synthesis, cell growth, and kidney tumor formation.

Topics: Animals; Carcinoma, Renal Cell; Carrier Proteins; Cell Culture Techniques; Cell Line; Cell Transformation, Neoplastic; Cyclin D1; Epithelial Cells; Extracellular Signal-Regulated MAP Kinases; Gene Expression Regulation, Neoplastic; Glutathione; Humans; Hydroquinones; Immunohistochemistry; Intracellular Signaling Peptides and Proteins; Kidney Neoplasms; Loss of Heterozygosity; Male; MAP Kinase Kinase Kinases; Phosphoproteins; Protein Biosynthesis; Proto-Oncogene Proteins B-raf; Proto-Oncogene Proteins c-raf; Rats; Rats, Mutant Strains; Receptor Cross-Talk; RNA, Small Interfering; Signal Transduction; Tuberous Sclerosis Complex 2 Protein; Tumor Suppressor Proteins

Renal cell carcinoma (RCC) is a rare tumor in the pediatric population. Recently, a phenotypically and genetically distinct kidney carcinoma, mainly prevalent in children and associated with an Xp11.2 translocation or TFE3 gene fusion, has been described. It has been advanced that in this subtype of RCC, there is an accumulation of cyclin D1, cyclin D3, and p21 ((wafl/cip1)). The aim of the present study was to figure out in two pediatric RCC recently diagnosed in our department (one clear cell-type RCC and one TFE3-positive RCC) whether those features are indeed specific of the latter tumor or occur in pediatric RCC irrespective of the tumor type. The following immunostains were performed in both cases: Ki67, p16(ink4a), p21 ((wafl/cip1)), p27(kip1), p53, p63, mdm2, cyclin D1, cyclin D3, TFE3, CD10, vimentin, E-cadherin, and RCC-antigen. We observed in the TFE3-positive carcinoma an intense immunoreaction for p21 ((wafl/cip1)), cyclin D1, and cyclin D3, without expression for p53, p16, p27(kip1), and mdm2, whereas the immunoexpression profile observed in the classic RCC was similar to that of clear cell, adult-type RCC. Our study confirms that TFE3-positive RCC exhibits a deregulation of the cell cycle apparently unrelated to the young age of the patients.

Topics: Antineoplastic Combined Chemotherapy Protocols; Basic Helix-Loop-Helix Leucine Zipper Transcription Factors; Carcinoma, Renal Cell; Cell Cycle Proteins; Child; Chromosomes, Human, X; Cyclin D1; Cyclin D3; Cyclin-Dependent Kinase Inhibitor p21; Cyclins; Female; Gene Expression Regulation, Neoplastic; Genotype; Humans; Immunohistochemistry; In Situ Hybridization, Fluorescence; Infant; Kidney Neoplasms; Nephrectomy; Phenotype; Translocation, Genetic; Treatment Outcome

The lack of effective anti-tumor therapy for renal cell carcinoma (RCC) has stimulated the search for novel target whose inhibition could block tumorigenesis. Recently, reduced DLC-1 has been shown to be associated with aggressive and highly metastatic renal cell carcinoma. In this study, the biological role of DLC-1 on cell growth, migration and cell cycle progression in RCC cells was investigated. Over-expression of DLC-1 was associated with a marked inhibition of cell growth (P<0.01). The inhibitory effect was partly due to the induction of apoptosis and cell cycle arrest in G(0)/G(1) accompanied by up-regulation of the intracellular signal proteins of p27 and down-regulation of cyclin D1 and cyclin E. Furthermore, DLC-1 induced FAK dephosphorylation of focal adhesion proteins inhibited cell migration (P<0.05). Decreased DLC-1 expression strongly correlated with proliferative activity, as indicated by the elevated levels of Ki67. Restoration of DLC-1 expression in RCC cells led to Bcl-2 and caspase-3 mediated apoptosis as well as attenuated the ability of the cells to form RCC tumors in athymic nude mice (P<0.05). Taken together, these results suggest that DLC-1 plays a crucial role in signal transduction pathway regulating the cell proliferation, migration, and carcinogenesis of human RCC.

Topics: Animals; Apoptosis; Blotting, Western; Carcinoma, Renal Cell; Caspase 3; Cell Line, Tumor; Cell Movement; Cell Proliferation; Cyclin D1; Cyclin E; Flow Cytometry; Focal Adhesion Kinase 1; GTPase-Activating Proteins; Humans; Immunohistochemistry; Kidney Neoplasms; Mice; Mice, Nude; Reverse Transcriptase Polymerase Chain Reaction; Signal Transduction; Transfection; Tumor Suppressor Proteins; Xenograft Model Antitumor Assays

