Microarray-based CGH in human lymphoma cell lines.. Homo sapiens

This series represents the data set of Array CGH from the paper (submit for publication): “Homozygous deletions localize novel tumor suppressor genes in B-cell lymphomas. Mestre et all. ” and it's web supplement. The molecular nature of many secondary events in the pathogenesis of B-cell non Hodgkin lymphoma (B-NHL) remains unknown. We used high-resolution CGH to BAC microarrays to characterize the genomes of 48 B-cell non-Hodgkin lymphoma (B-NHL) cell lines of different origins. Array CGH identified, on average, 20 genomic alterations per cell line, including regional gains and losses as well as previously uncharacterized aberrations. Different genomic patterns were observed among the B-NHL subtypes. To search for possible oncogenic target genes, gene expression profiling was performed in the cell lines models. Integrative genomic and gene expression analyses have identified amplified oncogenes in B-cell non-Hodgkin lymphoma (B-NHL), but the capability of such technologies to localize tumor suppressor genes within homozygous deletions remains unexplored. Array-based comparative genomic hybridization (CGH) and gene expression microarray analysis of 48 cell lines derived from patients with different B-NHLs delineated twenty homozygous deletions at seven chromosome areas, all of which contained tumor suppressor gene targets. Further investigation revealed that only a fraction of primary biopsies presented inactivation of these genes by point mutation or intragenic deletion, but instead some of them were frequently silenced by epigenetic mechanisms. Our microarray strategy has identified novel candidate suppressors inactivated by genetic and epigenetic mechanisms that substantially vary among the B-NHL subtypes. Keywords: Array CGH, B-cell non-Hodgkin lymphoma, genomic amplification, homozygous deletion Overall design: Genome-wide analysis of DNA-copy number changes of the 48 cell lines was performed using array CGH on a microchip with 2.400 BAC and P1 clones in triplicate. The array provides an average resolution of 1.4 Mb across the genome. Production and validation of the array, hybridization methods and analytical procedures have been described elsewhere in detail.( Snijder AM, Nowak N, Segraves R, et al. Assembly of microarrays for genome-wide measurement of DNA copy number. Nat Genet. 2001;29:263-264.) Briefly, 0.2 µg of test (tumor) and reference genomic DNAs were labeled by random priming using Cy3 and Cy5, respectively. After 48 hour hybridization, slides were washed and mounted with DAPI. The images of the arrays were captured using a CCD camera, and the “UCSF SPOT” software was used to analyze the images. A second program “SPROC” was used to associate clones with each spot and a mapping information file that allows the data to be plotted relative to the position of the BACs on the draft human genome sequence (http://genome.cse.ucsc.edu; July 2003 freeze). A formal data filtering procedure was then performed, and a SPROC output file consisting of averaged ratios of the triplicate spots for each clone, standard deviations of the replicates and plotting positions for each clone on the array, was obtained. For visualization of genomic data, the TreeView program 1.60 (Stanford, CA) was used. Mean log2 ratios (tumoral DNA vs. control DNA) were plotted and the resultant graphs were converted to the log2 domain. The observed log2 ratios were excluded from further analysis if there were fewer than two replicate spots (out of 3) or if the standard deviation of the replicates was above 0.2. The clones that were present in fewer than 50% of the samples were also removed from the dataset. The following mean log2 ratios (tumoral DNA vs. control DNA) were used to classify genomic aberrations: genomic gain (between 0.39 and 0.95), genomic amplification (greater than 0.95) or genomic loss (lower than -0.5 for heterozygosity, and lower than –1.4 for homozygous deletion). Statistically significant correlations between the genomic imbalances and the B-NHL subgroups were analyzed using Fisher’s exact test. For delineation of common regions of genomic imbalance in the lymphoma genomes, the position of the BAC/PACs on the draft human genome sequence according to the May 2004 freeze was used (http://genome.cse.ucsc.edu; May 2004 freeze).