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Copy number alterations in urothelial carcinomas: their clinicopathological significance and correlation with DNA methylation alterations.

Nishiyama N, Arai E, Nagashio R, Fujimoto H, Hosoda F, Shibata T, Tsukamoto T, Yokoi S, Imoto I, Inazawa J, Kanai Y - Carcinogenesis (2010)

Bottom Line: Losses of 1p32.2-p31.3, 10q11.23-q21.1 and 15q21.3 were correlated with tumor recurrence.Unsupervised hierarchical clustering analysis based on copy number alterations clustered UCs into three subclasses: copy number alterations associated with genome-wide DNA hypomethylation, regional DNA hypermethylation on C-type CpG islands and genome-wide DNA hypo- and hypermethylation were accumulated in clusters A, B(1) and B(2), respectively.Both genetic and epigenetic events appear to accumulate during urothelial carcinogenesis, reflecting the clinicopathological diversity of UCs.

View Article: PubMed Central - PubMed

Affiliation: Pathology Division, National Cancer Center Research Institute, Tokyo 104-0045, Japan.

ABSTRACT
The aim of this study was to clarify the genetic backgrounds underlying the clinicopathological characteristics of urothelial carcinomas (UCs). Array comparative genomic hybridization analysis using a 244K oligonucleotide array was performed on 49 samples of UC tissue. Losses of 2q33.3-q37.3, 4p15.2-q13.1 and 5q13.3-q35.3 and gains of 7p11.2-q11.23 and 20q13.12-q13.2 were correlated with higher histological grade, and gain of 7p21.2-p21.12 was correlated with deeper invasion. Losses of 6q14.1-q27 and 17p13.3-q11.1 and gains of 19q13.12-q13.2 and 20q13.12-q13.33 were correlated with lymph vessel involvement. Loss of 16p12.2-p12.1 and gain of 3q26.32-q29 were correlated with vascular involvement. Losses of 5q14.1-q23.1, 6q14.1-q27, 8p22-p21.3, 11q13.5-q14.1 and 15q11.2-q22.2 and gains of 7p11.2-q11.22 and 19q13.12-q13.2 were correlated with the development of aggressive non-papillary UCs. Losses of 1p32.2-p31.3, 10q11.23-q21.1 and 15q21.3 were correlated with tumor recurrence. Unsupervised hierarchical clustering analysis based on copy number alterations clustered UCs into three subclasses: copy number alterations associated with genome-wide DNA hypomethylation, regional DNA hypermethylation on C-type CpG islands and genome-wide DNA hypo- and hypermethylation were accumulated in clusters A, B(1) and B(2), respectively. Tumor-related genes that may encode therapeutic targets and/or indicators useful for the diagnosis and prognostication of UCs should be explored in the above regions. Both genetic and epigenetic events appear to accumulate during urothelial carcinogenesis, reflecting the clinicopathological diversity of UCs.

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Validation of array CGH analysis by FISH. (A) Array CGH profiles of representative tissue specimens (T1 to T4). The signal ratios of the CDKN2 locus in T1, T2 and T3 corresponded to copy numbers of 0, 1 and 2, respectively, whereas the signal ratio in T4 did not correspond to any whole number. (B) Although the LSI p16 (9p21) SpectrumOrange/CEP 9 SpectrumGreen Probe corresponding to the CDKN2A gene revealed two signals in stromal cells and adjacent non-cancerous urothelial cells, it revealed no signal in cancer cells in T1. (C) FISH analysis using the same probe revealed one signal in cancer cells in T2. (D) FISH analysis using the same probe revealed two signals in cancer cells in T3. (E) FISH analysis using the same probe revealed copy number heterogeneity in T4: cancer cells in areas 1 and 2 showed two signals and one signal within a tumor, respectively. These findings can explain the array CGH profile of T4 in panel (A).
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fig1: Validation of array CGH analysis by FISH. (A) Array CGH profiles of representative tissue specimens (T1 to T4). The signal ratios of the CDKN2 locus in T1, T2 and T3 corresponded to copy numbers of 0, 1 and 2, respectively, whereas the signal ratio in T4 did not correspond to any whole number. (B) Although the LSI p16 (9p21) SpectrumOrange/CEP 9 SpectrumGreen Probe corresponding to the CDKN2A gene revealed two signals in stromal cells and adjacent non-cancerous urothelial cells, it revealed no signal in cancer cells in T1. (C) FISH analysis using the same probe revealed one signal in cancer cells in T2. (D) FISH analysis using the same probe revealed two signals in cancer cells in T3. (E) FISH analysis using the same probe revealed copy number heterogeneity in T4: cancer cells in areas 1 and 2 showed two signals and one signal within a tumor, respectively. These findings can explain the array CGH profile of T4 in panel (A).

