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The expression of MMP-2 and MMP-9 protein was controlled by CREB in renal cell carcinoma cell lines. (A) Western blot and (B) quantitative polymerase chain reaction analysis demonstrated that MMP-2 and MMP-9 expression in siNC groups was significantly higher compared with expression in siCREB groups in 786-O and OS-RC-2 cells. In ACHN cells, knockdown of CREB increased MMP-2 and MMP-9 expression. * P<0.05 and ** P<0.01 vs. siNC group. Results are presented as the mean ± standard deviation from 3 independent experiments. MMP, matrix metallopeptidase; CREB, cyclic AMP responsive element-binding protein; siRNA, small interfering RNA; siNC, siRNA negative control; siCREB, siRNA targeting CREB.
Source publication
Renal cell carcinoma (RCC) is the most frequently occurring malignancy of the kidney worldwide. Anti-angiogenic targeted therapies inhibit the progression of RCC, however, limited effects on the invasion or metastasis of tumor cells have been observed. Cyclic AMP responsive element‑binding protein (CREB) is a serine/threonine kinase that has been i...
Contexts in source publication
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... mean ± standard deviation from 3 independent experiments. siRNA, small interfering RNA; CREB, cyclic AMP responsive element-binding protein; tCREB, total CREB; pCREB, phosphorylated CREB; NC, negative control; NS, not significant. and OS-RC-2 cells, following siCREB transfection, expressed significantly lower levels of MMP-2/9 protein (P<0.01; Fig. 4A) and mRNA (P<0.01; Fig. 4B) compared with the siNC group, however, the opposite was observed in ACHN cells where protein and mRNA levels of MMP-2/9 were significantly increased compared with the siNC group (P<0.01; Fig. 4). Combined, the results demonstrate that CREB regulated the migration and invasion of the 3 types of RCC cells, ...
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... from 3 independent experiments. siRNA, small interfering RNA; CREB, cyclic AMP responsive element-binding protein; tCREB, total CREB; pCREB, phosphorylated CREB; NC, negative control; NS, not significant. and OS-RC-2 cells, following siCREB transfection, expressed significantly lower levels of MMP-2/9 protein (P<0.01; Fig. 4A) and mRNA (P<0.01; Fig. 4B) compared with the siNC group, however, the opposite was observed in ACHN cells where protein and mRNA levels of MMP-2/9 were significantly increased compared with the siNC group (P<0.01; Fig. 4). Combined, the results demonstrate that CREB regulated the migration and invasion of the 3 types of RCC cells, however, the effects of CREB ...
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... and OS-RC-2 cells, following siCREB transfection, expressed significantly lower levels of MMP-2/9 protein (P<0.01; Fig. 4A) and mRNA (P<0.01; Fig. 4B) compared with the siNC group, however, the opposite was observed in ACHN cells where protein and mRNA levels of MMP-2/9 were significantly increased compared with the siNC group (P<0.01; Fig. 4). Combined, the results demonstrate that CREB regulated the migration and invasion of the 3 types of RCC cells, however, the effects of CREB knockdown were different between the 3 RCC cell ...
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... cells, cholangiocarcinoma cells and macrophages, CREB induced expression of MMPs (38-40). The results of the present study further demonstrated that, in 786-O and OS-RC-2 cells, CREB knockdown induced reduced MMP-2/9 expression, as determined by western blot and qPCR analysis. However, CREB silencing increased MMP-2/9 expression in ACHN cells (Fig. 4). The results of western blot and qPCR analysis corresponded with the results of wound healing and invasion assays. The results of the present study indicated that CREB-induced MMP-2/9 expression may be involved in tumor metastasis ...
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... mean ± standard deviation from 3 independent experiments. siRNA, small interfering RNA; CREB, cyclic AMP responsive element-binding protein; tCREB, total CREB; pCREB, phosphorylated CREB; NC, negative control; NS, not significant. and OS-RC-2 cells, following siCREB transfection, expressed significantly lower levels of MMP-2/9 protein (P<0.01; Fig. 4A) and mRNA (P<0.01; Fig. 4B) compared with the siNC group, however, the opposite was observed in ACHN cells where protein and mRNA levels of MMP-2/9 were significantly increased compared with the siNC group (P<0.01; Fig. 4). Combined, the results demonstrate that CREB regulated the migration and invasion of the 3 types of RCC cells, ...
