Search

INTRODUCTION AND OBJECTIVES: There are more than 200,000 cases of renal cell carcinoma (RCC) each year in the world, accounting for approximately 2% of all cancers. RCC clear cell type (ccRCC) is the most common subtype of RCC and accounts for 75 to 80% of the cases with highest rates of local invasion, development of metastasis and mortality. New target therapy is based on antiangiogenic antibodies and molecules, changing the course of the disease, but the results so far are disappointing. Our aim is to study miRNAs and their target genes related to angiogenesis in ccRCC trying to bring some new knowledge to the molecular pathways related to the disease METHODS: The expression levels of miRNAs miR-99a, 99b, 100; 199a; 106a; 106b; 29a; 29b; 29c; 126; 200a, 200b and their respective target genes: mTOR, HIF1-a, VHL, PDGF, VEGF, VEGFR1 and VEGFR2 were evaluated using qRT-PCR.in snap-frozen tumor tissue samples from 56 patients diagnosed with ccRCC and 5 samples of benign renal tissue as control. The results were related to tumor size, Fuhrman nuclear grade and microvascular invasion, considering the risk criteria proposed by Dall’Oglio et al. (2007). RESULTS: We found an underexpression of the most genes except the VEGFA and PDGF, while the analysis of miRNAs showed underexpression only the miR 199a, 100 and 126. We compared the expression of genes with their possible regulatory miRNAs, and we found that mTOR was underexpressed while miR99a was overexpressed in most samples. This relationship also occurs between VEGFA and miR126 and between miRNAs 106a, 106b, and their target gene VHL. Considering the risk groups the overexpression of miR200b was associated with high-risk patients (p 1⁄4 0.01) and the overexpression of miR126 was associated with lower Fuhrman grade (I-II) (p 1⁄4 0.03). CONCLUSIONS: Our results show that in ccRCC there is an unbalance in the expression of genes and miRNAs related to angiogenesis and cell proliferation and survival. Furthermore with our findings we can speculate the role of miR200b and miR126 in the prognostic of ccRCC. We believe that the relationship between miRNAs and their respective genes should be more profoundly searched as markers and possible therapeutic agents in this neoplasia.

Lower pole renal stones are well known to show a poor stone clearance rate after extracorporeal shock wave lithotripsy (ESWL) (Dornier Medical Systems, Marietta, Georgia). In the current study we analyzed several anatomical factors as predictors of lower pole stone clearance that may be used to indicate the usefulness of ESWL in such patients.Between January 1993 and October 2000, 63 patients with a unilateral single lower pole stone of 2 cm. or less were included in the study. Excretory urography was done to determine the lower infundibulopelvic angle, caliceal pelvic height, lower infundibular length and diameter, lower infundibular length-to-diameter ratio and number of lower pole minor calices. Stone-free status was assessed by x-ray.The overall stone clearance rate was 54% (34 of 63 patients). While caliceal pelvic height, lower infundibular diameter, length, length-to-diameter ratio and number of minor calices were different in the stone-free and residual stone groups on univariate analysis, multivariate logistic analysis revealed that the lower infundibular length-to-diameter ratio, diameter and number of minor calices were independent predictors of successful stone clearance. The 13 patients (20.6%) who showed all 3 favorable anatomical factors (lower infundibular length-to-diameter ratio less than 7, diameter greater than 4 mm. and a single minor calix) achieved an impressive 84.6% stone clearance rate. In patients with only 1 or 2 favorable factors the stone clearance rate was still greater than 60%. In contrast, the stone clearance rate was only 6.7% in the 15 patients who showed none of these factors.From this study it is apparent that successful ESWL is highly sensitive to the anatomy of the lower pole of the kidney. If a patient with a lower pole stone has at least 1 of the favorable factors identified in this study, ESWL may be advocated as first line therapy with a greater than 60% prospect of a successful outcome. Other treatment options should be considered in those without any of these predictive factors.

growth rates, protein, and gene expression were analyzed. Effects of combination AR-ASO and OGX-011 on ENZ-R LNCaP progression rates were assessed in castrated male athymic mice. RESULTS: ENZ-R LNCaP-derived sub-lines expressed high levels of CLU and AR compared to CRPC LNCaP cell lines. LNCaP cells overexpressing CLU acquire resistance to AR-ASO induced apoptosis as shown by decreased cell growth inhibition rates and apoptosis induction. AR-ASO induced timeand dose-dependent increases in CLU levels in parental and ENZ-R LNCaP cells. Co-targeted suppression of CLU and AR synergistically inhibited ENZ-R cell growth rates compared to monotherapy. Increased apoptosis (PARP and caspase-3 cleavage, sub G0/G1 fraction) together with reduced AKT phosphorylation was most enhanced with combined therapy. In vivo, combined OGX-011 + AR-ASO significantly delayed ENZ-R LNCaP tumor progression compared to scramble ASO + AR-ASO. CONCLUSIONS: Suppression of CLU combined with AR knockdown inhibited ENZ-R LNCaP cell growth, providing pre-clinical proof-ofprinciple as a promising AR co-targeting approach in ENZ-R CRPC.