Renal oncocytoma is a benign tumor occurring singly or as multiple synchronous lesions. The histologic features of renal oncocytoma may overlap with those of chromophobe renal cell carcinoma. Chromosomal translocations involving the CCND1 locus at 11q13 and overexpression of cyclin D1 occur in a subset of renal oncocytomas. We evaluated a series of 63 renal oncocytomas and 36 chromophobe renal cell carcinomas and assessed the clinical features, cyclin D1 overexpression by immunohistochemistry, and alterations of the CCND1 gene by fluorescence in situ hybridization. All 36 chromophobe renal cell carcinomas were negative for cyclin D1 overexpression and alterations of CCND1. Of the 63 renal oncocytomas, 21 (33%) showed cyclin D1 overexpression. Of 21 renal oncocytomas with cyclin D1 overexpression, a CCND1 rearrangement was detected in 12 (57%). A CCND1 rearrangement was also identified in 1 (2%) of the 42 renal oncocytomas without cyclin D1 overexpression. Of 42 renal oncocytomas without cyclin D1 overexpression, 16 (38%) were from patients with multiple renal oncocytomas at nephrectomy. Of 21 renal oncocytomas with cyclin D1 overexpression, only 1 (5%) patient had multiple renal oncocytomas (P = .006). Of the 25 patients whose original tumor showed no cyclin D1 overexpression, 8 (32%) developed a subsequent renal oncocytoma. None of 15 patients whose original tumor showed cyclin D1 overexpression had a subsequent renal oncocytoma (P = .016). The findings of this study suggest that renal oncocytomas lacking cyclin D1 overexpression may be associated with the development of multiple renal oncocytomas and that these patients are more likely to develop subsequent renal oncocytomas suggesting the need for more frequent clinical for these patients and little need for follow-up in patients with renal oncocytomas overexpressing cyclin D1. The data also show that cyclin D1 overexpression and CCND1 rearrangements by fluorescence in situ hybridization are absent in chromophobe renal cell carcinoma, suggesting that these are useful when differentiating between renal oncocytoma and chromophobe renal cell carcinoma.

Topics: Adenoma, Oxyphilic; Carcinoma, Renal Cell; Case-Control Studies; Chromosomes, Human, Pair 11; Cyclin D1; Cytogenetic Analysis; Diagnosis, Differential; Female; Gene Rearrangement; Humans; Immunohistochemistry; In Situ Hybridization, Fluorescence; Karyotyping; Kidney Neoplasms; Male; Nephrectomy; Translocation, Genetic; Tumor Cells, Cultured

Coactivator activator (CoAA) has been reported to be a coactivator that regulates steroid receptor-mediated transcription and alternative RNA splicing. Herein, we show that CoAA is a dual-function coregulator that inhibits G(1)-S transition in human kidney cells and suppresses anchorage-independent growth and xenograft tumor formation. Suppression occurs in part by down-regulating c-myc and its downstream effectors ccnd1 and skp2 and causing accumulation of p27/Kip1 protein. In this cellular setting, CoAA directly represses the proto-oncogene c-myc by recruiting HDAC3 protein and decreasing both the acetylation of histone H3 and the presence of RNA polymerase II on the c-myc promoter. Interestingly, a splicing isoform of CoAA, coactivator modulator (CoAM), antagonizes CoAA-induced G(1)-S transition and growth inhibition by negatively regulating the mRNA levels of the endogenous CoAA isoform. In addition, we found that expression of CoAA protein is significantly decreased in human renal cell carcinoma compared with normal kidney. Our study presents evidence that CoAA is a potential tumor suppressor in renal carcinoma and that CoAM is a counterbalancing splice isoform. This is, thus far, the only example of a nuclear receptor coregulator involved in suppression of kidney cancer and suggests potentially significant new roles for coregulators in renal cancer biology.

Topics: Animals; Carcinoma, Renal Cell; Cell Cycle; Cell Line; Cell Proliferation; Cell Transformation, Neoplastic; Cyclin D1; Gene Expression Regulation, Neoplastic; Genes, myc; Genes, Tumor Suppressor; Humans; Intracellular Signaling Peptides and Proteins; Kidney; Kidney Neoplasms; Mice; Mice, Nude; Models, Biological; Protein Isoforms; Proto-Oncogene Mas; S-Phase Kinase-Associated Proteins; Transplantation, Heterologous

To simultaneously analyze multiple biologic markers to identify strong prognostic markers for disease-specific survival of patients with clear cell renal cell carcinoma (ccRCC).. The expression of Ki-67, p53, bcl-2, cyclin-D1, caveolin-1, vascular endothelial growth factor, and HER-2 was evaluated in 119 paraffin-embedded ccRCC specimens using immunohistochemistry. The clinical significance of these markers in relation to disease-specific survival was analyzed.. On univariate analysis, high-level staining for Ki-67 (P <0.0001), p53 (P = 0.0029), vascular endothelial growth factor (P = 0.0062), and caveolin-1 (P = 0.0396) was associated with decreased survival, but high-level staining for bcl-2 (P <0.0001) and cyclin-D1 (P = 0.0002) was associated with increased survival. Only HER-2 expression was not related to survival (P = 0.1131). Multivariate analysis revealed the following independent predictors of disease-specific survival: expression of p53 (P = 0.0059) or bcl-2 (P = 0.0413) in all cases of ccRCC; expression of p53 (P = 0.0043) or bcl-2 (P = 0.0227) in cases of grade 1-2 disease; and expression of p53 (P = 0.0207) in cases with metastasis at surgery.. Of the seven markers reviewed, p53 and bcl-2 were strong prognostic factors in all cases and in cases of grade 1-2 ccRCC. Only p53 attained independent prognostic significance in metastatic ccRCC. This information could prove useful in selecting markers to predict for survival and plan therapy for patients with ccRCC.