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... from 3 independent experiments. siRNA, small interfering RNA; CREB, cyclic AMP responsive element-binding protein; tCREB, total CREB; pCREB, phosphorylated CREB; NC, negative control; NS, not significant. and OS-RC-2 cells, following siCREB transfection, expressed significantly lower levels of MMP-2/9 protein (P<0.01; Fig. 4A) and mRNA (P<0.01; Fig. 4B) compared with the siNC group, however, the opposite was observed in ACHN cells where protein and mRNA levels of MMP-2/9 were significantly increased compared with the siNC group (P<0.01; Fig. 4). Combined, the results demonstrate that CREB regulated the migration and invasion of the 3 types of RCC cells, however, the effects of CREB ...
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... and OS-RC-2 cells, following siCREB transfection, expressed significantly lower levels of MMP-2/9 protein (P<0.01; Fig. 4A) and mRNA (P<0.01; Fig. 4B) compared with the siNC group, however, the opposite was observed in ACHN cells where protein and mRNA levels of MMP-2/9 were significantly increased compared with the siNC group (P<0.01; Fig. 4). Combined, the results demonstrate that CREB regulated the migration and invasion of the 3 types of RCC cells, however, the effects of CREB knockdown were different between the 3 RCC cell ...
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... cells, cholangiocarcinoma cells and macrophages, CREB induced expression of MMPs (38-40). The results of the present study further demonstrated that, in 786-O and OS-RC-2 cells, CREB knockdown induced reduced MMP-2/9 expression, as deter- mined by western blot and qPCR analysis. However, CREB silencing increased MMP-2/9 expression in ACHN cells (Fig. 4). The results of western blot and qPCR analysis corre- sponded with the results of wound healing and invasion assays. The results of the present study indicated that CREB-induced MMP-2/9 expression may be involved in tumor metastasis ...
Citations
... Moreover, the beta-adrenergic antagonist propranolol has been shown to suppress tumor growth by inhibiting tumor angiogenesis and promoting T-cell recruitment in sarcoma [32], and the nonselective blockade of beta-adrenoceptors in prostate and pancreatic cancer in vivo suppressed tumor growth [33]. Additionally, beta-adrenergic activation has been associated with remodeling of the extracellular matrix to promote invasive phenotypes in breast cancer [34], which may be, in part, due to the potential cAMP-mediated epithelial-mesenchymal transition (EMT) [35,36]; however, whether ADRA2A inhibition of this signaling pathway results in suppression of the EMT is still to be determined. Those results and ours here suggest a complex relationship among adrenergic receptor subtypes and the processes involved in tumor development in a context-dependent manner. ...
Recurrence of ovarian cancer (OvCa) following surgery and standard carboplatin/paclitaxel first-line therapy signifies poor median progression-free survival (<24 months) in the majority of patients with OvCa. The current study utilized unbiased high-throughput screening (HTS) to evaluate an FDA-approved compound library for drugs that could be repurposed to improve OvCa sensitivity to carboplatin. The initial screen revealed six compounds with agonistic activity for the adrenoceptor alpha-2a (ADRA2A). These findings were validated in multiple OvCa cell lines (TYKnu, CAOV3, OVCAR8) using three ADRA2A agonists (xylazine, dexmedetomidine, and clonidine) and two independent viability assays. In all the experiments, these compounds enhanced the cytotoxicity of carboplatin treatment. Genetic overexpression of ADRA2A was also sufficient to reduce cell viability and increase carboplatin sensitivity. Taken together, these data indicate that ADRA2A activation may promote chemosensitivity in OvCa, which could be targeted by widely used medications currently indicated for other disease states.
... Furthermore, increased expression of CREBBP, EP300, and MMP9 has been reported in OSCC, and upregulation in esophageal squamous cell carcinomas has been associated with lymph node metastasis [43,44]. CREBP upregulation increased cell migration, invasion, and metastasis via upregulation of MMP2 and MMP9 [45]. With signals originating from pathways such as MAPK and PKA, phosphorylated CREBP binds to histone acetyl transferase, CREB-binding protein (CBP) and p300, to initiate CREBdependent transcription of genes that aid in cell survival and tumorigenesis [46]. ...
