Gain of a region on 7p22.3, containing MAD1L1, is the most frequent event in small-cell lung cancer cell lines

Small-cell lung cancer (SCLC) is a highly aggressive lung neoplasm, which accounts for 20% of yearly lung cancer cases. The lack of knowledge of the progenitor cell type for SCLC precludes the definition of a normal gene expression profile and has hampered the identification of gene expression changes, while the low resolution of conventional genomic screens such as comparative genomic hybridization (CGH) and loss of heterozygosity analysis limit our ability to fine-map genetic alterations. The recent advent of whole genome tiling path array CGH enables profiling of segmental DNA copy number gains and losses at a resolution 100 times that of conventional methods. Here we report the analysis of 14 SCLC cell lines and six matched normal B-lymphocyte lines. We detected 7p22.3 copy number gain in 13 of the 14 SCLC lines and 0 of the 6 matched normal lines. In 4 of the 14 cell lines, this gain is present as a 350 kbp gene specific copy number gain centered at MAD1L1 (the human homologue of the yeast gene MAD1). Fluorescence in situ hybridization validated the array CGH finding. Intriguingly, MAD1L1 has been implicated to have tumor-suppressing functions. Our data suggest a more complex role for this gene, as MAD1L1 is the most frequent copy number gain in SCLC cell lines.     

Identification of novel regions of altered DNA copy number in small cell lung tumors

Identification of the genetic alterations that occur in tumors is an important approach to understanding tumorigenesis. We have used comparative genomic hybridization (CGH), a novel molecular cytogenetic method, to identify the gross DNA copy number changes that commonly occur in small cell lung cancer (SCLC). We analyzed ten SCLC tumors (seven primary tumors and three metastases) from eight patients. We found frequent increases in DNA copy number on chromosome arms Sp, 8q, 3q, and Xq and frequent decreases in copy number on chromosome arms 3p. 17p, Sq, 8p, 13q, and 4p. The increase in copy number at 8q24 (MYC) and decreases at 17p 13 (TP53), 13q 14 (RB), and 3p have previously been identified in SCLC with other methods. Many of the other regions in which we detected common copy number changes have not been reported to be regions of common alteration in SCLC tumors. Comparison of copy number changes between a primary tumor and a metastasis from the same patient showed that they were more closely related to each other than to any of the other tumors. The results of direct CGH analysis of SCLC tumors reported here confirm the existence of copy number changes that we identified previously by using cell lines. © 1995 Wiley-Liss, Inc.     

Recurrent allelic deletions of chromosome arms 15q and 16q in human small cell lung carcinomas

The genetic lesions that lead to the development of small cell lung carcinoma (SCLC) remain incompletely defined. To identify recurrent allelic deletions in specific chromosomal regions that could serve as markers for tumor suppressor gene (TSG) inactivation in SCLC, we performed a comprehensive allelotype analysis of all 39 nonacrocentric autosomal arms. Alterations in 158 polymorphic microsatellite alleles were examined in 24 pairs of human SCLC tumor and normal control DNA samples. A total of 2,107 informative reactions were analyzed. This analysis revealed allelic losses of 100% on chromosome arm 3p, >85% loss within chromosome arms 13q and 17p, and >70% loss within chromosome arms 4q, 5q, 15q, and 16q. The allelic deletions on chromosome arms 15q and 16q have not been defined previously for SCLC and are candidate regions to harbor novel TSGs. Genes Chromosomes Cancer 27:323-331, 2000.     

Molecular cytogenetic characterization of human papillomavirus16-transformed foreskin keratinocyte cell line 16-MT

Anogenital cancers are closely associated with human papillomavirus (HPV), and HPV-infected individuals, particularly those with high-grade dysplasias, are at increased risk for cervical and anal cancers. Although genomic instability has been documented in HPV-infected keratinocytes, the full spectrum of genetic changes in HPV-associated lesions has not been fully defined. To address this, we examined an HPV16-transformed foreskin keratinocyte cell line, 16-MT, by GTG-banding, spectral karyotyping (SKY), and array comparative genomic hybridization (array CGH); these analyses revealed multiple numerical, complex, and cryptic chromosome rearrangements. Based on GTG-banding, the 16-MT karyotype was interpreted as 78-83,XXY,+add(1)(p36.3),+3,+4,+5,+5,+7,+8,+i(8)(q10)x2,+10,?der(12),der(13;14)(q10;q10),+15,+16,add(19)(q13.3),+21,+21,-22[cp20]. Multicolor analysis by SKY confirmed and further characterized the anomalies identified by GTG banding. The add(1) was identified as a der(1)(1qter-->1q25::1p36.1-->1qter), the add(19) as a dup(19), and the der(12) interpreted as a der(11) involving a duplication of chromosome 11 material and rearrangement with chromosome 19. In addition, previously unidentified der(9)t(9;22), der(3)t(3;19), and der(4)t(4;9) were noted. The 16-MT cell line showed losses and gains of DNA due to unbalanced translocations and complex rearrangements of regions containing known tumor suppressor genes. Chromosomal changes in these regions might explain the increased risk of cancer associated with HPV. Also, array CGH detected copy-number gains or amplifications of chromosomes 2, 8, 10, and 11 and deletions of chromosomes 3, 4, 11, and 15. These results provide the basis for the identification of candidate oncogenes responsible for cervical and anal cancer in amplified regions, and for putative tumor suppressor genes in commonly deleted regions like 11q22-23. Furthermore, these data represent the first full characterization of the HPV-positive cell line 16-MT.     