Topics: Adult; Age Factors; Aged; Aged, 80 and over; Analysis of Variance; Antibodies, Neoplasm; Biomarkers, Tumor; Biopsy, Needle; Carcinoma, Renal Cell; Cyclin D1; Female; Genes, p53; Humans; Immunohistochemistry; Ki-67 Antigen; Kidney Neoplasms; Male; Middle Aged; Neoplasm Staging; Nephrectomy; Probability; Prognosis; Proportional Hazards Models; Proto-Oncogene Proteins c-bcl-2; Retrospective Studies; Risk Assessment; Sampling Studies; Sensitivity and Specificity; Survival Analysis

To investigate the role of cyclin D1 and p27(kip1) in the occurrence and development of conventional renal cell carcinoma(CRCC).. RT-PCR and Western-blot were used to detect mRNA and protein contents of cyclin D1 and p27(kip1) in 25 CRCCs and 10 normal renal tissue distant to tumor. Immunohistochemistry was used to investigate the expression of cyclin D1 and p27(kip1) in pathological tissue sections of 76 CRCCs. The relationship between those index and clinicopathological parameters was analyzed statistically.. In CRCC, the expression of cyclin D1 was higher than that of the control group. The higher cyclin D1 content was related to big tumor size (P<0.05); The expression of p27(kip1) was lower than that of the control group, and the lower p27(kip1) was related to higher nuclear grade and TNM stage (P<0.01). The 5-year cumulative survival rate of the p27(kip1) high expression group is longer than that of the low group (P<0.01).. The excessive expression of cyclin D1 and lower expression of p27(kip1) play an important role in the carcinogenesis of CRCC. The lower expression of p27(kip1) may affect the progression of the tumors. The detection of p27(kip1) may be as a reference marker in the prognosis of CRCC.

Topics: Adolescent; Adult; Aged; Aged, 80 and over; Blotting, Western; Carcinoma, Renal Cell; Cyclin D1; Cyclin-Dependent Kinase Inhibitor p27; Female; Gene Expression Regulation, Neoplastic; Humans; Kidney Neoplasms; Male; Middle Aged; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger

The mutation of von Hippel-Lindau (VHL) tumor suppressor gene is closely related with tumorigenesis of renal clear cell carcinoma (RCCC). Cyclin D1 gene plays an important role in progression of RCCC by stimulating cell proliferation. This study was to determine the mutation of VHL gene in RCCC, and explore its correlation to overexpression of Cyclin D1.. The specimens of RCCC and adjacent normal renal tissue from 50 patients were collected after surgery. Total RNA and genomic DNA were extracted from each sample. Variant exons in VHL gene were amplified by polymerase chain reaction (PCR) and sequenced, and DNA hypermethylation was detected by restriction analysis. Reverse transcription-PCR (RT-PCR) and Western blot were used to detect the expression of Cyclin D1.. Of the 50 specimens, 42 (84.0%) had various VHL gene mutations, 12 (24.0%) had more than 1 kind of gene mutation. Of the 57 cases of exon mutation of VHL gene, 17 (29.8%) were located in exon 1, 26 (45.6%) in exon 2, and 14 (24.6%) in exon 3. The expression of Cyclin D1 in the 42 cases with VHL gene mutation was increased to 2-10 (3.91+/-1.54) times that of normal controls (P<0.01). Cyclin D1 expression in the other 8 cases was normal.. There are variant mutations of VHL gene in RCCC, which may lead to overexpression of Cyclin D1.

Topics: Carcinoma, Renal Cell; Cyclin D1; Exons; Gene Expression Regulation, Neoplastic; Humans; Kidney Neoplasms; Mutation; RNA, Messenger; Von Hippel-Lindau Tumor Suppressor Protein

The L1 cell adhesion molecule is overexpressed in many human carcinomas. The objectives of the study were to provide a comprehensive description of L1 distribution in human kidney and to establish the prognostic relevance of L1 expression in renal cell carcinomas (RCC).. Using two antibodies to the extracellular part and the cytoplasmic domain, respectively, we first compared L1 expression in normal kidney and renal tumors of diverse histopathologic origin, then we studied L1 expression together with tumor stage, grade, molecular prognostic biomarkers, and metastatic behavior.. In normal kidney, L1 immunoreactive with both antibodies was expressed in all epithelial cells originating from the ureteric bud except for intercalated cells. In renal tumors, L1 was mainly detected in those originating from cells that do not express L1 in the normal kidney [i.e., 33 of 72 clear cell RCC (ccRCC) and 25 of 88 papillary RCC (papRCC)]. Both in ccRCC and papRCC, L1 reacted only with the antibody to the extracellular domain, suggesting that the protein was truncated. In these carcinomas, L1 expression was strongly correlated with Ki-67 proliferation index (ccRCC, P = 0.0059; papRCC, P = 0.0039), but only in ccRCC, the presence of L1 was associated with the risk of metastasis (P = 0.0121). This risk was higher if cyclin D1 was concurrently absent in tumor cells (P < 0.0001). The L1(+)/cyclin D1(-) profile was an independent prognostic factor of metastasis occurrence in multivariate analysis (P = 0.0023).. We have found a combination of markers that can serve to identify a subgroup of high-risk patients with ccRCC that may require more aggressive therapies.