Introduction: Oral carcinogenesis is a multistage process, featuring genetic and molecular alterations leading to rapid cell division, invasion, metastasis, and increased cell survival. Many of these alterations are due to perturbations in the cell signaling networks, which in turn lead to constitutive deregulation of the proteins involved in the regulatory pathways. Our recent reports show that the silencing of dentin sialophosphoprotein (DSPP) and its cognate matrix metalloproteinases 20 (MMP20) alters key tumorigenic hallmarks of oral squamous cell carcinoma (OSCC). Objective: This study, intended to advance our recent findings, focuses on determining the effects of silencing DSPP and its cognate MMP20 on the signaling pathways that control cell proliferation, differentiation, invasion and metastasis. Materials and Methods: DSPP and MMP20 were silenced individually and in combination, using adenovirus-mediated short hairpin RNA (shRNA) in OSCC cell line, OSC2, and the effects of silencing on the following pathways: EFGR; RAS-RAF; MEK; MAPK; ERK; JNK; NF-kB; TGFβ; and GSK3β, were analysed by western blot. Results: DSPP and MMP20 silencing decreased EGFR, KRAS, MEK1/2, MAPK, ERK, MEEK1, JNK, CREBP, p300, NF-kB,TGF β, SMAD7, GSK3 β, and β-catenin expressions. In contrast, the expression of IKKα and SMAD4 were increased in DSPP/MMP20-silenced group, compared with control group. Furthermore, DSPP-silencing alone was more effective than MMP20, or combined DSPP-MM20 silencing, in altering the levels of key proteins of each signaling pathway investigated. Conclusion: Our findings provide the basis for further studies aimed at verifying the effects of these alterations in the profiles of these proteins on the various hallmarks of oral carcinogenesis, and for understanding the molecular role of DSPP and MMP20 in OSCC. This is with a view to evaluating their diagnostic and prognostic utility as well as the values of DSPP/MMP20 as potential targets for design of chemotherapeutic agents for the treatment of OSCC patients.
... This was demonstrated by increased pCREB signal (by IF) in PNCK overexpressing ( Figure 1) and a marked decrease in pCREB levels when PNCK is knocked down (Figure 4). CREB is a transcription factor known to play a critical role in oncogenesis and progression in multiple cancers, including RCC. 23,35,36 Wang et al. reported that targeting CREB is associated with the inhibition of RCC growth and metastatic abilities. 36 CREB regulates the expression of several cell cycle checkpoints, such as cyclin A and D1, and is crucial in cell cycle progression in cancer. ...
Renal Cell Carcinoma (RCC) is a fatal disease when advanced. While immunotherapy and tyrosine kinase inhibitor-based combinations are associated with improved survival, the majority of patients eventually succumb to disease. Through a comprehensive pan-cancer, pan-kinome analysis of the Cancer Genome Atlas (TCGA), pregnancy upregulated non-ubiquitous calcium-calmodulin dependent kinase (PNCK), was identified as the most differentially overexpressed kinase in RCC. PNCK overexpression correlated with tumor stage, grade and poor survival. PNCK overexpression in RCC cells was associated with increased CREB phosphorylation, increased cell proliferation and cell cycle progression. PNCK down-regulation, conversely, was associated with the opposite, in addition to increased apoptosis. Pathway analyses in PNCK knockdown cells showed significant down-regulation of hypoxia and angiogenesis pathways, as well as modulation of cell cycle, DNA damage and apoptosis pathways. These results demonstrate for the first time the biological role of PNCK, an understudied kinase, in RCC and validate PNCK as a druggable target.
... In addition to the above molecular markers, MMP-2 and MMP-9 are believed to be involved in the EMT process of RCC. 39 High expression of MMPs is an important feature of tumor cells escaping from the primary tumor and entering the secondary site. 40 Therefore, MMP-2 and MMP-9 can be used as key effectors for ECM remodeling and potential targets in anti-tumor therapies. ...
