Gain Of 5q Research Articles

Gain of Chromosome 5q Predicts a Favorable Prognosis in Localized Renal Cell Carcinoma

Approximately 65% of renal cell carcinomas (RCC) are diagnosed at a localized stage. We investigated the chromosome 5q gain impact on disease-free survival (DFS) in RCC patients. Overall, 676 patients with stages 1–2 RCC and having cytogenetic analysis were included. Gain of 5q was observed in 108 patients, more frequently in clear cell (ccRCC) than non-clear cell tumors. Gain of 5q is likely an independent prognostic factor since the concerned patients had a decreased recurrence risk in stages 1–2 RCC, confirmed in multivariable analysis. Detecting 5q gain could enhance recurrence risk assessment, allowing tailored post-surgery surveillance, and reducing unnecessary treatments.

Risk prediction for metastasis of clear cell renal cell carcinoma using digital multiplex ligation-dependent probe amplification.

Precise quantification of copy‐number alterations (CNAs) in a tumor genome is difficult. We have applied a comprehensive copy‐number analysis method, digital multiplex ligation‐dependent probe amplification (digitalMLPA), for targeted gene copy‐number analysis in clear cell renal cell carcinoma (ccRCC). Copy‐number status of all chromosomal arms and 11 genes was determined in 60 ccRCC samples. Chromosome 3p loss and 5q gain, known as early changes in ccRCC development, as well as losses at 9p and 14q were detected in 56/60 (93.3%), 31/60 (51.7%), 11/60 (18.3%), and 33/60 (55%), respectively. Through gene expression analysis, a significant positive correlation was detected in terms of 14q loss determined using digitalMLPA and downregulation of mRNA expression ratios with HIF1A and L2HGDH (P = .0253 and .0117, respectively). Patients with early metastasis (<1 y) (n = 18) showed CNAs in 6 arms (in median), whereas metastasis‐free patients (n = 34) showed those in significantly less arms (3 arms in median) (P = .0289). In particular, biallelic deletion of CDKN2A/2B was associated with multiple CNAs (≥7 arms) in 3 tumors. Together with sequence‐level mutations in genes VHL, PBRM1, SETD2, and BAP1, we performed multiple correspondence analysis, which identified the association of 9p loss and 4q loss with early metastasis (both P < .05). This analysis indicated the association of 4p loss and 1p loss with poor survival (both, P < .05). These findings suggest that CNAs have essential roles in aggressiveness of ccRCC. We showed that our approach of measuring CNA through digitalMLPA will facilitate the selection of patients who may develop metastasis.

Cancer Genetics and Therapeutic Opportunities in Urologic Practice.

This article aims to summarize the current literature on genetic alterations related to tumors of the genitourinary tract. Novel associations have recently been reported between specific DNA alterations and genitourinary malignancies. The most common cause of chromosome 3p loss in clear cell renal cell carcinoma is a chromothripsis event, which concurrently generates a chromosome 5q gain. Specific patterns of clear cell renal cell carcinoma metastatic evolution have been uncovered. The first therapy targeting a specific molecular alteration has now been approved for urothelial carcinoma. Germline mutations in DNA damage repair genes and the transcription factor HOXB13 are associated with prostate cancer and may be targeted therapeutically. The genetic associations noted across different genitourinary cancers can inform potential screening approaches and guide novel targeted treatment strategies.

Prognostic value of gain of chromosome 5q in localized renal cell carcinoma.

623 Background: While gain of chromosome 5q is a frequently seen cytogenetic abnormality noted to occur in patients with renal cell carcinoma (RCC), little is known about its prognostic significance. We investigated the association of gain of 5q with disease-free survival (DFS) in patients with localized (non-metastatic T1-2) RCC. Methods: All patients from UCLA with primary tumor stage T1-2 RCC who had tumor cytogenetic analysis on tumor specimen were included. Alterations in chromosome 5q was specifically reviewed in this study. Logistic regression analyses were used to assess association of gain of 5q with final histopathology, ISUP grading, and T-stage. Cox proportional hazard modeling and Kaplan-Meier analyses were used to assess the impact of gain of 5q on DFS. Recurrence was defined as any local recurrence or development of new metastasis. Results: A total of 676 patients were included in this study. Gain of 5q occurred in 108 (16%) patients and was more commonly seen in clear cell versus non-clear cell tumors (19% vs. 9%, p = .002). Gain of 5q was associated with low grade ISUP (1 or 2) of clear-cell RCC (p = 0.011). On survival analysis, there was a 67% decreased risk of RCC recurrence for patients with gain of 5q (HR = 0.33, p = 0.018). The 5- and 10-year risk of recurrence was 2% vs. 16% and 14% vs. 28% for patients with and without gain of 5q, respectively (p = 0.013). In multivariable Cox analysis, the gain of 5q was an independent prognostic factor (p = .026 with ISUP grade and p = .025 with T-stage). These findings were confirmed among clear-cell RCC subgroup. Conclusions: Gain of chromosome 5q is an independent prognostic factor associated with decreased risk of recurrence in patients with localized T1-2 renal cell carcinoma. Identifying patients with a gain of 5q will improve the stratification of the risk of recurrence, could allow to adapt the follow-up protocols and avoid adjuvant treatment.

