Identification and Validation of Suitable Endogenous Reference Genes for Gene Expression Studies of Human Bladder Cancer

Ohl, Falk; Jung, Monika; Radonić, Aleksandar; Sachs, Murray B.; Loening, Stefan A.; Jung, Klaus

doi:10.1016/s0022-5347(05)00919-5

Cited by 106 publications

(106 citation statements)

References 20 publications

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“…Further, using the eight genes as internal reference, we observed that the prognostic accuracy of B2M was 0.82 (95% CI ¼ 0.63 -0.94; data not shown). In a separate study conducted with a small set of tissues (n ¼ 14), we observed that B2M maintained its high prognostic accuracy for lymph node metastases (AUC ¼ 0.79) when more classical reference control genes TBP (Ohl et al, 2006) or UBP (Andersen et al, 2004) were substituted for Spint2 (not shown). Area under the curve values were obtained using the mean C t value of the patient sample as an internal reference.…”

Section: Map7mentioning

confidence: 90%

β2microglobulin mRNA expression levels are prognostic for lymph node metastasis in colorectal cancer patients

et al. 2008

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Colorectal cancer (CRC) is the fourth most common non-cutaneous malignancy in the United States and the second most frequent cause of cancer-related death. One of the most important determinants of CRC survival is lymph node metastasis. To determine whether molecular markers might be prognostic for lymph node metastases, we measured by quantitative real-time RT -PCR the expression levels of 15 cancer-associated genes in formalin-fixed paraffin-embedded primary tissues derived from stage I -IV CRC patients with (n ¼ 20) and without (n ¼ 18) nodal metastases. Using the mean of the 15 genes as an internal reference control, we observed that low expression of b 2 microglobulin (B2M) was a strong prognostic indicator of lymph node metastases (area under the curve (AUC) ¼ 0.85; 95% confidence interval (CI) ¼ 0.69 -0.94). We also observed that the expression ratio of B2M/Spint2 had the highest prognostic accuracy (AUC ¼ 0.87; 95% CI ¼ 0.71 -0.96) of all potential two-gene combinations. Expression values of Spint2 correlated with the mean of the entire gene set at an R 2 value of 0.97, providing evidence that Spint2 serves not as an independent prognostic gene, but rather as a reliable reference control gene. These studies are the first to demonstrate a prognostic role of B2M at the mRNA level and suggest that low B2M expression levels might be useful for identifying patients with lymph node metastasis and/or poor survival.

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Section: Map7mentioning

confidence: 90%

β2microglobulin mRNA expression levels are prognostic for lymph node metastasis in colorectal cancer patients

et al. 2008

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“…This issue was investigated by Ohl et al and found that the most commonly used endogenous control GAPDH is not stably expressed in bladder cancer. They found TATA box-binding protein and succinate dehydrogenase complex, subunit A, the most appropriate endogenous control in bladder caner (17). Therefore, we used TATA box-binding protein to normalize our target genes expression.…”

Section: Methodsmentioning

confidence: 99%

Angiogenic Switch of Angiopietins-Tie2 System and Its Prognostic Value in Bladder Cancer