Spectrum of genetic changes in gastro-esophageal cancer cell lines determined by an integrated molecular cytogenetic approach

Adenocarcinomas arising around the gastro-esophageal junction (GEJ) are highly malignant, and their incidence has risen rapidly in the last decades. Cell lines are the basic in vitro system for functional and therapeutic studies in GEJ tumors, but only a small number of cell lines are currently available, and none of them has been fully karyotyped. We analyzed 5 GEJ tumor cell lines using a combination of 24-color fluorescence in situ hybridization (FISH), comparative genomic hybridization (CGH) and genomic microarrays. Using CGH we demonstrated that these cell lines present imbalances similar to those we had previously observed in primary GEJ tumors, namely gains on 1q, 7q, 8q, 17q, 19q, 20, and X, and losses on 3p, 4, 5q, 9p, 18q, and 21. Multicolor FISH karyotyping revealed multiple structural rearrangements involving chromosomes 1, 5, 6, 7, 8, 9, 13, 17, 18, and 22. Rearrangements of chromosome 8 involved 10 different chromosomes, while rearrangements of chromosome 17 involved 5. Different rearrangements resulted in imbalances of similar chromosome regions, suggesting that similar genomic imbalances are constitutively important but are achieved through different pathways. The use of a commercially available genomic array excluded TOP2A (17q), and MYBL2, PTPT1, CSE1L, and ZNF217 (20q) as candidate genes for frequently amplified areas on these chromosomes, and contributed to refining the limits of chromosome regions involved in genomic imbalances.     

The application of comparative genomic hybridization as an additional tool in the chromosome analysis of acute myeloid leukemia and myelodysplastic syndromes

In acute myeloid leukemia (AML) and myelodysplastic syndromes (MDS) there are frequently complex karyotypes with multiple structurally altered chromosomes, many of which are marker chromosomes of unknown origin. The aim of this study was to apply comparative genomic hybridization (CGH) to cases of AML or MDS in transformation submitted for routine cytogenetic analysis to investigate whether this approach would yield any further information and, if possible, to predict which cases would benefit from CGH analysis. Nineteen cases with AML or MDS in transformation were analyzed. CGH revealed nine cases with gains or losses of chromosomal material. In six of these cases the chromosomal location of this material was not apparent from cytogenetic analysis especially when multiple markers were present. By using fluorescence in situ hybridization (FISH) with specific libraries for the chromosome regions that showed discordance between CGH and conventional cytogenetics, we were able to identify the chromosome location of material within the karyotype. In this group of six patients, four cases of an unbalanced translocation involving regions of chromosomes 5 and 17 were characterized. Three of these cases had additional abnormalities, including two cases with regions of amplification in which oncogenes are located (MYC, MLL) and one case with a dic(7;21)(p10;p10). In all six cases it was possible to characterize complex chromosomal aberrations such as derivative chromosomes, marker chromosomes, and ring chromosomes. This study demonstrates that CGH can detect true gain and loss of critical chromosome regions more accurately than conventional karyotyping in cases with very complex karyotypes, and can thus prove useful in predicting prognosis and pinpointing areas of the genome that require further study. Also, CGH can be a useful technique to identify the origin of marker chromosomes, and it can assist in choice of probes for confirmatory FISH, when there is no clue provided from the analysis of G-banded chromosomes.     

Detection of chromosomal DNA gains and losses in testicular germ cell tumors by comparative genomic hybridization

To extend the results of conventional cytogenetic analysis of testicular germ cell tumors (TGCTs), we applied the new molecular cytogenetic method of comparative genomic hybridization (CGH), which enables the detection of chromosomal imbalances without the need for dividing cells. DNA from II TGCTs was studied by CGH. In all tumors examined, gain of 12p, mostly of the whole p arm, could be demonstrated. However, in three tumors, an amplification of 12p material restricted to the chromosomal bands 12p11.2-p12.1 was found. Further fluorescence in situ hybridization (FISH) analysis using a yeast artificial chromosome (YAC) that was previously mapped to that region revealed multiple copies of that chromosomal segment in interphase nuclei of these tumors. This finding is an important clue to the localization of candidate protooncogenes at 12p involved in TGCTs. Gains of small chromosomal regions at 2p, 4q, 6p, and 19p were also detected recurrently. Furthermore, gains of chromosomes 8, 14, 21, and X as well as loss of chromosome 13 were frequent findings. In conclusion, CGH provides new insights into genetic alterations of TGCTs. By using CGH, chromosomal subregions could be identified that may harbor genes involved in the pathogenesis of this malignancy. Genes Chromosom Cancer 17:78–87 (1996). © 1996 Wiley-Liss, Inc.     