"High-grade oncocytic renal tumor": morphologic, immunohistochemical, and molecular genetic study of 14 cases.

The spectrum of the renal oncocytic tumors has been expanded in recent years to include several novel and emerging entities. We describe a cohort of novel, hitherto unrecognized and morphologically distinct high-grade oncocytic tumors (HOT), currently diagnosed as "unclassified" in the WHO classification. We identified 14 HOT by searching multiple institutional archives. Morphologic, immunohistochemical (IHC), molecular genetic, and molecular karyotyping studies were performed to investigate these tumors. The patients included 3 men and 11 women, with age range from 25 to 73years (median 50, mean 49years). Tumor size ranged from 1.5 to 7.0cm in the greatest dimension (median 3, mean 3.4cm). The tumors were all pT1 stage. Microscopically, they showed nested to solid growth, and focal tubulocystic architecture. The neoplastic cells were uniform with voluminous oncocytic cytoplasm. Prominent intracytoplasmic vacuoles were frequently seen, but no irregular (raisinoid) nuclei or perinuclear halos were present. All tumors demonstrated prominent nucleoli (WHO/ISUP grade 3 equivalent). Nine of 14 cases were positive for CD117 and cytokeratin (CK) 7 was either negative or only focally positive in of 6/14 cases. All tumors were positive for AE1-AE3, CK18, PAX 8, antimitochondrial antigen, and SDHB. Cathepsin K was positive in 13/14 cases and CD10 was positive in 12/13 cases. All cases were negative for TFE3, HMB45, Melan-A. No TFEB and TFE3 genes rearrangement was found in analyzable cases. By array CGH, complete chromosomal losses or gains were not found in any of the cases, and 3/9 cases showed absence of any abnormalities. Chromosomal losses were detected on chromosome 19 (4/9), 3 with losses of the short arm (p) and 1 with losses of both arms (p and q). Loss of chromosome 1 was found in 3/9 cases; gain of 5q was found in 1/9 cases. On molecular karyotyping, 3/3 evaluated cases showed loss of heterozygosity (LOH) on 16p11.2-11.1 and 2/3 cases showed LOH at 7q31.31. Copy number (CN) losses were found at 7q11.21 (3/3), Xp11.21 (3/3), Xp11.22-11.21 (3/3), and Xq24-25 (2/3). CN gains were found at 13q34 (2/3). Ten patients with available follow up information were alive and without disease progression, after a mean follow-up of 28months (1 to 112months). HOT is a tumor with unique morphology and its IHC profile appears mostly consistent. HOT should be considered as an emerging renal entity because it does not meet the diagnostic criteria for other recognized eosinophilic renal tumors, such as oncocytoma, chromophobe renal cell carcinoma (RCC), TFE3 and TFEB RCC, SDH-deficient RCC, and eosinophilic solid and cystic RCC.

Timing the Landmark Events in the Evolution of Clear Cell Renal Cell Cancer: TRACERx Renal

SummaryClear cell renal cell carcinoma (ccRCC) is characterized by near-universal loss of the short arm of chromosome 3, deleting several tumor suppressor genes. We analyzed whole genomes from 95 biopsies across 33 patients with clear cell renal cell carcinoma. We find hotspots of point mutations in the 5′ UTR of TERT, targeting a MYC-MAX-MAD1 repressor associated with telomere lengthening. The most common structural abnormality generates simultaneous 3p loss and 5q gain (36% patients), typically through chromothripsis. This event occurs in childhood or adolescence, generally as the initiating event that precedes emergence of the tumor’s most recent common ancestor by years to decades. Similar genomic changes drive inherited ccRCC. Modeling differences in age incidence between inherited and sporadic cancers suggests that the number of cells with 3p loss capable of initiating sporadic tumors is no more than a few hundred. Early development of ccRCC follows well-defined evolutionary trajectories, offering opportunity for early intervention.

Versican Promotes Tumor Progression, Metastasis and Predicts Poor Prognosis in Renal Carcinoma.