Topics: Adenocarcinoma, Clear Cell; Adolescent; Adult; Aged; Aged, 80 and over; Carcinoma, Renal Cell; Cyclin D1; Female; Gene Expression Regulation, Neoplastic; Humans; Immunohistochemistry; In Situ Hybridization; Ki-67 Antigen; Kidney; Kidney Neoplasms; Male; Middle Aged; Neoplasm Metastasis; Neural Cell Adhesion Molecule L1; Prognosis; RNA, Messenger; Survival Analysis

To investigate the effect of saponins from Tribulus terrestris (STT) on the renal carcinoma cell (786-0) in vitro, and inhibitory mechanisms.. Effects of SIT on the cytotoxicity, morphological changes of apoptosis, cell cycle and expression of Bcl-2 protein in the 786-0 were tested respectively by MTT method, Wright and acridine orange stain assay, as well as flow cytometry (FCM).. After the 786-0 was treated by STY, it was shown that: 1) A significant cytotoxic effect was observed by MTT assay; 2) Apoptosis-induced was viewed by Wright and acridine orange stain assay; 3) The distribution of 786-0 on S phase was increased; 4.) The expression of Bcl-2 protein and cyclin D1 was decreased.. STT can significantly inhibit the growth of 786-0 in vitro, partially, by apoptosis.

Topics: Antineoplastic Agents, Phytogenic; Apoptosis; Carcinoma, Renal Cell; Cell Line, Tumor; Cell Proliferation; Cyclin D1; Drugs, Chinese Herbal; Humans; Kidney Neoplasms; Plants, Medicinal; Proto-Oncogene Proteins c-bcl-2; S Phase; Saponins; Tribulus

We report the case of a 43-year-old male with multiple tumor foci showing microscopic features of chromophobe renal carcinoma (ChRCC) arising in an oncocytoma. Conventional cytogenetics of fresh tumor cells and fluorescence in situ hybridization (FISH) revealed the following abnormal karyotype: 46,XY,der(8)ins(8;11)(p?;q13),der(11)ins(8;11)inv(11)(q12?p15) with CCND1 (11q13) rearrangement. To our knowledge, chromosome 8 has not been reported as a partner involved in structural rearrangements of 11q13 in oncocytomas. FISH in paraffin tissue sections revealed a rearrangement of CCND1 (11q13) in the oncocytoma cells. The multiple foci of chromophobe carcinoma presented multiple copies of CCND1, suggesting that they represented a transformation from oncocytoma into ChRCC. There was immunohistochemical overexpression of CCND1 in both oncocytoma and chromophobe carcinoma cells. In this case, the correlation of the microscopic findings with changes in CCND1 gene associated to CCND1 overexpression in both components suggest that the ChRCC would have originated from the preexisting oncocytoma. It is not possible to detect, by cytogenetic techniques alone, if the ChRCC component have also the CCND1 rearrangement in addition to the detected polysomy. FISH techniques on paraffin tissue sections may help to identify genetic aberrations such as CCND1 rearrangement in order to establish a diagnosis of oncocytoma.

Topics: Adenoma, Oxyphilic; Adult; Carcinoma, Renal Cell; Cell Transformation, Neoplastic; Chromosome Banding; Chromosomes, Human, Pair 11; Chromosomes, Human, Pair 8; Cyclin D1; Humans; Immunohistochemistry; In Situ Hybridization, Fluorescence; Karyotyping; Kidney Neoplasms; Male

The retinoblastoma gene product (pRb) is the main substrate for cyclin-dependent kinases (CDKs) during the G1/S transition. Aberrations in cell cycle regulatory proteins, which have been observed in many malignancies, can theoretically cause increased phosphorylation of pRb due to unbalanced CDK activities. The expression and phosphorylation of pRb and potential associations to cell cycle aberrations in renal cell carcinomas (RCC) has only partly been clarified. We therefore evaluated the presence of pRb and the level of pRb-phosphorylation in 216 RCCs arranged in tissue microarrays by using different pRb-antibodies, including pRb-phosphospecific antibodies. Most RCCs (95%) expressed pRb, while cases with the low pRb levels, potentially indicative for pRb-inactivation, were few. In order to detect secondary alterations to a potential pRb-inactivation, the p16 expression was also monitored. None of the tumors exhibited increased p16 levels, confirming that pRb-inactivation is rare in RCC. Phosphorylated pRb was detected in approximately 50% of the RCCs, using Western blotting or immunohistochemistry. The immunohistochemical ppRb(ser807/811) levels were associated with high proliferation, cyclin D1, cyclin E and p27 protein content. Surprisingly, there was no association between pRb-phosphorylation and clinicopathological data. In summary, pRb seemed to be functional and aberrations in G1/S-regulatory proteins were associated with increased phosphorylation of pRb and proliferation. The data supports that pRb might be one of the main cell cycle regulators in RCC.

