Genome-wide genetic characterization of bladder cancer: a comparison of high-density single-nucleotide polymorphism arrays and PCR-based microsatellite analysis.

Most human cancers are characterized by genomic instability, the accumulation of multiple genetic alterations, and allelic imbalance throughout the genome. Loss of heterozygosity (LOH) is a common form of allelic imbalance, and the detection of LOH has been used to identify genomic regions that harbor tumor suppressor genes and to characterize different tumor types, pathological stages and progression. Global patterns of LOH can be discerned by allelotyping of tumors with polymorphic genetic markers. Microsatellites are reliable genetic markers for studying LOH, but typically only a modest number of microsatellites are tested in LOH studies because the genotyping procedure can be laborious. Here we describe the use of a new alternative approach to comprehensive allelotyping in which samples are genotyped for nearly 1500 single-nucleotide polymorphism (SNP) loci distributed across all human autosomal arms. We examined the pattern of allelic imbalance in human transitional cell carcinomas of the urinary bladder including 36 primary tumors and 1 recurrent tumor with matched normal DNAs. The call rate for all SNPs was 78.5 +/- 1.87% overall samples. Overall, the median number of allelic imbalance was 47.5, ranging from 20 to 118. The mean number of allelic imbalances was 36.58, 51.30, and 67.78 for pT(a), pT(1), and > or =pT(2), respectively, and also increased by grade. The SNP microarray analysis result was validated by comparison with microsatellite allelotype analysis of 118 markers in the same tumors. Overall, the two methods produced consistent loss patterns at informative loci. The SNP assay discovered previously undiscovered allelic imbalances at chromosomal arms 12q, 16p, 1p, and 2q. The detection of LOH and other chromosomal changes using large numbers of SNP markers should enable rapid and accurate identification of allelic imbalance patterns that will facilitate the mapping and identification of important cancer genes. Moreover, SNP analysis raises the possibility of individual tumor genome-wide allelotyping with potential prognostic and diagnostic applications.

[1]  C. Greer,et al.  PCR amplification from paraffin-embedded tissues: recommendations on fixatives for long-term storage and prospective studies. , 1991, PCR methods and applications.

[2]  C. Busch,et al.  Comparison of comparative genomic hybridization, fluorescence in situ hybridization and flow cytometry in urinary bladder cancer. , 1999, Anticancer Research.

[3]  Z. Gibas,et al.  Cytogenetics of bladder cancer. , 1997, Cancer genetics and cytogenetics.

[4]  R. Knuechel,et al.  Occurrence of chromosome 9 and p53 alterations in multifocal dysplasia and carcinoma in situ of human urinary bladder. , 2002, Cancer research.

[5]  M. Knowles,et al.  Deletion mapping implicates two tumor suppressor genes on chromosome 8p in the development of bladder cancer. , 1996, Oncogene.

[6]  H. Moch,et al.  Patterns of chromosomal imbalances in advanced urinary bladder cancer detected by comparative genomic hybridization. , 1998, The American journal of pathology.

[7]  B. Vogelstein,et al.  A genetic model for colorectal tumorigenesis , 1990, Cell.

[8]  S. Devries,et al.  Identification of gains and losses of DNA sequences in primary bladder cancer by comparative genomic hybridization , 1995, Genes, chromosomes & cancer.

[9]  R. Blelloch,et al.  A molecular genetic model of human bladder cancer pathogenesis. , 1996, Seminars in oncology.

[10]  T. Polascik,et al.  Novel suppressor loci on chromosome 14q in primary bladder cancer. , 1995, Cancer research.

[11]  Eric S. Lander,et al.  Loss-of-heterozygosity analysis of small-cell lung carcinomas using single-nucleotide polymorphism arrays , 2000, Nature Biotechnology.

[12]  D J Lockhart,et al.  Genome-wide detection of allelic imbalance using human SNPs and high-density DNA arrays. , 2000, Genome research.

[13]  G. Gyapay,et al.  A second-generation linkage map of the human genome , 1992, Nature.

[14]  Ash A. Alizadeh,et al.  Genome-wide analysis of DNA copy-number changes using cDNA microarrays , 1999, Nature Genetics.

[15]  Kathleen R. Cho,et al.  Frequency of homozygous deletion at p16/CDKN2 in primary human tumours , 1995, Nature Genetics.

[16]  J. Cairns,et al.  A comparison between microsatellite and quantitative PCR analyses to detect frequent p16 copy number changes in primary bladder tumors. , 1998, Clinical cancer research : an official journal of the American Association for Cancer Research.

[17]  Dirk Zaak,et al.  Clonality and Genetic Divergence in Multifocal Low-Grade Superficial Urothelial Carcinoma as Determined by Chromosome 9 and p53 Deletion Analysis , 2000, Laboratory Investigation.

[18]  O. Kallioniemi,et al.  Genome screening by comparative genomic hybridization. , 1997, Trends in genetics : TIG.

[19]  T. Koyanagi,et al.  Gelsolin: a candidate for suppressor of human bladder cancer. , 1995, Cancer research.

[20]  B. Reid,et al.  Single nucleotide polymorphism array analysis of flow-sorted epithelial cells from frozen versus fixed tissues for whole genome analysis of allelic loss in breast cancer. , 2002, The American journal of pathology.

[21]  M. Knowles,et al.  Preliminary mapping of the deleted region of chromosome 9 in bladder cancer. , 1993, Cancer research.

[22]  David Sidransky,et al.  Microsatellite alterations in serum DNA of head and neck cancer patients , 1996, Nature Medicine.

[23]  C. Nusbaum,et al.  Large-scale identification, mapping, and genotyping of single-nucleotide polymorphisms in the human genome. , 1998, Science.

[24]  C. Ihm,et al.  Detection of genetic alterations in bladder tumors by comparative genomic hybridization and cytogenetic analysis. , 1999, Cancer genetics and cytogenetics.

[25]  C. Cordon-Cardo,et al.  Chromosome 9 allelic losses and microsatellite alterations in human bladder tumors. , 1994, Cancer research.

[26]  D. Sidransky,et al.  Rates of p16 (MTS1) mutations in primary tumors with 9p loss. , 1994, Science.

[27]  M. Williamson,et al.  p16 (CDKN2) is a major deletion target at 9p21 in bladder cancer. , 1995, Human molecular genetics.

[28]  Leonid Kruglyak,et al.  The use of a genetic map of biallelic markers in linkage studies , 1997, Nature Genetics.

[29]  H. von der Maase,et al.  Allelic imbalances in human bladder cancer: genome-wide detection with high-density single-nucleotide polymorphism arrays. , 2002, Journal of the National Cancer Institute.

[30]  Y. Nakamura,et al.  Allelotype of colorectal carcinomas. , 1989, Science.

[31]  J. Rossi,et al.  Effect of fixation on the amplification of nucleic acids from paraffin-embedded material by the polymerase chain reaction. , 1991, The journal of histochemistry and cytochemistry : official journal of the Histochemistry Society.

[32]  P. Nichols,et al.  Allelic losses of chromosomes 9, 11, and 17 in human bladder cancer. , 1990, Cancer research.

[33]  P. Devilee,et al.  PCR-based microsatellite polymorphisms in the detection of loss of heterozygosity in fresh and archival tumour tissue. , 1993, British Journal of Cancer.

[34]  A. Knudson.  Hereditary cancer, oncogenes, and antioncogenes. , 1985, Cancer research.