The proteoglycan versican (VCAN) promotes tumor progression and enhances metastasis in several cancers; however, its role in clear cell renal cell carcinoma (ccRCC) remains unknown. Recent evidence suggests that VCAN is an important target of chromosomal 5q gain, one of the most prevalent genetic abnormalities in ccRCC. Thus, we investigated whether VCAN expression is associated with the pathogenesis of ccRCC. VCAN expression was analyzed using three RCC and normal kidney cell lines as well as a clinical cohort of 84 matched ccRCC and normal renal tissues. Functional analyses on growth and progression properties were performed using VCAN-depleted ccRCC cells. Microarray expression profiling was employed to investigate the target genes and biologic pathways involved in VCAN-mediated ccRCC carcinogenesis. ccRCC had elevated VCAN expression in comparison with normal kidney in both cell lines and clinical specimens. The elevated expression of VCAN was significantly correlated with metastasis (P < 0.001) and worse 5-year overall survival after radical nephrectomy (P = 0.014). In vitro, VCAN knockdown significantly decreased cell proliferation and increased apoptosis in Caki-2 and 786-O cells, and this was associated with alteration of several TNF signaling-related genes such as TNFα, BID, and BAK Furthermore, VCAN depletion markedly decreased cell migration and invasion which correlated with reduction of MMP7 and CXCR4. These results demonstrate that VCAN promotes ccRCC tumorigenesis and metastasis and thus is an attractive target for novel diagnostic, prognostic, and therapeutic strategies.Implications: This study highlights the oncogenic role of VCAN in renal cell carcinogenesis and suggests that this gene has therapeutic and/or biomarker potential for renal cell cancer. Mol Cancer Res; 15(7); 884-95. ©2017 AACR.

Abstract 5749: VCAN promotes clear cell renal cell carcinoma tumor progression and metastasis, and predicts poor prognosis

Abstract Introduction and Objectives: Although versican (VCAN) is known to promote tumor progression and enhance metastasis of several types of cancers, its role in clear cell renal cell carcinoma (ccRCC) remains unknown. Interestingly, a recent report showed that VCAN is an important target of chromosomal 5q gain, one of the most prevalent genetic abnormalities in ccRCC cases. In the present study, we investigated whether VCAN expression is associated with the pathogenesis of ccRCC. Methods: VCAN expression was analyzed in 3 RCC and normal kidney cell lines, as well as 84 matched ccRCC and normal renal tissues. We also performed various functional analyses of growth and progression properties using VCAN-depleted ccRCC cells. Microarray analysis was then employed to investigate the target genes of the pathway involved in ccRCC tumorigenesis and development. Results: There are 4 isoforms of VCAN containing the N-terminal globular (G1 domain) and C-terminal globular (G3 domain) domains, each of which was found to be over-expressed in the ccRCC samples as compared to the controls. Higher VCAN expression was significantly correlated with metastasis (p&amp;lt;0.001) and worse 5-year overall survival after a radical nephrectomy (p=0.014). In vitro, VCAN knockdown by siRNA in Caki-2 and 786-O cells significantly decreased cell proliferation and increased apoptosis, and was also found to be associated with alteration of several TNF signaling-related genes, such as TNF-α, BID, and BAK. Furthermore, VCAN depletion markedly decreased cell migration and invasion associated with reduced MMP7 and CXCR4 levels. Conclusion: Our results demonstrate that VCAN promotes ccRCC tumorigenesis and metastasis, showing it to be an attractive novel target for diagnostic, prognostic, and therapeutic strategies for affected patients. Note: This abstract was not presented at the meeting. Citation Format: Yozo Mitsui, Taku Kato, Shigekatsu Maekawa, Yutaka Hashimoto, Marisa Shiina, Mitsuho Imai Sumida, Ryan Kenji Wong, Soichiro Yamamura, Varahram Shahryari, Shahana Majid, Sharanjot Saini, Guoren Deng, Rajvir Dahiya, Koichi Nakajima, Yuichiro Tanaka. VCAN promotes clear cell renal cell carcinoma tumor progression and metastasis, and predicts poor prognosis [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 5749. doi:10.1158/1538-7445.AM2017-5749

Genetic characterization of Polish ccRCC patients: somatic mutation analysis of PBRM1, BAP1 and KDMC5, genomic SNP array analysis in tumor biopsy and preliminary results of chromosome aberrations analysis in plasma cell free DNA

BackgroundMutation analysis and cytogenetic testing in clear cell renal cell carcinoma (ccRCC) is not yet implemented in a routine diagnostics of ccRCC.Material and methodsWe characterized the chromosomal alterations in 83 ccRCC tumors from Polish patients using whole genome SNP genotyping assay. Moreover, the utility of next generation sequencing of cell free DNA (cfDNA) in patients plasma as a potential tool for non-invasive cytogenetic analysis was tested. Additionally, tumor specific somatic mutations in PBRM1, BAP1 and KDM5C were determinedResultsWe confirmed a correlation between deletions at 9p and higher tumor size, and deletion of chromosome 20 and the survival time. In Fuhrman grade 1, only aberrations of 3p and 8p deletion, gain of 5q and 13q and gains of chromosome 7 and 16 were present. The number of aberrations increased with Fuhrman grade, all chromosomes displayed cytogenetic changes in G3 and G4. ccRCC specific chromosome aberrations were observed in cfDNA, although discrepancies were found between cfDNA and tumor samples. In total 12 common and 94 rare variants were detected in PBRM1, BAP1 and KDM5C, with four potentially pathogenic variants. We observed markedly lower mutation load in PBRM1.ConclusionsCytogenetic analysis of cfDNA may allow more accurate diagnosis of tumor aberrations and therefore the correlation between the chromosome aberrations in cfDNA and clinical outcome should be studied in larger cohorts. The functional studies on in BAP1, KDM5C, PBRM1 mutations in large, independent sample set would be necessary for the assessment of their prognostic and diagnostic potential.