Topics: Adult; Aged; Aged, 80 and over; Carcinoma, Renal Cell; Cell Cycle Proteins; Cyclin D1; Cyclin E; Cyclin-Dependent Kinase Inhibitor p16; Cyclin-Dependent Kinase Inhibitor p21; Cyclin-Dependent Kinase Inhibitor p27; Cyclins; DNA, Neoplasm; Female; G1 Phase; Genes, Tumor Suppressor; Humans; Immunoenzyme Techniques; In Situ Hybridization, Fluorescence; Kidney Neoplasms; Male; Middle Aged; Phosphorylation; Ploidies; Retinoblastoma Protein; S Phase; Tumor Suppressor Proteins

Gene expression analysis was performed on a human renal cancer cell line (786-0) with mutated VHL gene and a transfectant with wild-type VHL to analyse genes regulated by VHL and to compare with the gene programme regulated by hypoxia. There was a highly significant concordance of the global gene response to hypoxia and genes suppressed by VHL. Cyclin D1 was the most highly inducible transcript and 14-3-3 epsilon was downregulated. There were some genes regulated by VHL but not hypoxia in the renal cell line, suggesting a VHL role independent of hypoxia. However in nonrenal cell lines they were hypoxia regulated. These included several new pathways regulated by hypoxia, including RNase 6PL, collagen type 1 alpha 1, integrin alpha 5, ferritin light polypeptide, JM4 protein, transgelin and L1 cell adhesion molecule. These were not found in a recent SAGE analysis of the same cell line. Hypoxia induced downregulation of Cyclin D1 in nonrenal cells via an HIF independent pathway. The selective regulation of Cyclin D1 by hypoxia in renal cells may therefore contribute to the tissue selectivity of VHL mutation.

Topics: Carcinoma, Renal Cell; Cell Hypoxia; Cyclin D1; Gene Expression Profiling; Humans; Kidney Neoplasms; Tumor Cells, Cultured; Tumor Suppressor Proteins; Ubiquitin-Protein Ligases; Von Hippel-Lindau Tumor Suppressor Protein

Topics: Carcinoma, Renal Cell; Chromosome Aberrations; Cyclin D1; Disease Progression; Disease-Free Survival; Humans; Kidney; Kidney Neoplasms; Lymphatic Metastasis; Neoplasm Invasiveness; Neoplasm Recurrence, Local; Neoplasm Staging; Neoplastic Cells, Circulating; Prognosis; Proliferating Cell Nuclear Antigen; RNA, Messenger; Vascular Endothelial Growth Factor A

Previously, we showed that arsenic trioxide potently inhibited the growth of myeloma cells and head and neck cancer cells. Here, we demonstrate that arsenic trioxide inhibited the proliferation of all the renal cell carcinoma cell lines (ACHN, A498, Caki-2, Cos-7, and Renca) except only one cell line (Caki-1) with IC(50) of about 2.5-10 microM. Arsenic trioxide induced a G(1) or a G(2)-M phase arrest in these cells. When we examined the effects of this drug on A498 cells, arsenic trioxide (2.5 microM) decreased the levels of CDK2, CDK6, cyclin D1, cyclin E, and cyclin A proteins. Although p21 protein was not increased by arsenic trioxide, this drug markedly enhanced the binding of p21 with CDK2. In addition, the activities of CDK2- and CDK6-associated kinase were reduced in association with hypophosphorylation of Rb protein. Arsenic trioxide (10 microM) also induced apoptosis in A498 cells. Apoptotic process of A498 cells was associated with the changes of Bcl-(XL), caspase-9, caspase-3, and caspase-7 proteins as well as mitochondria transmembrane potential (Deltapsi(m)) loss. Taken together, these results demonstrate that arsenic trioxide inhibits the growth of renal cell carcinoma cells via cell cycle arrest or apoptosis.

Topics: Animals; Apoptosis; Arsenic Trioxide; Arsenicals; bcl-X Protein; Carcinoma, Renal Cell; Caspases; CDC2-CDC28 Kinases; Cell Cycle; Cell Division; Cyclin A; Cyclin D1; Cyclin E; Cyclin-Dependent Kinase 2; Cyclin-Dependent Kinase 6; Cyclin-Dependent Kinase Inhibitor p21; Cyclin-Dependent Kinases; Cyclins; Humans; Kidney Neoplasms; Oxides; Protein Serine-Threonine Kinases; Proto-Oncogene Proteins c-bcl-2; Retinoblastoma Protein; Tumor Cells, Cultured