Detection of amplified oncogenes by genome DNA microarrays in human primary esophageal squamous cell carcinoma: comparison with conventional comparative genomic hybridization analysis

Oncogene amplification in 20 surgically resected esophageal squamous cell carcinomas (ESCC) was examined with DNA microarrays that detected 57 oncogenes and two reference DNA. Alterations in DNA copy numbers detected by microarrays were compared to those obtained by conventional comparative genomic hybridization (CGH). Amplification of eight oncogenes (CCND1, FGF3/FGF4, EMS1, SAS, ERBB2, PDGFRA, MYC, and BCL2) was detected by DNA microarrays in 9 of 20 tumors. Although ERBB2 was 23.2 times higher than the control level in one case, the average magnitude of gene amplification was approximately two to four times that of the control level. EMS1, CCND1, and FGF3/FGF4, which are all located on 11q13, were amplified in 7, 5, and 4 of 20 ESCC, respectively, and they were coamplified in 3 tumors. A comparison of genome DNA microarrays and CGH data revealed that although most amplified oncogenes were included in chromosomal regions for which DNA copy number gains were detected by conventional CGH, not all amplified genes on microarrays showed concomitant DNA copy number gains on CGH. In conclusion, microarrays of oncogenes are useful for the comprehensive identification of amplified oncogenes and for analysis of areas of specific amplification within chromosomal regions with DNA copy number increases detected by CGH analysis.     

Frequent gain of chromosome 19 in megakaryoblastic leukemias detected by comparative genomic hybridization

Acute megakaryocytic leukemia is a rare subtype of AML that is often difficult to diagnose; it is most commonly associated with Down syndrome in children. To identify chromosomal imbalances and rearrangements associated with acute megakaryocytic leukemia, we used G-banding, comparative genomic hybridization (CGH), and whole chromosome painting (WCP) on a variety of primary patients' samples and leukemia cell lines. The most common abnormality was gain of chromosome 19 or arm 19q, which was detected by CGH in four of 12 (33.3%) primary samples and nine of 11 (81.8%) cell lines. In none of the primary samples was this abnormality detected by G-banding analysis. WCP was used to define further the nature of the chromosome 19 gain in the cell lines, which was found to be due to the presence of additional 19q material on marker chromosomes or to cryptic translocations involving 19q. The most common chromosomal loss--detected only in the cell lines--was deletion of chromosomal band 13q14, which was seen in six of 11 (54.5%) cell lines. Other recurrent changes included gains of 1p, 6p, 8q, 11q, 15q, 17q, and 21q and losses of 2, 4q, 5q, 7q, 9p, and 11p. Combining conventional and molecular cytogenetic analyses defined recurrent clonal chromosomal abnormalities, which will aid in the identification of critical genes that are abnormal in acute megakaryocytic leukemia cells.     

Examination of oncogene amplification by genomic DNA microarray in hepatocellular carcinomas: comparison with comparative genomic hybridization analysis

To identify amplified oncogenes involved in hepatocellular carcinomas (HCC), we applied a genomic DNA microarray spotted with 57 oncogenes to 20 HCCs. Aberrations in DNA copy number also were analyzed by comparative genomic hybridization (CGH) using an aliquot of DNA samples. In 5 of 20 HCCs, only 6 oncogenes (CCND1, FGF3/FGF4, SAS/CDK4, TERC, MET, and MYC) were amplified, whereas in the remaining 15 tumors no oncogenes were amplified. A comparison of DNA microarray and conventional CGH analyses showed that, although 5 of 6 amplified oncogenes shown by microarray were located in chromosomal regions shown by CGH to have increased DNA copy numbers, not all genes located in such chromosomal regions were affected. One of the amplified oncogenes (SAS/CDK4) was found in a chromosomal region that was undetected by CGH. We, therefore, conclude that amplification of the oncogenes examined in this series is not directly implicated in hepatocellular carcinogenesis.     

