Definition of a 1-Mb homozygous deletion at 9q32-q33 in a human bladder-cancer cell line

Fujiwara, Hiromichi; Emi, Mitsuru; Nagai, Hisaki; Ohgaki, Kenji; Imoto, Issei; Akimoto, Masao; Ogawa, Osamu; Habuchi, Tomonori

doi:10.1007/s100380170056

Cited by 13 publications

(10 citation statements)

References 20 publications

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“…Although no mutations have yet been identified in this gene, methylation-based silencing of expression has been found in bladder cancer cell lines and primary tumors (Fujiwara et al, 2001). The finding in several studies of homozygous deletions including the DBCCR1 gene (Nishiyama et al, 1999;Fujiwara et al, 2001;Stadler et al, 2001) provides additional evidence for a tumor suppressor in this region. In this study, we confirmed the homozygous deletion of DBCCR1 in a cell line derived from one such tumor, DSH1, but none was detected in any of the other lines.…”

Section: Discussionmentioning

confidence: 98%

“…A third region spanning 9q13-31 contains the Gorlin syndrome gene PTCH. Common mutation of DBCCR1 has not been identified in bladder tumors, but frequently the gene is silenced by methylation, and homozygous deletion of the gene has been identified Nishiyama et al, 1999;Fujiwara et al, 2001;Stadler et al, 2001). Mutations of TSC1 and PTCH have been identified at low frequency in bladder tumors (McGarvey et al, 1998;Hornigold et al, 1999;van Tilborg et al, 2001), but none of these genes is altered at the same frequency as chromosome 9 LOH is found, leading to uncertainty about the role of different chromosome 9 genes or combinations of genes in different bladder tumors.…”

Section: Introductionmentioning

confidence: 95%

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Molecular genetic analysis of chromosome 9 candidate tumor‐suppressor loci in bladder cancer cell lines

Williams

Sibley

Davies

et al. 2002

Genes Chromosomes & Cancer

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Underrepresentation of chromosome 9 is a common finding in bladder cancer. Frequent loss of the whole chromosome suggests the presence of at least one relevant tumor suppressor gene on each arm. Candidate regions identified by loss of heterozygosity (LOH) analysis include a region at 9p21 containing CDKN2A, which encodes p16 and p14(ARF), a large region at 9q12-31 including PTCH and many other genes, a small region at 9q32-33, which includes the DBCCR1 gene, and a region at 9q34 including the TSC1 gene. Experimental replacement of genes or chromosomes into tumor cells with appropriate deletions or mutations represents an important approach to test the functional significance of candidate tumor suppressor genes. Loss of an entire copy of chromosome 9 in many bladder tumor cell lines provides no indication of which gene or genes are affected, and selection of appropriate recipient cells for gene replacement is difficult. We have investigated three candidate tumor suppressor genes on chromosome 9 (CDKN2A, DBCCR1, and TSC1), at the DNA level and by expression analysis in a panel of bladder tumor cell lines, many of which have probable LOH along the length of the chromosome, as indicated by homozygosity for multiple polymorphic markers. Cytogenetically, we found no reduction in the numbers of chromosomes 9 relative to total chromosome count. Homozygous deletion of the CDKN2A locus was frequent but homozygous deletion of TSC1 was not found. A new cell line, DSH1, derived from a pT1G2 transitional cell carcinoma with known homozygous deletion of DBCCR1, is described. This study identifies suitable cell lines for future functional analysis of both CDKN2A and DBCCR1.

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Section: Discussionmentioning

confidence: 98%

Section: Introductionmentioning

confidence: 95%

Molecular genetic analysis of chromosome 9 candidate tumor‐suppressor loci in bladder cancer cell lines

Williams

Sibley

Davies

et al. 2002

Genes Chromosomes & Cancer

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“…Hypermethylation of the DBCCR1 gene as well as LOH and homozygous deletions at the DBCCR1 locus have been shown to be frequent events in bladder cancer (Fujiwara et al, 2001;Habuchi et al, 1998Habuchi et al, , 2001Nishiyama et al, 1999). Previous studies of head and neck carcinomas have demonstrated LOH involving the 9q32-33 region, which covers the DBCCR1 gene (Ah-See et al, 1994).…”

Section: Discussionmentioning

confidence: 99%

Loss of heterozygosity at 9q33 and hypermethylation of the DBCCR1 gene in oral squamous cell carcinoma