Loss of the von Hippel-Lindau gene (VHL) expression ca-uses deregulation of contact inhibition of cell growth, which might be one of the bases of the tumor suppressor function of VHL. Here we show that this function of the VHL gene product (pVHL) depends on cell autonomous events. To identify the target gene of pVHL, which is directly involved in the contact inhibition, we compared the gene expression profile between VHL-deficient renal carcinoma 786-O cells and those infected with an adenovirus vector encoding VHL. In addition to known pVHL-regulated genes, such as vascular endothelial growth factor and carbonic anhydrase, we found cyclinD1 as a new target of pVHL at a high cell density. In VHL-expressing cells (VHL (+) cells), the cyclinD1 mRNA expression level diminishes at a high cell density, while it remains at a relatively high level in VHL-deficient cells (VHL (-) cells). The cyclinD1 expression level was also abnormally high in VHL (-) cells at a high cell density. Consequently, the phosporylation level of the retinoblastoma (Rb) protein remained high in these cells, whereas there was no phosporylated Rb in VHL (+) cells under the contact inhibition. The abnormal expression of cyclinD1 at a high cell density was observed even in VHL (+) cells under the hypoxic state. Moreover, ectopic expression of a HIF mutant resistant to pVHL-mediated proteolysis causes the abnormal cyclinD1 expression in VHL (+) cells. Taken together, these observations indicate that VHL is required for the downregulation of cyclinD1 at a high cell density through HIF.

Topics: Carbonic Anhydrases; Carcinoma, Renal Cell; Cell Division; Cell Hypoxia; Coculture Techniques; Cyclin D1; DNA-Binding Proteins; Endothelial Growth Factors; Gene Deletion; Gene Expression Regulation, Neoplastic; Humans; Hypoxia-Inducible Factor 1; Hypoxia-Inducible Factor 1, alpha Subunit; Intercellular Signaling Peptides and Proteins; Kidney Neoplasms; Kinetics; Ligases; Lymphokines; Nuclear Proteins; Phosphorylation; Transcription Factors; Transfection; Tumor Cells, Cultured; Tumor Suppressor Proteins; Ubiquitin-Protein Ligases; Vascular Endothelial Growth Factor A; Vascular Endothelial Growth Factors; von Hippel-Lindau Disease; Von Hippel-Lindau Tumor Suppressor Protein

Aberrations in the G1/S transition of the cell cycle have been observed in many malignancies and seem to be critical in the transformation process. Few studies have delineated the presence of G1/S regulatory defects and their clinical relevance in renal cell carcinoma (RCC). Therefore, we have examined the protein contents of cyclin D1, D3, E, and p27 in 218 RCCs, using tissue microarray and immunohistochemistry. The results from a subset of tumours were confirmed by Western blotting and immunohistochemical staining of regular tissue sections. Interestingly, low protein contents of cyclin D1 and p27 were associated with high nuclear grade, large tumour size, and poor prognosis for patients with conventional tumours. We further observed substantial differences in the pattern of G1/S regulatory defects between the different RCC subtypes. The majority of both conventional and papillary cases expressed p27; however, chromophobe tumours generally lacked p27 staining. In addition, conventional RCCs often expressed high cyclin D1 protein levels, while papillary RCCs exhibited high cyclin E. In summary, we have shown that G1/S regulatory defects are present in RCC and are associated with clinico-pathological parameters. The pattern of cell cycle regulatory defects also differed between RCC subtypes.

Topics: Actuarial Analysis; Adult; Aged; Aged, 80 and over; Carcinoma, Renal Cell; Cell Cycle; Cyclin D; Cyclin D1; Cyclin E; Cyclins; Female; Humans; Immunohistochemistry; Kidney Neoplasms; Male; Microfilament Proteins; Middle Aged; Multivariate Analysis; Muscle Proteins; Oligonucleotide Array Sequence Analysis; Survival Analysis; Time Factors

Renal cell carcinoma is associated with mutation of the von Hippel-Lindau (VHL) tumor suppressor gene. Cell lines derived from these tumors cannot exit the cell cycle when deprived of growth factors, and the ability to exit the cell cycle can be restored by the reintroduction of wild-type protein VHL (pVHL). Here, we report that cyclin D1 is overexpressed and remains inappropriately high in during contact inhibition in pVHL-deficient cell lines. In addition, hypoxia increased the expression of cyclin D1 specifically in pVHL-negative cell lines into which pVHL expression was restored. Hypoxic-induction of cyclin D1 was not observed in other pVHL-positive cell lines. This suggests a model whereby in some kidney cell types, pVHL may regulate a proliferative response to hypoxia, whereas the loss of pVHL leads to constitutively elevated cyclin D1 and abnormal proliferation under normal growth conditions.