Mentions: The array CGH analysis for copy number alterations was validated by FISH. Examples of array CGH profiles and FISH images of the four representative UCs (T1 to T4) are shown in Figure 1A–E, respectively. The signal ratios of the CDKN2A locus in T1, T2 and T3 corresponded to copy numbers of 0, 1 and 2, respectively, whereas the signal ratio in T4 did not correspond to any whole numbers (Figure 1A). The LSI p16 (9p21) SpectrumOrange/CEP 9 SpectrumGreen Probe corresponding to the CDKN2A gene revealed two signals in stromal cells and adjacent non-cancerous urothelial cells on the specimen of T1 (Figure 1B). The probe revealed zero, one and two signals in cancer cells in T1, T2 and T3, respectively (Figure 1B–D). FISH analysis revealed copy number heterogeneity within a UC: cancer cells showing two signals and those showing one signal were both observed in T4 (Figure 1E). These findings were able to explain the array CGH profile in T4 (Figure 1A). Similarly FISH analysis using the LSI p53 (17p13.1) SpectrumOrange Probe corresponding to the TP53 gene also validated the array CGH profiles (data not shown).


Copy number alterations in urothelial carcinomas: their clinicopathological significance and correlation with DNA methylation alterations.

Nishiyama N, Arai E, Nagashio R, Fujimoto H, Hosoda F, Shibata T, Tsukamoto T, Yokoi S, Imoto I, Inazawa J, Kanai Y - Carcinogenesis (2010)

Validation of array CGH analysis by FISH. (A) Array CGH profiles of representative tissue specimens (T1 to T4). The signal ratios of the CDKN2 locus in T1, T2 and T3 corresponded to copy numbers of 0, 1 and 2, respectively, whereas the signal ratio in T4 did not correspond to any whole number. (B) Although the LSI p16 (9p21) SpectrumOrange/CEP 9 SpectrumGreen Probe corresponding to the CDKN2A gene revealed two signals in stromal cells and adjacent non-cancerous urothelial cells, it revealed no signal in cancer cells in T1. (C) FISH analysis using the same probe revealed one signal in cancer cells in T2. (D) FISH analysis using the same probe revealed two signals in cancer cells in T3. (E) FISH analysis using the same probe revealed copy number heterogeneity in T4: cancer cells in areas 1 and 2 showed two signals and one signal within a tumor, respectively. These findings can explain the array CGH profile of T4 in panel (A).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3066412&req=5