Abstract 3559: Long non-coding RNA subtype classification of clear-cell renal cell carcinoma

Abstract Background: Long non-coding RNAs (lncRNA) play a key role in regulating cell physiology through different mechanisms including regulation of transcription and splicing, and recruitment of histone-modifying enzymes. The portrait of lncRNA in kidney cancers remains fully unknown. Material and Methods: we performed a genomic analysis of GENCODE lncRNAs in clear-cell renal cell carcinoma (ccRCC), using The Cancer Genome Atlas (TCGA) molecular RNAseq profiles of 475 primary tumors. Furthermore, we described global correlations between lncRNA and expression of cis- and trans-acting genes, and established lncRNA expression subtypes based on distinct signatures. In addition, lncRNA expression was correlated with miRNA, copy-number alterations and the chromatin landscape of human kidney. Results: Using stringent criteria (RPKM ≥1 in at least 10% of ccRCC), we identified 1,934 lncRNAs and predicted those that are potential drivers of cancer progression through integrative analysis. The expression of 1,057 (54.6%) lncRNAs showed a positive correlation with the mRNA expression of their neighboring genes, while only 13 (0.7%) lncRNAs showed a negative correlation. Overall, 844 (43.6%) lncRNAs were pre-marked by H3K4me3 in normal kidney, suggesting that those lncRNA might arise from promoter-associated elements. Conversely, only 81 (4.2%) and 182 (9.4%) lncRNAs were pre-marked by H3K36me3 and H3K27me3, respectively. Unsupervised clustering of lncRNA expression revealed four robust subtypes, which differ from the ccRCC transcriptomic subgroups. Interestingly, patients belonging to cluster II (n=111) displayed poor overall survival as compared to patients of other clusters (p=2.6x10-6). Intriguingly, there were 37 (7.8%) ccRCC belonging to cluster IV, and those were enriched for TFE3/TFEB RCC. Furthermore, tumors within cluster IV were devoid from 3p loss and 5q gain which represent the most frequent genomic aberrations in ccRCC. Finally, GREAT analysis reveals that overexpressed lncRNA in cluster IV were exclusively enriched for genes belonging to collecting duct (p=1.6x10-13) highly suggesting that their cell of origin might be different. Conclusion: This study describes the first portrait of lncRNA in ccRCC and proposes a robust lncRNA subtype classification of ccRCC, which may serve in predicting patient outcome. Furthermore, we identified a small ccRCC subgroup with distinct lncRNA and karyotypic phenotype. Citation Format: Gabriel G. Malouf, Jianping Zhang, Nizar Tannir, Erika J. Thompson, David Khayat, Jean-Philippe Spano, Xiaoping Su. Long non-coding RNA subtype classification of clear-cell renal cell carcinoma. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 3559. doi:10.1158/1538-7445.AM2014-3559

Multiplex ligation‐dependent probe amplification and fluorescence in situ hybridization are complementary techniques to detect cytogenetic abnormalities in multiple myeloma

Multiple myeloma (MM) is a genetically heterogeneous disease with diverse clinical outcomes. Interphase fluorescence in situ hybridization (i-FISH) is the most commonly used approach to detect recurrent cytogenetic abnormalities in this malignancy. We aimed to assess the performance of multiplex ligation-dependent probe amplification (MLPA) to reveal copy number abnormalities (CNAs) in MM. Diagnostic bone marrow samples from 81 patients were analyzed using 42 MLPA probes for the following regions: 1p32-31, 1p21, 1q21.3, 1q23.3, 5q31.3, 12p13.31, 13q14, 16q12, 16q23, and 17p13. All samples were also screened by i-FISH for the presence of hyperdiploidy, deletion/monosomy of chromosome 13, deletion of TP53, disruption of the immunoglobulin heavy-chain gene, t(4;14), t(11;14), t(14;16), t(8;14), gain of 5q and abnormalities of chromosome 1. A total of 245 alterations were detected in 79 cases (98%). Investigating the same aberrations, the two methods showed a congruency of higher than 90%. A low proportion of cells with the relevant abnormality, focal CNAs and unmatched probes were responsible for the discrepancies. MLPA revealed 95 CNAs not detected by i-FISH providing additional information in 53 cases (65%). Scrutiny of CNAs on chromosome 1, using more than 20 probes, revealed significant heterogeneity in size and location, and variable intra-chromosomal and intra-clonal rates of loss or gain. Our results suggest that MLPA is a reliable high-throughput technique to detect CNAs in MM. Since balanced aberrations are key to prognostic classification of this disease, MLPA and i-FISH should be applied as complementary techniques in diagnostic pathology.

Impact of chromosomal copy number variation on outcome in metastatic clear cell renal cell carcinoma patients treated with antiangiogenic agents.