Background: Renal cell carcinoma (RCC) is a common cancer of the urinary system. Chinese medicine is being extensively used in the treatment of various cancers. Recent studies found that Poria cocos, a widely used Chinese medicine, has an anti-tumor effect. Pachymic acid (PA) is a triterpenoid compound of P. cocos, and the present study investigated whether PA is a main anti-tumor active ingredient of this fungus. Methods: The effects of PA on cell viability, proliferation, adhesion, invasion, and metastasis were assessed by 3-(4,5-dimethylthiazole-2-yl)-2,5-diphenyltetrazole bromide, colony formation, cell adhesion, wound healing, and Transwell assays. High-content imaging was used to dynamically observe and verify the inhibitory effect of PA on cell motility, and Western blot was used to detect the expression of matrix metalloproteinase-related proteins and epithelial–mesenchymal transformation proteins. Result: The in vitro assays demonstrated that PA was able to reduce the viability, proliferation, adhesion number, wound healing rate, and transmembrane number of the RCC cells. In addition, high-intensive imaging indicated that PA shortened the traveling distance and slowed the movement of the RCC cells. Western blots showed that in the PA-treated RCC cells, E-cadherin expression was increased, whereas N-cadherin and Vimentin expression was decreased. Furthermore, the expression levels of matrix metalloproteinase-2, matrix metalloproteinase-9, and tissue inhibitor of metalloproteinase-1 were all decreased by the PA administration. Conclusion: PA inhibited the invasion and metastasis of RCC, and these results suggest that PA may be an effective anti-tumor component of P. cocus.
... Consistently, the most frequent consequences of suppressing CREB1 by either knockdown or inhibitors are cell death, metastatic defect, failure to form colony and filopodia, and inhibition of tumor growth [15,[20][21][22][23]. Contrary to the widely acknowledged essential role of CREB1 in cancers, a comprehensive and in-depth investigation of the gene network mediated by CREB1 is less explored. Only a limited number of CREB1-targeted genes, including CCNA1, CCND1, BCL2, MMP2, MMP9, GSK3A [12,20,21,[24][25][26], were shown to contribute to tumor development. Many of them are tumor typespecific. ...
The transcription factor cyclic-AMP response element-binding protein 1 (CREB1) responds to cAMP level and controls the expression of target genes, which regulates nutrition partitioning. The promoters of CREB1-targeted genes responsive to cAMP have been extensively investigated and characterized with the presence of both cAMP response element and TATA box. Compelling evidence demonstrates that CREB1 also plays an essential role in promoting tumor development. However, only very few genes required for cell survival, proliferation and migration are known to be constitutively regulated by CREB1 in tumors. Their promoters mostly do not harbor any cAMP response element. Thus, it is very likely that CREB1 regulates the expressions of distinct sets of target genes in normal tissues and tumors. The whole gene network constitutively regulated by CREB1 in tumors has remained unrevealed. Here, we employ a systematical and integrative approach to decipher this gene network in the context of both tissue cultured cancer cells and patient samples. We combine transcriptomic, Rank-Rank Hypergeometric Overlap, and Chipseq analysis, to define and characterize CREB1-regulated genes in a multidimensional fashion. A strong cancer relevance of those top-ranked targets, which meet the most stringent criteria, is eventually verified by overall survival analysis of cancer patients. These findings strongly suggest the importance of genes constitutively regulated by CREB1 for their implicative involvement in promoting tumorigenesis.
... MMP-2 and MMP-9 were more abundantly detected in cystic fluids from cystic renal carcinomas than from benign kidney cysts [275]. The cAMP responsive element binding protein induces metastatic renal cell carcinoma via potentiating the expression of MMP-2 and MMP-9, and the proteins involved with EMT [276]. MMP-7 has been implicated in both renal cell carcinoma development and progression [277,278]. ...
Matrix metalloproteinases (MMPs) are a group of zinc and calcium endopeptidases which cleave extracellular matrix (ECM) proteins. They are also involved in the degradation of cell surface components and regulate multiple cellular processes, cell to cell interactions, cell proliferation, and cell signaling pathways. MMPs function in close interaction with the endogenous tissue inhibitors of matrix metalloproteinases (TIMPs), both of which regulate cell turnover, modulate various growth factors, and participate in the progression of tissue fibrosis and apoptosis. The multiple roles of MMPs and TIMPs are continuously elucidated in kidney development and repair, as well as in a number of kidney diseases. This chapter focuses on the current findings of the significance of MMPs and TIMPs in a wide range of kidney diseases, whether they result from kidney tissue changes, hemodynamic alterations, tubular epithelial cell apoptosis, inflammation, or fibrosis. In addition, the potential use of these endopeptidases as biomarkers of renal dysfunction and as targets for therapeutic interventions to attenuate kidney disease are also explored in this review.