fig1: Validation of array CGH analysis by FISH. (A) Array CGH profiles of representative tissue specimens (T1 to T4). The signal ratios of the CDKN2 locus in T1, T2 and T3 corresponded to copy numbers of 0, 1 and 2, respectively, whereas the signal ratio in T4 did not correspond to any whole number. (B) Although the LSI p16 (9p21) SpectrumOrange/CEP 9 SpectrumGreen Probe corresponding to the CDKN2A gene revealed two signals in stromal cells and adjacent non-cancerous urothelial cells, it revealed no signal in cancer cells in T1. (C) FISH analysis using the same probe revealed one signal in cancer cells in T2. (D) FISH analysis using the same probe revealed two signals in cancer cells in T3. (E) FISH analysis using the same probe revealed copy number heterogeneity in T4: cancer cells in areas 1 and 2 showed two signals and one signal within a tumor, respectively. These findings can explain the array CGH profile of T4 in panel (A).
Mentions: The array CGH analysis for copy number alterations was validated by FISH. Examples of array CGH profiles and FISH images of the four representative UCs (T1 to T4) are shown in Figure 1A–E, respectively. The signal ratios of the CDKN2A locus in T1, T2 and T3 corresponded to copy numbers of 0, 1 and 2, respectively, whereas the signal ratio in T4 did not correspond to any whole numbers (Figure 1A). The LSI p16 (9p21) SpectrumOrange/CEP 9 SpectrumGreen Probe corresponding to the CDKN2A gene revealed two signals in stromal cells and adjacent non-cancerous urothelial cells on the specimen of T1 (Figure 1B). The probe revealed zero, one and two signals in cancer cells in T1, T2 and T3, respectively (Figure 1B–D). FISH analysis revealed copy number heterogeneity within a UC: cancer cells showing two signals and those showing one signal were both observed in T4 (Figure 1E). These findings were able to explain the array CGH profile in T4 (Figure 1A). Similarly FISH analysis using the LSI p53 (17p13.1) SpectrumOrange Probe corresponding to the TP53 gene also validated the array CGH profiles (data not shown).

Bottom Line: Losses of 1p32.2-p31.3, 10q11.23-q21.1 and 15q21.3 were correlated with tumor recurrence.Unsupervised hierarchical clustering analysis based on copy number alterations clustered UCs into three subclasses: copy number alterations associated with genome-wide DNA hypomethylation, regional DNA hypermethylation on C-type CpG islands and genome-wide DNA hypo- and hypermethylation were accumulated in clusters A, B(1) and B(2), respectively.Both genetic and epigenetic events appear to accumulate during urothelial carcinogenesis, reflecting the clinicopathological diversity of UCs.

View Article: PubMed Central - PubMed

Affiliation: Pathology Division, National Cancer Center Research Institute, Tokyo 104-0045, Japan.

ABSTRACT
The aim of this study was to clarify the genetic backgrounds underlying the clinicopathological characteristics of urothelial carcinomas (UCs). Array comparative genomic hybridization analysis using a 244K oligonucleotide array was performed on 49 samples of UC tissue. Losses of 2q33.3-q37.3, 4p15.2-q13.1 and 5q13.3-q35.3 and gains of 7p11.2-q11.23 and 20q13.12-q13.2 were correlated with higher histological grade, and gain of 7p21.2-p21.12 was correlated with deeper invasion. Losses of 6q14.1-q27 and 17p13.3-q11.1 and gains of 19q13.12-q13.2 and 20q13.12-q13.33 were correlated with lymph vessel involvement. Loss of 16p12.2-p12.1 and gain of 3q26.32-q29 were correlated with vascular involvement. Losses of 5q14.1-q23.1, 6q14.1-q27, 8p22-p21.3, 11q13.5-q14.1 and 15q11.2-q22.2 and gains of 7p11.2-q11.22 and 19q13.12-q13.2 were correlated with the development of aggressive non-papillary UCs. Losses of 1p32.2-p31.3, 10q11.23-q21.1 and 15q21.3 were correlated with tumor recurrence. Unsupervised hierarchical clustering analysis based on copy number alterations clustered UCs into three subclasses: copy number alterations associated with genome-wide DNA hypomethylation, regional DNA hypermethylation on C-type CpG islands and genome-wide DNA hypo- and hypermethylation were accumulated in clusters A, B(1) and B(2), respectively. Tumor-related genes that may encode therapeutic targets and/or indicators useful for the diagnosis and prognostication of UCs should be explored in the above regions. Both genetic and epigenetic events appear to accumulate during urothelial carcinogenesis, reflecting the clinicopathological diversity of UCs.

Show MeSH
Related in: MedlinePlus