393 Background: Antiangiogenic agents (AA) are used to treat metastatic clear cell renal cell carcinoma (ccRCC). Currently there are no biomarkers of therapeutic efficacy with these agents. We have previously shown that chromosome copy number variation (CNV) is associated with poor (14q loss) or more favorable outcomes (5q31-ter gain) in nonmetastatic patients. The purpose of the current study was the impact of CNV on response, progression free survival (PFS) and on overall survival (OS) after AA in patients with metastatic ccRCC. Methods: We obtained archival FFPE or frozen tumor specimens from patients with mRCC treated with sorafenib (after tumor removal), bevacizumab or sunitinib (neoadjuvant treatment). DNA was analyzed with Affymetrix 250K Nsp SNP microarrays. We identified the presence of genomic imbalances and loss of heterozygosity (LOH) to obtain “virtual karyotypes”. We then compared CNV to outcome using Wilcoxon-Gehan statistical analysis. Results: A total of 83 patients were used to analyze CNV after treatment with AA: 22 sorafenib, 29 sunitinib, 31 bevacizumab. Gain of 8q (p = 0.036) and loss of 16q (p = 0.0031), 20p (p = 0.038) or 20q (p = 0.022) were associated with a shorter OS, whereas gain of 1q (p = 0.037) and 5q (p = 0.019) were associated with longer OS in this patient cohort. When 14q loss was combined with 8q gain, median OS was further decreased (p 0.01). Surprisingly, when assessed as a group, no specific CNV was associated with PFS. However, when compared separately, 5q gain was predictive for better PFS in sorafenib or bevacizumab treated patients (p = 0.006), but no such effect was seen in the sunitinib treated cohort. Conclusions: Our results show that chromosomal imbalances are associated with divergent clinical outcome in metastatic ccRCC patients treated with AA, and are predominantly prognostic. Dissecting out driver tumor suppressor or tumor activating genes within these chromosomal regions will guide our understanding of what defines a particularly lethal phenotype. The possibility exists that specific CNV profiles predict for response to therapy, but a larger sample size is required to validate this possibility.

Correlation of chromosome (Chr) 14 loss and 5q gain with outcomes of pazopanib treatment in patients (pts) with metastatic clear cell renal cell carcinoma (mRCC).

4605 Background: The identification of molecular prognostic and/or predictive determinants of outcome in pts with mRCC is an important challenge. We hypothesized that specific tumor DNA copy number alterations (CNAs) - loss of chr 14 or 14q (14/14q-), and gain of chr 5q (5q+) may predict likelihood of pazopanib treatment benefit in pts with mRCC. Methods: Pts DNA samples from the Phase II (VEG102616) pazopanib trial were analyzed by using Affymetrix OncoScan. Copy no. data were moving smoothed and normalized. The copy no. and allele difference were profiled according to their chromosome locations to interrogate CNA and LOH. Objective response (OR, RECIST) and progression free survival (PFS) were determined by investigators (IN) and an independent review (IR) committee. OR and PFS data were analyzed using exact and Kaplan-Meier tests. Results: Tumor DNA samples from 75 pts were adequate for CNA analysis. 14/14q- was found in 35/75 pts (46.7%), and 5q+ was found in 37/75 (49.3%). 14/14q- was present in the tumors of 12/31 pts (39%) with OR by IR compared with 23/44 (52%) nonresponding pts (p=0.347). 14/14q- did not affect PFS. 5q+ was present in tumors of 17/31 pts (55%) with OR and 20/44 (45%) nonresponding pts (p=0.486). PFS was longer in pts with 5q+ tumors compared with those without 5q+ (log rank p=0.026), with median PFS of 66 vs 35 wks, respectively. The odds of achieving OR decreased as the total number of chromosomal gains/losses increased (p=0.032, odds ratio 0.49), but this had no effect on PFS. In exploratory analysis, we examined the combined effect of both 14/14q- and 5q+ on PFS (Table). Conclusions: Pts with 5q+ tumors have significantly longer PFS, with no effect on OR rate. While 14/14q- alone had no effect on outcomes, the combination of 5q+ and no 14/14q- was associated with significantly greater PFS. Pts with more genetically complex tumors were less likely to obtain OR with pazopanib. [Table: see text]

Copy number profiling in von hippel‐lindau disease renal cell carcinoma

Germline mutations in the VHL tumor suppressor gene cause von Hippel-Lindau (VHL) disease and somatic VHL mutations occur in the majority of clear cell renal cell carcinoma (cRCC). To compare copy number abnormalities (CNAs) between cRCC from VHL patients and sporadic cRCC cases without detectable somatic VHL mutations, we analyzed 34 cRCC with Affymetrix 250K arrays. To increase the power of the study, we then combined our results with those of a previously published study and compared CNAs in VHL and non-VHL related cRCC using the genomic identification of significant targets in cancer (GISTIC) program. In VHL, cRCC GISTIC analysis identified four statistically significant regions of copy number gain and four statistically significant regions of deletion that occurred in >10% of tumors analyzed. Sporadic cRCC without detectable VHL mutations had, on average, more copy number abnormalities than VHL cRCC though the most common regions of loss/gain (e.g., 3p and 14q loss and 5q gain) were present in both tumor sets. However, CNAs on chromosome arms 7p (gain) and 8p (loss) were only detected in VHL RCC. Although individual copy number abnormality peaks contained clear candidate cancer genes in some cases (e.g., the 3p loss peak in VHL cRCC contained only six genes including VHL), most peaks contained many genes. To date, only a minority of the candidate genes included in these peaks have been analyzed for mutation or epigenetic inactivation in cRCC but TNFRSF10C and DUSP4 map to the 8p region deleted in VHL cRCC and TP53 and HIF2A (EPAS1) mapped to CNA loss and gain peaks (chromosomes 17 and 2, respectively) detected in sporadic VHL wild-type cRCC.