... Of these, the p38 MAPK pathway is reported to play an important role in the pro-inflammatory activation of endothelial cells [32,33]. The cAMP response element binding protein (CREB) plays an important role downstream of p38MAPK in modulating expression of oxidative stress and inflammatory proteins [34][35][36]. ...
... To further determine the mechanisms modulated by thrombin in brain endothelial cells we explored the p38MAPK and CREB signaling pathways. These pathways were selected based on evidence that the p38 MAPK pathway plays an important role in the pro-inflammatory activation of endothelial cells [47] and that CREB may be involved in modulating expression of oxidative stress and inflammatory proteins downstream of p38MAPK [34][35][36]. ...
Background: Diabetes is one of the strongest disease-related risk factors for Alzheimer's disease (AD). In diabetics, hyperglycemia-induced microvascular complications are the major cause of end-organ injury, contributing to morbidity and mortality. Microvascular pathology is also an important and early feature of AD. The cerebral microvasculature may be a point of convergence of both diseases. Several lines of evidence also implicate thrombin in AD as well as in diabetes. Objective: Our objective was to investigate the role of thrombin in glucose-induced brain microvascular endothelial injury. Methods: Cultured Human brain microvascular endothelial cells (HBMVECs) were treated with 30 mM glucose ± 100 nM thrombin and ± 250 nM Dabigatran or inhibitors of PAR1, p38MAPK, MMP2, or MMP9. Cytotoxicity and thrombin activity assays on supernatants and western blotting for protein expression in lysates were performed. Results: Treatment of HBMVECs with 30 mM glucose increased thrombin activity and expression of inflammatory proteins TNF␣, IL-6, and MMPs 2 and 9; this elevation was reduced by the thrombin inhibitor dabigatran. Direct treatment of brain endothelial cells with thrombin upregulated p38MAPK and CREB, and induced TNF␣, IL6, MMP2, and MMP9 as well as oxidative stress proteins NOX4 and iNOS. Inhibition of thrombin, thrombin receptor PAR1 or p38MAPK decrease expression of inflammatory and oxidative stress proteins, implying that thrombin may play a central role in glucose-induced endothelial injury. Conclusion: Since preventing brain endothelial injury would preserve blood-brain barrier integrity, prevent neuroinflamma-tion, and retain intact functioning of the neurovascular unit, inhibiting thrombin, or its downstream signaling effectors, could be a therapeutic strategy for mitigating diabetes-induced dementia.
... CREB is a 43 kDa transcription factor, which binds after phosphorylation to the cAMP responsive element (CRE), a sequence that is localized in several gene promoters. Indeed, CREB functions are implicated amongst others in the regulation of cell proliferation, apoptosis, cycle progression and metastasis [24]. Indeed, CREB knock down in RCC cell lines suppressed RCC proliferation and decreased their tumor formation in nude mice [25]. ...
Background
The non-classical human leukocyte antigen (HLA)-G is a strong immunomodulatory molecule. Under physiological conditions, HLA-G induces immunological tolerance in immune privileged tissues, while under pathophysiological situations it contributes to immune escape mechanisms. Therefore, HLA-G could act as a potential immune checkpoint for future anti-cancer immunotherapies. Recent data suggest an aberrant expression of the cAMP response element binding protein (CREB) in clear cell renal cell carcinoma (ccRCC), which is correlated with tumor grade and stage. Furthermore, preliminary reports demonstrated a connection of CREB as a control variable of HLA-G transcription due to CREB binding sites in the HLA-G promoter region. This study investigates the interaction between CREB and HLA-G in different renal cell carcinoma (RCC) subtypes and its correlation to clinical parameters.
