We investigated expression profiles of microRNA (miRNA) in renal cell carcinoma [clear cell carcinomas (CCC) and chromophobe renal cell carcinomas (ChCC)] and in normal kidneys by using a miRNA microarray platform which covers a total of 470 human miRNAs (Sanger miRBase release 9.1). Unsupervised hierarchical cluster analysis revealed that CCC and ChCC were separable and that no subgroups were identified in CCCs. We found that 43 miRNAs were differentially expressed between CCC and normal kidney, of which 37 were significantly down-regulated in CCC and the other 6 were up-regulated. We also found that 57 miRNAs were differentially expressed between ChCC and normal kidney, of which 51 were significantly down-regulated in ChCC and the other 6 were up-regulated. Together, these observations indicate that expression of miRNAs tends to be down-regulated in both CCC and ChCC compared with normal kidney. We observed that miR-141 and miR-200c were the most significantly down-regulated miRNAs in CCCs. Indeed, in all cases of CCC analysed, both miR-141 and miR-200c were down-regulated in comparison with normal kidney. Microarray data and quantitative RT-PCR showed that these two miRNAs were expressed concordantly. TargetScan algorithm revealed that ZFHX1B mRNA is a hypothetical target of both miR-141 and -200c. We established by quantitative RT-PCR that, in CCCs in which miR-141 and miR-200c were down-regulated, ZFHX1B, a transcriptional repressor for CDH1/E-cadherin, tended to be up-regulated. Furthermore, we found that overexpression of miR-141 and miR-200c caused down-regulation of ZFHX1B and up-regulation of E-cadherin in two renal carcinoma cell lines, ACHN and 786-O. On the basis of these findings, we suggest that down-regulation of miR-141 and miR-200c in CCCs might be involved in suppression of CDH1/E-cadherin transcription via up-regulation of ZFHX1B.

BackgroundClinical outcome of patients with high-grade ccRCC (clear cell renal cell carcinoma) remains still poor despite recent advances in treatment strategies. Molecular mechanism of pathogenesis in developing high-grade ccRCC must be clarified. In the present study, we found that SAV1 was significantly downregulated with copy number loss in high-grade ccRCCs. Therefore, we investigated the SAV1 function on cell proliferation and apoptosis in vitro. Furthermore, we attempted to clarify the downstream signaling which is regulated by SAV1.MethodsWe performed array CGH and gene expression analysis of 8 RCC cell lines (786-O, 769-P, KMRC-1, KMRC-2, KMRC-3, KMRC-20, TUHR4TKB, and Caki-2), and expression level of mRNA was confirmed by quantitative RT-PCR (qRT-PCR) analysis. We next re-expressed SAV1 in 786-O cells, and analyzed its colony-forming activity. Then, we transfected siRNAs of SAV1 into the kidney epithelial cell line HK2 and renal proximal tubule epithelial cells (RPTECs), and analyzed their proliferation and apoptosis. Furthermore, the activity of YAP1, which is a downstream molecule of SAV1, was evaluated by western blot analysis, reporter assay and immunohistochemical analysis.ResultsWe found that SAV1, a component of the Hippo pathway, is frequently downregulated in high-grade ccRCC. SAV1 is located on chromosome 14q22.1, where copy number loss had been observed in 7 of 12 high-grade ccRCCs in our previous study, suggesting that gene copy number loss is responsible for the downregulation of SAV1. Colony-forming activity by 786-O cells, which show homozygous loss of SAV1, was significantly reduced when SAV1 was re-introduced exogenously. Knockdown of SAV1 promoted proliferation of HK2 and RPTEC. Although the phosphorylation level of YAP1 was low in 786-O cells, it was elevated in SAV1-transduced 786-O cells. Furthermore, the transcriptional activity of the YAP1 and TEAD3 complex was inhibited in SAV1-transduced 786-O cells. Immunohistochemistry frequently demonstrated nuclear localization of YAP1 in ccRCC cases with SAV1 downregulation, and it was preferentially detected in high-grade ccRCC.ConclusionsTaken together, downregulation of SAV1 and the consequent YAP1 activation are involved in the pathogenesis of high-grade ccRCC. It is an attractive hypothesis that Hippo signaling could be candidates for new therapeutic target.

The purpose of the present study was to determine the genomic profile of renal cell carcinoma (RCC) in end‐stage renal disease (ESRD) by analyzing genomic copy number aberrations. Seventy‐nine tumor samples from 63 patients with RCC‐ESRD were analyzed by array comparative genomic hybridization using the Agilent Whole Human Genome 4 × 44K Oligo Micro Array (Agilent Technologies Inc., Palo Alto, CA, USA). Unsupervised hierarchical clustering analysis revealed that the 63 cases could be divided into two groups, Clusters A and B. Cluster A was comprised mainly of clear cell RCC (CCRCC), whereas Cluster B was comprised mainly of papillary RCC (PRCC), acquired cystic disease (ACD)‐associated RCC, and clear cell papillary RCC. Analysis of the averaged frequencies revealed that the genomic profiles of Clusters A and B resembled those of sporadic CCRCC and sporadic PRCC, respectively. Although it has been proposed on the basis of histopathology that ACD‐associated RCC, clear cell papillary RCC and PRCC‐ESRD are distinct subtypes, the present data reveal that the genomic profiles of these types, categorized as Cluster B, resemble one another. Furthermore, the genomic profiles of PRCC, ACD‐associated RCC and clear cell papillary RCC admixed in one tissue tended to resemble one another. On the basis of genomic profiling of RCC‐ESRD, we conclude that the molecular pathogenesis of CCRCC‐ESRD resembles that of sporadic CCRCC. Although various histologic subtypes of non‐clear cell RCC‐ESRD have been proposed, their genomic profiles resemble those of sporadic PRCC, suggesting that the molecular pathogenesis of non‐CCRCC‐ESRD may be related to that of sporadic PRCC. (Cancer Sci 2012; 103: 569–576)

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Purpose: Glucose is a major energy resource for tumor cell survival and growth, and its influx into cells is mainly carried out by facilitative glucose transporters (GLUTs). Sodium - dependent glucose transporters (SGLTs) have been highlighted as playing important roles in diabetic treatment. However, their potential roles in cancer remain unclear. We examined expression patterns of SGLTs in tumor tissues together with conventional pathological variables to determine prognostic significance in patients with renal cell carcinoma (RCC). Materials and Methods: Nephrectomy specimens were obtained from 68 patients. GLUT - 1, - 2 and SGLT - 1, - 2 expression in tumor and adjacent normal tissues were analyzed by immunohistochemical staining, and intensity was quantified using an image analyzer. Results: The four glucose transporters evaluated were broadly distributed in tumor tissues as well as throughout the normal parenchyma. There was no significant correlation between transporter expression and conventional pathological variables. However, increased SGLT - 2 expression was significantly associated with shorter overall survival (p < 0.01), regardless of metastatic status. Conclusions: We propose possible prognostic significance of SGLT - 2 expression in human RCC. Given that glucose is a major energy resource for tumor cells and that glucose transport is largely mediated by SGLT, SGLT - 2 may serve as a possible therapeutic target in RCC.