Topics: Carcinoma, Renal Cell; Cell Cycle; Cell Cycle Proteins; Cell Hypoxia; Cyclin D1; Gene Expression Regulation, Neoplastic; Humans; Kidney Neoplasms; Ligases; Transfection; Tumor Suppressor Proteins; Ubiquitin-Protein Ligases; Von Hippel-Lindau Tumor Suppressor Protein

Malignancy is a dreaded complication of organ transplantation. Immunosuppressive drug therapy-induced impairment of the organ graft recipient's immune surveillance is considered to be the mechanism for the heightened incidence and metastatic progression. We identified a cell-autonomous and host-immunity independent mechanism for cyclosporine-associated tumor progression. In this study, we investigated the effect of rapamycin on tumor progression, in the presence and absence of cyclosporine.. A spontaneously arising renal adenocarcinoma (renal cancer) of BALB/c origin was used as the model tumor. The effect of rapamycin on renal cancer cell phenotype, molecules (E-cadherin, p27 kip1, cyclin D1) implicated in tumor progression, and the effect of rapamycin on in vivo tumor progression were explored in BALB/c mice and in T-cell, B-cell, and natural killer (NK) cell-deficient severe combined immune deficiency (SCID)-beige mice. In the SCID-beige mice, T24 human bladder transitional cell carcinoma also was used as the tumor inoculum.. Rapamycin conditioning of renal cancer cells upregulated E-cadherin expression and induced phenotypic transition from invasive spindle, or dome-shaped cells, with exploratory pseudopodia to noninvasive cuboidal cells that formed cell-to-cell adhesions. Rapamycin increased p27 kip1, reduced cyclinD1, and arrested the growth of renal cancer cells in G1/S phase. In vivo, rapamycin prevented tumor growth and metastatic progression in syngeneic BALB/c or SCID-beige mice, and in BALB/c or SCID-beige mice treated with cyclosporine. Rapamycin treatment alone, or with cyclosporine, prolonged the survival of mice inoculated with renal cancer cells or T24 human bladder cancer cells.. Our findings, in addition to unlinking mechanisms of immunosuppression from that of tumor progression, suggest that rapamycin may be of value for the management of posttransplant malignancy.

Topics: Adenocarcinoma; Animals; Antibiotics, Antineoplastic; B-Lymphocytes; Cadherins; Carcinoma, Renal Cell; Cell Cycle Proteins; Cyclin D1; Cyclin-Dependent Kinase Inhibitor p27; Disease Progression; DNA Primers; Genes, Tumor Suppressor; Immunosuppression Therapy; Immunosuppressive Agents; Kidney Neoplasms; Killer Cells, Natural; Mice; Mice, Inbred BALB C; Mice, SCID; Sirolimus; T-Lymphocytes; Tacrolimus; Tumor Cells, Cultured; Tumor Suppressor Proteins

Germ-line mutations in the von Hippel-Lindau (VHL) tumor suppressor disease are associated with a high risk of retinal and cerebellar hemangioblastomas, renal cell carcinoma (RCC), and, in some cases, pheochromocytoma (PHE). In addition, somatic mutation or epigenetic inactivation of the VHL gene occurs in most clear cell RCCs. VHL protein (pVHL) has a critical role in regulating proteasomal degradation of the HIF transcription factor, and VHL inactivation results in overexpression of many hypoxia-inducible mRNAs including vascular endothelial growth factor (VEGF). To identify novel pVHL target genes we investigated the effect of wild-type (WT) pVHL on the expression of 588 cancer-related genes in two VHL-defective RCC cell lines. Expression array analysis identified nine genes that demonstrated a >2-fold decrease in expression in both RCC cell lines after restoration of WT pVHL. Three of the nine genes (VEGF, PAI-1, and LRP1) had been reported previously as pVHL targets and are known to be hypoxia-inducible. In addition, six novel targets were detected: cyclin D1 (CCND1), cell division protein kinase 6, collagen VIII alpha 1 subunit, CD59 glycoprotein precursor, integrin beta8, and interleukin 6 precursor IFN-beta2. We found no evidence that CCND1, cell division protein kinase 6, CD59, and integrin beta8 expression was influenced by hypoxia suggesting that pVHL down-regulates these targets by a HIF-independent mechanism. A type 2C pVHL mutant (V188L), which is associated with a PHE only phenotype (and had been shown previously to retain the ability to promote HIF ubiquitylation), retained the ability to suppress CCND1expression suggesting that loss of pVHL-mediated suppression of cyclin D1 is not necessary for PHE development in VHL disease. Other studies have suggested that: (a) genetic modifiers influence the phenotypic expression of VHL disease; and (b) polymorphic variation at a CCND1 codon 242 A/G single nucleotide polymorphism (SNP) may influence cancer susceptibility or prognosis in some situations. Therefore, we analyzed the relationship between CCND1 genotype and phenotypic expression of VHL disease. There was an association between the G allele and multiple retinal angiomas (P = 0.04), and risk of central nervous system hemangioblastomas (P = 0.05). These findings suggest that a variety of HIF-independent mechanisms may contribute to pVHL tumor suppressor activity and that polymorphic variation at one pVHL target influences the phenotypic expre

Topics: Blotting, Northern; Carcinoma, Renal Cell; Cyclin D1; Gene Expression Profiling; Gene Expression Regulation, Neoplastic; Genes, Tumor Suppressor; Humans; Kidney Neoplasms; Ligases; Oligonucleotide Array Sequence Analysis; Oxygen; Transfection; Tumor Cells, Cultured; Tumor Suppressor Proteins; Ubiquitin-Protein Ligases; von Hippel-Lindau Disease; Von Hippel-Lindau Tumor Suppressor Protein