Methods
The direct interaction of CREB with the HLA-G promoter was investigated by chromatin immunoprecipitation in RCC cell systems. Furthermore, the expression of CREB and HLA-G was determined by immunohistochemistry using a tissue microarray (TMA) consisting of 453 RCC samples of distinct subtypes. Staining results were assessed for correlations to clinical parameters as well as to the composition of the immune cell infiltrate.
Results
There exists a distinct expression pattern of HLA-G and CREB in the three main RCC subtypes. HLA-G and CREB expression were the lowest in chromophobe RCC lesions. However, the clinical relevance of CREB and HLA-G expression differed. Unlike HLA-G, high levels of CREB expression were positively associated to the overall survival of RCC patients. A slightly, but significantly elevated number of tumor infiltrating regulatory T cells was observed in tumors of high CREB expression. Whether this small increase is of clinical relevance has to be further investigated.
Conclusions
An interaction of CREB with the HLA-G promoter could be validated in RCC cell lines. Thus, for the first time the expression of CREB and its interaction with the HLA-G in human RCCs has been shown, which might be of clinical relevance.
... MMP-2/9 and proteins associated with EMT in Renal cell carcinoma(Wang et al., 2017). ...
The CREB1 gene encodes a pleiotropic transcription factor that frequently dysregulated in cancers. CREB1 can regulates tumour cell status of proliferation or migration, however, the molecular basis for this switch involvement in cell plasticity has not been fully understood. Here, we show that knocking out CREB1 triggered a remarkable effect of epithelial-mesenchymal transition (EMT) and led to the occurrence of inhibited proliferation and enhanced motility in cancer cells. Mechanistically, CREB1-knockout cells showed arrest in the G0/G1 phase as a result of impaired CREB1-dependent transcription of CCAT1 and E2F1. Interestingly, the competition between the coactivator CBP/p300 for CREB1 and p65 leads to the activation of the NF-κB pathway in cells with CREB1 disrupted, which induces an EMT phenotype and enhances motility. These studies identified previously unknown mechanisms of CREB1 in cell plasticity via its lncRNA and protein effector pathways, revealing an important feature that should be considered in CREB1-targeted tumour therapies.
... The overexpression of SKA2 by up-regulated CREB1 promotes RCC cell proliferation in vitro and in vivo 19 . In 2017, Wang and co-worker found CREB1 to be a regulator of epithelial to mesenchymal transition (EMT) in RCC probably by controlling the expression of matrix metallopeptidase (MMP) 2/9 and further proteins involved in EMT 20 . In a follow-up study, MMP 2/9 were found to be decreased in sorafenib-treated RCC cells characterized by inhibited cell migration and invasion while the artificial overexpression of CREB1 reversed sorafenib-induced effects in these cells 21 . ...
... In these types of cancer deregulation of CREB1 was shown to be associated with enhanced proliferation, reduced apoptosis and increased angiogenesis 17 . First reports for RCC demonstrated very similar observations 19,20 . In addition, our analysis showed a significant number of RCC samples with reduced CREB1 protein levels. ...
The transcription factor cAMP response element-binding protein (CREB1) has been shown to be involved in diverse biological pathways including the regulation of cell proliferation, apoptosis, cell cycle progression, and metastasis. In this context, aberrant expression of CREB1 and the functional consequences are well investigated in a number of hematopoietic and solid tumors. However, CREB1 expression and underlying control mechanisms are only poorly analyzed in renal cell carcinoma (RCC). The present study confirmed a deregulation of CREB1 protein in the clear cell type of RCC (ccRCC) and analysis of in-house ccRCC cell lines suggested a post-transcriptional control. The combination of miRNA enrichment assay, in silico analysis and molecular biological approaches revealed four novel CREB1-regulating miRNAs, namely miR-22-3p, miR-26a-5p, miR-27a-3p, and miR-221-3p. Categorizing RCC samples as CREB1 negative or positive, respectively, the expression of these miRNAs was found to be inversely correlated with CREB1 protein levels. Analyzing 453 consecutive RCC tumors by immunohistochemistry, weakly negative, but significant correlations of CREB1 with tumor stage and grade, vascular invasion (V1) and lymphovascular invasion (L1) were found. In this respect, ccRCC might differ from other solid tumors like esophageal squamous-cell carcinoma or glioma.