113 GENOMIC PROFILING OF RENAL CELL CARCINOMA IN PATIENTS WITH END-STAGE RENAL DISEASE

You have accessJournal of UrologyKidney Cancer: Basic Research1 Apr 2011113 GENOMIC PROFILING OF RENAL CELL CARCINOMA IN PATIENTS WITH END-STAGE RENAL DISEASE Toru Inoue, Keiko Matsuura, Taichiro Yoshimoto, Takahiro Narimatsu, Fuminori Sato, Yoji Nagashima, Shingo Hatakeyama, Chikara Ohyama, Tomonori Habuchi, Masayuki Nakagawa, Masatsugu Moriyama, and Hiromitsu Mimata Toru InoueToru Inoue Oita, Japan More articles by this author , Keiko MatsuuraKeiko Matsuura Oita, Japan More articles by this author , Taichiro YoshimotoTaichiro Yoshimoto Tokyo, Japan More articles by this author , Takahiro NarimatsuTakahiro Narimatsu Oita, Japan More articles by this author , Fuminori SatoFuminori Sato Oita, Japan More articles by this author , Yoji NagashimaYoji Nagashima Yokohama, Japan More articles by this author , Shingo HatakeyamaShingo Hatakeyama Hirosaki, Japan More articles by this author , Chikara OhyamaChikara Ohyama Hirosaki, Japan More articles by this author , Tomonori HabuchiTomonori Habuchi Akita, Japan More articles by this author , Masayuki NakagawaMasayuki Nakagawa Kagoshima, Japan More articles by this author , Masatsugu MoriyamaMasatsugu Moriyama Oita, Japan More articles by this author , and Hiromitsu MimataHiromitsu Mimata Oita, Japan More articles by this author View All Author Informationhttps://doi.org/10.1016/j.juro.2011.02.179AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookTwitterLinked InEmail INTRODUCTION AND OBJECTIVES Patients with end-stage renal disease (ESRD) show an increased incidence of renal cell carcinoma (RCC) compared with the general population. RCCs in patients with ESRD (RCC-ESRD) have a wide variety of histological subtypes. The copy number aberrations (CNAs) of these RCC-ESRD have not been fully investigated. Therefore, we performed array comparative genomic hybridization (array CGH) to determine the CNAs of RCC-ESRD. METHODS A total of 65 patients with RCCs in ESRD were analyzed. Genomic DNA was extracted selectively from formalin-fixed, paraffin-embedded tissues using laser microdissection. High resolution array CGH was performed using Agilent Whole Human Genome 4x44K Oligo Micro Array Kit. RESULTS We analyzed 65 cases of RCC in ESRD (32 clear cell RCCs, 9 papillary RCCs, 17 ACD-associated RCCs, 3 clear cell-papillary RCCs, 1 chromophobe RCC, 1 MTSCC, 2 unclassified RCCs) by array CGH. Unsupervised hierarchical cluster analysis revealed that RCC-ESRD could be subclassified into two main groups (Cluster A and B). The cluster A contained mainly papillary RCC, ACD-associated RCC, clear cell-papillary RCC. These 3 histologic subtypes did not constitute any independent cluster, suggesting that they shared similar CNAs. On the other hand, the cluster B mainly contained clear cell RCC. Next, we analyzed the patterns of CNAs on each cluster and found that the cluster A showed similar CNA patterns with sporadic papillary RCC, such as 7pq gain and 16 pq gain, suggesting that, as a whole, CNAs of papillary RCC-ESRD resemble those of sporadic papillary RCC. On the other hand, the cluster B had similar CNA patterns with sporadic clear cell RCC including 3p, 14q losses, and 5q gain. Next, we compared the CNAs of papillary RCC, ACD-associated RCC and clear cell-papillary RCC, all of which were categorized in the cluster A. Interestingly, we found that all the three histologic subtypes had similar patterns of CNAs, which resemble those of sporadic papillary RCC, suggesting that they shared similar molecular mechanisms of the carcinogenesis of these tumors, although their histologic phenotypes were clearly different. Moreover, duration of dialysis in cluster A was significantly longer than that of cluster B (p=0.015). CONCLUSIONS RCCs-ESRD can be classified into two categories based on their CNAs. Papillary RCC-ESRD, ACD-associated RCC, and clear cell-papillary RCC may share similar oncogenic mechanism. © 2011 by American Urological Association Education and Research, Inc.FiguresReferencesRelatedDetails Volume 185Issue 4SApril 2011Page: e48 Peer Review Report Advertisement Copyright & Permissions© 2011 by American Urological Association Education and Research, Inc.MetricsAuthor Information Toru Inoue Oita, Japan More articles by this author Keiko Matsuura Oita, Japan More articles by this author Taichiro Yoshimoto Tokyo, Japan More articles by this author Takahiro Narimatsu Oita, Japan More articles by this author Fuminori Sato Oita, Japan More articles by this author Yoji Nagashima Yokohama, Japan More articles by this author Shingo Hatakeyama Hirosaki, Japan More articles by this author Chikara Ohyama Hirosaki, Japan More articles by this author Tomonori Habuchi Akita, Japan More articles by this author Masayuki Nakagawa Kagoshima, Japan More articles by this author Masatsugu Moriyama Oita, Japan More articles by this author Hiromitsu Mimata Oita, Japan More articles by this author Expand All Advertisement Advertisement PDF downloadLoading ...