Szarvas

Jäger

Tötsch³

et al. 2008

Clinical Cancer Research

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Purpose: Vascular endothelial growth factor (VEGF), angiopoietins (Ang-1and Ang-2), and their receptor Tie2 are critically involved in both normal and pathologic angiogenesis. The aim of this study was to explore the role of Ang-1, Ang-2,VEGF, andTie2 in the development and progression of bladder cancer as well as to examine their prognostic value in this tumor type. Experimental Design:Tumor samples of 113 bladder cancer patients, normal bladder epithelium of 5 noncancer patients, and two low-grade (UMUC3 and RT4) and two high-grade (J82 and T24) bladder cancer cell lines were analyzed by quantitative real-time PCR. The expression data were analyzed performing Wilcoxon rank-sum and Kaplan-Meier log-rank tests as well as univariate Cox analyses and Cox proportional hazards regression model. Results: In tissues of noninvasive bladder tumors, Ang-1 expression was significantly lower (P < 0.001), whereasVEGF expression was significantly higher (P = 0.031) than in normal bladder tissue. These findings were also confirmed at the protein level by immunohistochemistry. In contrast, Tie2 and Ang-2 abundance in tumor did not differ significantly from that in normal bladder tissue. Multivariate analysis identified Ang-2 as a strong and independent predictor of tumor recurrence [hazard ratio (HR), 10.18; 95% confidence interval (95% CI), 2.69-38.49; P < 0.001] andTie2 expression as an independent favorable prognostic factor for both metastasis (HR, 0.31; 95% CI, 0.11-0.89; P = 0.029) and disease-specific survival (HR, 0.25; 95% CI, 0.10-0.62; P = 0.003). Conclusions: These data show the strongest change in expression of VEGF and Ang-1 in superficial bladder cancer in comparison with normal bladder epithelium and the invasive tumor stages. The prognostic significance of Ang-2 and Tie2 underlines the essential role of angiopoietins-Tie2 system in progression of bladder cancer.

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“…Cycling parameters were as follows: initial denaturation at 95°C for 12 min; 45 cycles of 1 s at 95°C for denaturation, 5 s at 62°C for annealing and 10 s at 72 °C for extension. Primer sequences were as follows: Axin2-FW: 5'-agtcagcagagggacaggaa-3', Axin2-RW; 5'-agctctgagccttcagcatc-3'; G6PD-FW: 5'-atcgaccactacctgggcaa-3', G6PD-RW: 5'-ttctgcatcacgtcccgga-3' [19]. All primer sequences exclude amplification of genomic DNA under the parameters described above.…”

Section: Quantitative Real-time Rt-pcrmentioning

confidence: 99%

Quantitative and kinetic profile of Wnt/β-catenin signaling components during human neural progenitor cell differentiation

Mazemondet¹,

Hübner

Frahm

et al. 2011

Cellular and Molecular Biology Letters

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ReNcell VM is an immortalized human neural progenitor cell line with the ability to differentiate in vitro into astrocytes and neurons, in which the Wnt/β-catenin pathway is known to be involved. However, little is known about kinetic changes of this pathway in human neural progenitor cell differentiation. In the present study, we provide a quantitative profile of Wnt/β-catenin pathway dynamics showing its spatio-temporal regulation during ReNcell VM cell differentiation. We show first that T-cell factor dependent transcription can be activated by stabilized β-catenin. Furthermore, endogenous Wnt ligands, pathway receptors and signaling molecules are temporally controlled, demonstrating changes related to differentiation stages. During the first three hours of differentiation the signaling molecules LRP6, Dvl2 and β-catenin are spatio-temporally regulated between distinct cellular compartments. From 24 h onward, components of the Wnt/β-catenin pathway are strongly activated and regulated as shown by mRNA up-regulation of Wnt ligands (Wnt5a and Wnt7a), receptors including Frizzled-2, -3, -6, -7, and -9, and co-receptors, and target genes including Axin2. This detailed temporal profile of the Wnt/β-catenin pathway is a first step to understand, control and to orientate, in vitro, human neural progenitor cell differentiation.

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Identification and Validation of Suitable Endogenous Reference Genes for Gene Expression Studies of Human Bladder Cancer

Abstract: For normalization purposes in gene profiling studies of bladder cancer the genes SDH and TBP are recommended as single reference genes depending on the expression level of the target gene or more favorably in combination.

Cited by 106 publications

References 20 publications

β2microglobulin mRNA expression levels are prognostic for lymph node metastasis in colorectal cancer patients

β2microglobulin mRNA expression levels are prognostic for lymph node metastasis in colorectal cancer patients

Angiogenic Switch of Angiopietins-Tie2 System and Its Prognostic Value in Bladder Cancer

Quantitative and kinetic profile of Wnt/β-catenin signaling components during human neural progenitor cell differentiation

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