INTRODUCTION AND OBJECTIVES: Although it is well known that deregulation of VHL-HIF1 pathway is responsible for the carcinogenesis of clear cell renal cell carcinoma (ccRCC), molecular mechanisms responsible for the development of metastasis are still poorly investigated. The first aim of this study is to identify copy number alteration (CNA) that is specifically detected in metastasis but not in primary lesion. The second aim is to identify a gene whose deregulation is responsible for the development of metastasis. METHODS: For the array comparative genomic hybridization (CGH) analysis, genomic DNAs were extracted from the formalin-fixed, paraffin-embedded tissues. Twenty cases of primary ccRCCs and their corresponding metastases were analyzed using 4x44K oligonucleotide CGH array (Agilent Inc.). For the gene expression analysis, total RNA was extracted from frozen samples of 30 primary ccRCCs. Expression levels of the 58 genes were analyzed by qRT-PCR. To investigate the biologic function of candidate genes, RCC cell lines, 786-O and 769-P were used. Cell proliferation, invasiveness and apoptosis were analyzed before and after the infection of lenti-virus encoding candidate genes to the RCC cell lines. RESULTS: By comparing CNAs of primary ccRCCs with their corresponding metastases, we found that gene copy number loss at 9p24.1-p13.3 was more frequently found in metastases than in primary lesions. Out of 58 genes located at 9p24.1-p13.3, we identified 2 genes, NDUFB6 and LRRC19, both of which were down-regulated due to copy number loss. Next, using lenti-virus encoding each gene, we introduced the two genes into 786-O and 769-P cells in which both the 2 genes were down-regulated. We found that, when NDUFB6 was overexpressed in both cell lines, cell proliferation was significantly suppressed (figure shown below), although apoptotic activity and invasion activity were unchanged. CONCLUSIONS: Down-regulation of NDUFB6 caused by gene copy number loss at 9p24.1-p13.3 may bring about growth advantage to metastatic ccRCC cells. We propose that NDUFB6 is a new candidate for a tumor suppressor and that its down-regulation may play an important role in the development of metastasis of ccRCC.

Toru Inoue*, Keiko Matsuura, Chisato Nakada, Takeo Nomura, Fuminori Sato, Yufu, Japan; Yoji Nagashima, Yokohama, Japan; Chikara Ohyama, Hirosaki, Japan; Tomonori Habuchi, Akita, Japan; Yasuhiko Tomita, Yamagata, Japan; Satoru Takahashi, Tokyo, Japan; Osamu Ogawa, Kyoto, Japan; Norio Nonomura, Osaka, Japan; Akio Matsubara, Hiroshima, Japan; Masayuki Nakagawa, Kagoshima, Japan; Masatsugu Moriyama, Hiromitsu Mimata, Yufu, Japan

INTRODUCTION AND OBJECTIVES: Distant metastasis is one of the most important prognostic factors of clear cell renal cell carcinoma (ccRCC). To identify genomic copy number alterations (CNAs) responsible for metastasis, we compared genomic profiles between primary lesions of ccRCC and corresponding metastatic lesions using array comparative genomic hybridization (array CGH). METHODS: Genomic DNAs for samples were extracted from formalin fixed paraffin embedded tissues of 19 primary ccRCCs and corresponding metastases. Genomic DNAs for controls were from normal renal cortexes of the same patients. Array CGH was performed using Agilent Whole Human Genome 4x44K Oligo Micro Array. RESULTS: To identify CNAs correlated with metastasis of ccRCC, we compared CNAs of 19 primary lesions and corresponding metastatic lesions. In 9 cases, loss at 9p24.1-p13.3 was detected in both primary and metastatic lesions. While, in 8 of the remaining 10 cases without 9p24.1-p13.3 loss at primary lesions, 9p24.1-p13.3 loss was detected at metastatic lesions. In the remaining 2 of the 10 cases, 9p24.1-p13.3 loss was detected in neither primary nor metastatic lesions. Similar results were detected in terms of 8p12-p11.23 loss. These results indicate that frequencies of losses at 8p12-p11.23 and 9p24.1-p13.3 were significantly higher in metastatic lesions than in primary lesions (p 0.01), leading us to speculate that these CNAs may be added in the process of metastasis. Next, we investigated whether these CNAs are useful to predict metastasis. In primary lesions of 15 cases that were free of metastasis at first diagnosis, 9p24.1-p13.3 loss was detected in 5 cases but undetected in the other 10 cases. Between the 2 groups, we compared the recurrence free survival rate. We found that, in cases with 9p24.1-p13.3 loss at primary lesion, metastasis tends to occur earlier than those without 9p24.1-p13.3 loss at primary lesions (p 0.05). While, there was no significant difference in terms of 8p12-p11.23 loss. These results suggest that the presence of 9p24.1p13.3 loss at primary lesion may be a predictive indicator of metastasis. CONCLUSIONS: Genomic losses at 8p12-p11.23 and 9p24.1p13.3 are correlated with metastases of ccRCCs. Investigation of the presence or absence of loss at 9p24.1-p13.3 may be useful as an indicator to predict metastasis of ccRCC.