Chromosomal imbalances as biomarkers for recurrence and antiangiogenic resistance in clear cell renal cell carcinoma.

e15012 Background: A key dilemma in the management of clear cell renal cell carcinoma (ccRCC) is the lack of reliable biomarkers for recurrence, prognosis, and prediction of resistance to anti-angiogenic therapy. Chromosome copy number alterations (CCNAs) by conventional cytogenetics have been shown to be associated with outcomes. In this study, the goal was to determine whether CCNAs identified with SNP arrays could be used as prognostic biomarkers for recurrence and predictors of antiangiogenic therapy response in ccRCC. Methods: We obtained archival formalin fixed paraffin embedded (FFPE) tumor samples from 32 patients with organ confined ccRCC, and 56 patients with metastatic ccRCC treated with sorafenib (after tumor removal, n=23) or bevacizumab (neoadjuvant treatment, n=33). DNA from the FFPE tissue sections was analyzed with Affymetrix 250K Nsp SNP microarrays to identify genomic CCNAs. Marker associations with tumor recurrence were evaluated using the Fisher's exact test. Logistic regression was used to evaluate the association between markers and response. Results: In the organ confined cohort, 14q loss showed a significant association with tumor recurrence (log rank P= 0.017), and a 3-locus model (gains of 1q or 12p or loss of 14q) showed a stronger association with recurrence (P= 0.0014). Gain of 5q was strongly associated with longer progression free survival (PFS) in both sorafenib and bevacizumab treated cohorts (HR = 0.25, 95% CI 0.08 to 0.81, P = 0.021 and HR = 0.82 95% CI 0.65- 1, P < 0.0001 respectively). In the bevacizumab cohort, 14q loss showed a significant association with worse response to treatment (CR or PR vs SD or PD, Fisher exact test, P = 0.0473). There was a nonsignificant trend toward a higher hazard of progression among patients who had loss of 9p or loss of 14q in the sorafenib cohort. Conclusions: Our results show that whole genome analysis with virtual karyotyping identifies genetic alterations associated with outcomes in ccRCC. These data suggest that genomic analysis of renal tumors with SNP arrays provides important prognostic information and may lead to a mechanistic understanding of progression and resistance to antiangiogenic therapy of ccRCC. Author Disclosure Employment or Leadership Position Consultant or Advisory Role Stock Ownership Honoraria Research Funding Expert Testimony Other Remuneration Bayer/Onyx, Genentech, iKaryos Diagnostics iKaryos Diagnostics Bayer/Onyx Genentech

Combined classical cytogenetics and microarray‐based genomic copy number analysis reveal frequent 3;5 rearrangements in clear cell renal cell carcinoma

Karyotypic analysis and genomic copy number analysis with single nucleotide polymorphism (SNP)-based microarrays were compared with regard to the detection of recurrent genomic imbalances in 20 clear cell renal cell carcinomas (ccRCCs). Genomic imbalances were identified in 19 of 20 tumors by DNA copy number analysis and in 15 tumors by classical cytogenetics. A statistically significant correlation was observed between the number of genomic imbalances and tumor stage. The most common genomic imbalances were loss of 3p and gain of 5q. Other recurrent genomic imbalances seen in at least 15% of tumors included losses of 1p32.3-p33, 6q23.1-qter and 14q and gain of chromosome 7. The SNP-based arrays revealed losses of 3p in 16 of 20 tumors, with the highest frequency being at 3p21.31-p22.1 and 3p24.3-p25.3, the latter encompassing the VHL locus. One other tumor showed uniparental disomy of chromosome 3. Thus, altogether loss of 3p was identified in 17 of 20 (85%) cases. Fourteen tumors showed both overlapping losses of 3p and overlapping gains of 5q, and the karyotypic assessment performed in parallel revealed that these imbalances arose via unbalanced 3;5 translocations. Among the latter, there were common regions of loss at 3p21.3-pter and gain at 5q34-qter. These data suggest that DNA copy number analysis will supplant karyotypic analysis of tumor types such as ccRCC that are characterized by recurrent genomic imbalances, rather than balanced rearrangements. These findings also suggest that the 5q duplication/3p deficiency resulting from unbalanced 3;5 translocations conveys a proliferative advantage of particular importance in ccRCC tumorigenesis.

