Integrated genomic and transcriptional analysis of the in vitro evolution of telomerase-immortalized urothelial cells (TERT-NHUC).
Much progress has been made in identifying the molecular genetic alterations that occur in bladder cancer. However, in many cases the genes targeted by these alterations are not known. Telomerase immortalized human urothelial cells (TERT-NHUC) are a useful resource for in vitro studies of genes involved in urothelial transformation. When cultured under standard conditions they remain genetically stable but when cultured under low-density conditions they exhibit genetic instability and acquire chromosomal alterations. TERT-NHUC from three donors were cultured at low plating density and examined at four time-points during a culture period of 600 days. Analyses included population doubling kinetics, array-based CGH (aCGH), chromosome counts, fluorescence in situ hybridization (FISH), mutation analysis, Affymetrix gene expression analysis, Western blotting for p16, anchorage-independent growth and tumorigenicity assays. Alterations acquired during continued culture of TERT-NHUC at low density (TERT-NHUC-L) included some observed in urothelial carcinoma (UC) cell lines and primary UC. Examination of gene expression in TERT-NHUC with distinct acquired genetic aberrations may pinpoint genes targeted by these alterations. Data from an aCGH study of UC cell lines and primary tumors were examined for changes in chromosomal regions that also showed alterations in TERT-NHUC-L. Loss of a region on 2q including BOK was identified in UC cell lines and primary tumors. DNER and FRAS1 were identified as potential candidate genes, whose expression is altered independently of the acquisition of any genetic event.
Chapman
Emma J
EJ
Cancer Research UK Clinical Centre, St James's University Hospital, Leeds LS97TF, UK.
Williams
Sarah V
SV
Platt
Fiona M
FM
Hurst
Carolyn D
CD
Chambers
Philip
P
Roberts
Paul
P
Knowles
Margaret A
MA
D002452:Cell Count; D018929:Cell Culture Techniques; D045744:Cell Line, Tumor; D049109:Cell Proliferation; D002478:Cells, Cultured; D002869:Chromosome Aberrations; D055028:Comparative Genomic Hybridization; D019941:Cyclin-Dependent Kinase Inhibitor p16 / Q000235:genetics / Q000378:metabolism; D018628:Gene Dosage; D015870:Gene Expression; D020869:Gene Expression Profiling / Q000379:methods*; D006801:Humans; D019656:Loss of Heterozygosity; D010641:Phenotype; D011003:Ploidies; D011518:Proto-Oncogene Proteins / Q000235:genetics / Q000378:metabolism; D019098:Telomerase / Q000235:genetics* / Q000378:metabolism; D001749:Urinary Bladder Neoplasms / Q000235:genetics* / Q000378:metabolism; D019459:Urothelium / Q000166:cytology* / Q000378:metabolism*
D019941:Cyclin-Dependent Kinase Inhibitor p16; D011518:Proto-Oncogene Proteins; D019098:Telomerase