Aberrations in the G1/S checkpoint are common in malignancies and are probably important for tumor development. Few G1/S studies have been performed on renal cell carcinoma (RCC) and therefore in this study the cyclin D3 protein content in 80 RCCs and that in 12 corresponding normal kidney cortex tissues are characterized using Western blotting. High cyclin D3 protein content was observed in 16% of the tumors and was significantly associated with aneuploidy, high TNM stage, high nuclear grade, high proliferation and young age. There was no association between tumor cyclin D3 and patient survival. The cyclin D3 overexpression was confirmed by immunohistochemical staining of 72 tumors, showing both nuclear and cytoplasmic localization of cyclin D3 in a fraction of the tumors. The cyclin D1 content has earlier been characterized in this tumor material and there was no relation between cyclin D1 and cyclin D3 protein expression. In summary, a fraction of the tumors overexpressed cyclin D3, supporting that various aberrations in the G1/S transition are implicated in RCCs.

Topics: Adrenal Cortex; Aged; Carcinoma, Papillary; Carcinoma, Renal Cell; Cell Nucleus; Cyclin D1; Cyclin D3; Cyclins; Cytoplasm; Female; Humans; Immunoenzyme Techniques; Kidney Neoplasms; Male; Middle Aged; Neoplasm Invasiveness; Neoplasm Staging; Ploidies; Survival Rate

Cyclin-D1 over-expression represents one of several common alterations in the G1-S transition associated with malignancies. Conclusive evidences indicate that cyclin D1 is a proto-oncogene and the gene is amplified or rearranged in different tumour types. Since very little is known about aberrations in the G1-S transition in human renal-cell carcinoma (RCC), we have characterized the expression of cyclin D1 in 80 human renal-cell carcinomas and 12 normal kidney cortex tissues using Western blotting. The cyclin-D1-protein content varied considerably and 75% of the tumours expressed higher levels than normal kidney cortex, in contrast to 25% of the tumours either lacking cyclin D1 or with low protein levels. Although it is difficult to define aberrant expression of cyclin D1, the results might indicate that the proto-oncogene was activated in a sub-set of RCC. It is also possible that low expression of cyclin D1 represents an aberrant down-regulation of the protein. Immunohistochemical assessment of cyclin D1 in a sub-set of the tumours showed large variations in the fraction of cyclin-D1-positive cells, supporting the Western-blot analyses. Surprisingly, cyclin-D1 expression did not correlate with proliferation determined by Ki-67-antigen expression or S-phase analyses. In non-papillary renal-cell carcinomas, high cyclin-D1 expression was associated with a diploid DNA profile and smaller tumour size, but there was no association between cyclin-D1 expression and tumour stage or nuclear grade. In nonpapillary tumours, high cyclin-D1 expression was further significantly associated with a better prognosis according to univariate and multivariate analyses (p = 0.005 and 0.002 respectively), as compared with highly aggressive tumours with low cyclin-D1 levels.

Topics: Adult; Aged; Carcinoma, Renal Cell; Cell Division; Cyclin D1; Female; Humans; Kidney Neoplasms; Male; Middle Aged; Proto-Oncogene Mas; Survival Rate

Cyclin D1 plays an important role in cell cycle progression from G1 to S phase. Cyclin D1 overexpression has been identified in many human neoplasms, including a variety of carcinomas. A systematic study of cyclin D1 expression in renal carcinomas and oncocytomas has not been reported. Ninety-six renal epithelial neoplasms, 78 renal carcinomas (45 clear-cell, 18 papillary, and 15 chromophobe), and 18 oncocytomas were analyzed immunohistochemically using routinely fixed tissue sections and a cocktail of two monoclonal anti-cyclin D1 antibodies. One thousand cells were manually counted, and the percentage of cyclin D1 positive cells was calculated. Fluorescence in situ hybridization studies using chromosome 11 centromeric and 11q13 specific probes were performed on a subset of clear-cell carcinomas and oncocytomas. Cyclin D1 immunoreactivity was observed in 23 (51%) of 45 clear-cell, 5 (28%) of 18 papillary, and 2 (13%) of 15 chromophobe carcinomas. Nine (50%) of 18 oncocytomas were positive for cyclin D1. Cyclin D1 expression in clear-cell carcinomas did not correlate with survival. Fluorescence in situ hybridization studies on eight clear-cell carcinomas and seven oncocytomas revealed normal chromosome 11 number and no evidence of amplification of the 11q13 locus. Thus, cyclin D1 can be immunohistochemically demonstrated in approximately one-half of renal oncocytomas and clear-cell carcinomas and is less frequent in papillary and chromophobe carcinomas. The mechanism of cyclin D1 expression is unknown, but it does not seem to be related to extra copies of chromosome 11 or to gene amplification.

Topics: Adenocarcinoma; Adenocarcinoma, Clear Cell; Adenoma, Oxyphilic; Carcinoma, Papillary; Carcinoma, Renal Cell; Cell Nucleus; Cyclin D1; Disease-Free Survival; Humans; Immunohistochemistry; In Situ Hybridization, Fluorescence; Kidney Neoplasms; Survival Analysis