Worm

Guldberg

et al. 2004

Br J Cancer

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The DBCCR1 gene at chromosome 9q33 has been identified as a candidate tumour suppressor, which is frequently targeted by promoter hypermethylation in bladder cancer. Here, we studied the possible involvement of DBCCR1 in the development of oral squamous cell carcinoma. DNA from 34 tumours was examined for loss of heterozygosity (LOH) at three markers surrounding DBCCR1 and for hypermethylation of the DBCCR1 promoter, using methylation-specific PCR and methylation-specific melting-curve analysis. LOH was found in 10 of 31 cases (32%), and DBCCR1 hypermethylation was present in 15 of 34 cases (44%). Hypermethylation of DBCCR1 was also present in three of seven epithelial tissues adjacent to the tumours, including two hyperplastic and one histologically normal epithelia. Furthermore, of four oral leukoplakias with dysplasia, one showed LOH at 9q33 and two showed DBCCR1 hypermethylation. These data suggest that LOH at 9q33 and hypermethylation of the DBCCR1 promoter are frequent and possibly early events in oral malignant development. Oral cancer comprises about 3% of all newly diagnosed cancer cases in the Western countries. Despite advances in therapy, the 5-year survival rate after diagnosis is still poor and remains B50% (Landis et al, 1999;Silverman, 2001). Clinically, oral carcinomas often develop in a two-step process. The first step is characterised by the appearance of potentially malignant lesions such as leukoplakias and erythroplakias, and the second step is characterised by the development of carcinomas. However, clinical and histopathological features are insufficient measures for predicting the prognosis of potentially malignant lesions (Warnakulasuriya, 2000(Warnakulasuriya, , 2001. Furthermore, a recent study indicated that clinically and histologically normal mucosa adjacent to tumours may harbour patches of genetically altered cells (Braakhuis et al, 2003). It is, therefore, important to find molecular markers for identifying the minor fraction of oral lesions that will develop into carcinoma.Loss of heterozygosity (LOH) at multiple chromosome regions and genetic and epigenetic alterations of several proto-oncogenes and tumour suppressor genes have been demonstrated in oral carcinomas, including alterations of the TP53, p16, p15, MGMT, Ecadherin genes and RAS (Califano et al, 1996;Partridge et al, 1999;Williams, 2000;Ogi et al, 2002;Viswanathan et al, 2003). In addition, our previous study showed that hypermethylation of the ABO gene promoter was associated with loss of expression of A/B antigen in approximately one-third of oral squamous cell carcinomas (Gao et al, 2004). LOH at 9q34, in which the ABO gene is located, was also a frequent event in these tumours. However, a number of tumours from AO and BO heterozygotes showed deletion of the O allele, which does not encode a functional glycosyltransferase, suggesting the existence of an additional tumour suppressor gene on chromosome 9q. The DBCCR1 (deleted in bladder cancer chromosomal region candidate 1) gene at chromosome 9q33 was identified as...

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“…In addition to LOH studies, evidence supporting the hypothesis that DBCCR1 is a tumor suppressor gene includes chromosome copy number changes detected by fluorescence in situ hybridization (FISH) (Stadler et al, 2001), absence of DBCCR1 mRNA expression in tumor cell lines and bladder tumors, and the existence of a bladder tumor (Nishiyama et al, 1999) and a bladder tumor cell line with homozygous deletions at the DBCCR1 locus (9p33) (Fujiwara et al, 2001).Transfection and expression of DBCCR1 in NIH/3T3 cells resulted in an increase in the proportion of cells in the G1 phase of the cell cycle (Nishiyama et al, 2001). This phenomenon was observable only under serum starvation conditions and the mechanism of action remains unclear.…”

Section: Introductionmentioning

confidence: 99%

DBCCR1 mediates death in cultured bladder tumor cells

2004

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Chromosome 9, which is often partially or fully reduced to homozygosity in bladder cancer cells, harbors several tumor suppressor loci including deleted in bladder cancer chromosome region 1 (DBCCR1) at 9q32-33. To study DBCCR1 function, stable cell lines, inducible for DBCCR1 expression by tetracycline, were made, but the DBCCR1 protein was not expressed at detectable levels. To understand the fate of DBCCR1-expressing cells, human bladder tumor cells were transiently transfected with an expression vector containing DBCCR1 fused to enhanced green fluorescent protein (EGFP). Initially, DBCCR1-EGFP-expressing cells demonstrated diffuse cytoplasmic green fluorescence with nuclear exclusion patterns. After time, the intensity level of green fluorescence increased and a granular distribution of protein became visible in the cells. At this point, cells rounded up and detached from the tissue culture dish. Cells transfected with a control vector, containing only EGFP, and partial DBCCR1-EGFP fusion constructs did not demonstrate this behavior. DBCCR1-mediated cell death in cultured tumor cells was independent of caspase-3 activation and did not result in detectable DNA strand breaks by TUNEL staining that are hallmarks of the classical apoptotic pathway.

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Definition of a 1-Mb homozygous deletion at 9q32-q33 in a human bladder-cancer cell line

Cited by 13 publications

References 20 publications

Molecular genetic analysis of chromosome 9 candidate tumor‐suppressor loci in bladder cancer cell lines

Molecular genetic analysis of chromosome 9 candidate tumor‐suppressor loci in bladder cancer cell lines

Loss of heterozygosity at 9q33 and hypermethylation of the DBCCR1 gene in oral squamous cell carcinoma

DBCCR1 mediates death in cultured bladder tumor cells

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