CSF1R copy number changes, point mutations, and RNA and protein overexpression in renal cell carcinomas

Renal cell carcinomas comprise a heterogeneous group of tumors. Of these, 80% are clear cell renal cell carcinomas, which are characterized by loss of 3p, often with concomitant gain of 5q22qter. Although VHL is considered the main target gene of the 3p deletions, none has been identified as the relevant target gene for the 5q gain. We have studied 75 consecutive kidney tumors and paired normal kidney samples to evaluate at the genomic and expression levels the tyrosine kinase genes CSF1R and PDGFRB as potential targets in this region. Our findings show that RNA expression of CSF1R, but not of PDGFRB, was significantly higher in clear cell renal cell carcinomas than in normal tissue samples, something that was corroborated at the protein level by immunohistochemistry. The CSF1R staining pattern in clear cell renal cell carcinomas was clearly different from that observed in other renal cell carcinomas, suggesting its potential usefulness in differential diagnosis. FISH analysis demonstrated whole chromosomal gain and relative CSF1R/PDGFRB copy number gain in clear cell renal cell carcinomas, which might contribute to CSF1R overexpression. Finally, one polymorphism and two novel mutations were identified in CSF1R in clear cell renal cell carcinoma patients. Our data allow us to conclude that CSF1R plays a relevant role in clear cell renal cell carcinoma carcinogenesis and raise the possibility that CSF1R may represent a future valuable therapeutic target in these patients.

Cytogenetic and Molecular Tumor Profiling for Type 1 and Type 2 Papillary Renal Cell Carcinoma

The goal of this study was to evaluate immunohistochemical and cytogenetic features and their prognostic value in papillary renal cell carcinoma (PRCC) subtypes. One hundred fifty-eight cases of PRCC were identified and reclassified by subtype. Tumoral expression of 29 molecular markers was determined by immunohistochemistry. Cytogenetic analyses were done on a prospective series of 65 patients. Associations with clinicopathologic information and disease-specific survival were assessed. Fifty-one patients (32%) had type 1 and 107 (68%) type 2 PRCC. Type 2 patients had worse Eastern Cooperative Oncology Group performance status, higher T stages, nodal and distant metastases, higher grades, and a higher frequency of necrosis, collecting system invasion and sarcomatoid features. Type 2 showed greater expression of vascular endothelial growth factor (VEGF)-R2 in the tumor epithelium, and of VEGF-R3 in both tumor epithelium and endothelium. Loss of chromosome 1p, loss of 3p, and gain of 5q were exclusively observed in type 2, whereas type 1 more frequently had trisomy 17. Type 2 PRCC was associated with worse survival than type 1, but type was not retained as an independent prognostic factor. Lower PTEN, lower EpCAM, lower gelsolin, higher CAIX, and higher VEGF-R2 and VEGF-R3 expression, loss of 1p, 3p, or 9p, and absence trisomy 17 were all associated with poorer prognosis. Type 2 PRCC is associated with more aggressive clinicopathologic features and worse outcome. Molecular and chromosomal alterations can distinguish between PRCC subtypes and influence their prognosis. The effect of 3p loss on survival in PRCC is opposite to the relationship seen in clear cell RCC.

Cytogenetic Profile Predicts Prognosis of Patients With Clear Cell Renal Cell Carcinoma

The majority of cytogenetic studies in renal cell carcinoma (RCC) have been impaired by small sample size, retrospective character, and lack of a survival end point. We prospectively studied the prognostic impact of cytogenetic abnormalities on a larger cohort of patients having up to 108 months of follow-up. Tumors of 282 patients who underwent nephrectomy for clear cell RCC were cytogenetically analyzed. Results were correlated with pathological factors and disease-specific survival. The most frequently observed cytogenetic abnormalities were loss of 3p (60%), gain of 5q (33%), loss of 14q (28%), trisomy 7 (26%), loss of 8p (20%), loss of 6q (17%), loss of 9p (16%), loss of 4p (13%), and loss of chromosome Y in men (55%). Tumors with loss of 3p presented at lower TNM stages. Loss of 4p, 9p, and 14q were all associated with higher TNM stages, higher grade, and greater tumor size. A deletion of 3p was associated with better prognosis (P = .03), while loss of 4p (P < .001), loss of 9p (P < .01), and loss of 14q (P < .01) were each associated with worse prognosis. Loss of the Y chromosome led to improved progression-free survival in metastatic patients (P = .02). In multivariate analysis, loss of 9p was retained as an independent prognostic factor. This cytogenetic study serves as a proof of principal that genetic information, such as loss of chromosome 9, can be obtained from widely available technology, and can provide additional prognostic information to standard clinicopathologic variables.