A cytogeneticist's perspective on genomic microarrays

The identification of cytogenetic imbalance is an important component of clinical genetics. About 1 in 154 newborns has a chromosome abnormality. Conventional cytogenetic analysis has enabled the identification of microscopic alterations of the chromosomes. The development of fluorescence in situ hybridization (FISH) and other molecular methodologies has made possible the identification of submicroscopic aberrations. An additional development was comparative genomic hybridization (CGH), a method that directly compares two genomes for DNA copy differences. As first developed, the substrate for CGH analysis is normal metaphase chromosomes. Recently, CGH has been applied to microarrays (array CGH) constructed from large insert clones to identify chromosome imbalance. Array CGH has many advantages over conventional cytogenetic and molecular cytogenetic techniques. Array CGH can be comprehensive (genome-wide), high resolution, amenable to automation, rapid, and sensitive. We anticipate that array CGH will be employed in the clinical cytogenetics laboratory in the near future and will lead to the identification of the chromosomal basis of new syndromes and existing genetic conditions.     

Frequent gain of copy number on the long arm of chromosome 20 in human pancreatic adenocarcinoma

We have used comparative genomic hybridization (CGH) to survey genomic regions with aberrant copy numbers of DNA sequences in pancreatic adenocarcinoma. In 12 cell lines and 6 primary tumors from 18 patients with pancreatic adenocarcinomas, highly frequent losses (>60%) were observed on chromosome arms 6q, 9p, and 18q and the Y chromosome. Moderately frequent losses (40–60%) were observed on chromosome arms 3p, 4q, 8p, and 21q. Interestingly, these samples showed extremely high frequencies of increases in copy numbers of DNA sequences on the long arm of chromosome 20 (15/18, 83%). We further analyzed five cell lines by fluorescence in situ hybridization (FISH) with probes on chromosome 20 to define the increase in copy number more accurately, and we found that 20q was increased to between 5 and 8 copies per cell. These results suggest the existence of an oncogene or oncogenes on 20q that play a role in the development and/or the progression of pancreatic carcinogenesis. Genes Chromosom. Cancer 19:161–169, 1997. © 1997 Wiley-Liss Inc.     

Complex CGH alterations on chromosome arm 8p at candidate tumor suppressor gene loci in breast cancer cell lines

Loss of genetic material from chromosome arm 8p occurs frequently in human breast carcinomas, consistent with this region of the genome harboring one or more tumor suppressor genes (TSGs). We used the complementary techniques of microsatellite-based LOH, high-density FISH, and conventional CGH on 6 breast cancer cell lines (MCF7, SKBR3, T47D, MDA MB453, BT549, and BT474) to investigate the molecular cytogenetic changes occurring on chromosome 8 during tumorigenesis, with particular emphasis on 6 potential TSGs on 8p. We identified multiple alterations of chromosome 8, including partial or complete deletion of 8p or 8q, duplication of 8q, and isochromosome 8q. The detailed FISH analysis showed several complex rearrangements of 8p with differing breakpoints of varying proximity to the genes of interest. High rates of LOH were observed at markers adjacent to or within PCM1, DUSP4/MKP2, NKX3A, and DLC1, supporting their status as candidate TSGs. Due to the complex ploidy status of these cell lines, relative loss of 8p material detected by CGH did not always correlate with microsatellite-based LOH results. These results extend our understanding of the mechanisms accompanying the dysregulation of candidate tumor suppressor loci on chromosome arm 8p, and identify appropriate cellular systems for further investigation of their biological properties.     

Development of a high-density pericentromeric region BAC clone set for the detection and characterization of small supernumerary marker chromosomes by array CGH.

PURPOSE: Small supernumerary marker chromosomes are centric chromosomal segments that, by definition, cannot be characterized unambiguously by conventional chromosome banding. Marker chromosomes are of particular interest in clinical cytogenetics because they are nearly 10 times more frequent in individuals with mental retardation (0.426%) than in the normal population (0.043%). However, they are often found in only a small percentage of cells, making them difficult to detect and characterize in a diagnostic setting. We designed, constructed, and employed a bacterial artificial chromosome (BAC)-based microarray to demonstrate the utility of array-based comparative genomic hybridization (array CGH) for detecting and characterizing marker chromosomes in clinical diagnostic specimens. METHODS: We constructed a high-density microarray using 974 BAC clones that were mapped by fluorescence in situ hybridization and cover approximately 5 Mb of the most proximal unique sequence adjacent to the centromere on all 43 unique pericentromeric regions of the human genome (excluding the acrocentric short arms). This array was used to further characterize 20 previously identified marker chromosomes that were originally found with either conventional chromosome analysis or a targeted microarray. RESULTS: The enhanced coverage of this pericentromeric array not only identified the chromosomal origin of each marker in 15 cases, it also distinguished between the involvement of the short arm and/or the long arm of each chromosome, defined the sizes of many of the markers, and revealed complex rearrangements or multiple markers in single individuals. However, in five cases, the markers could not be identified by this assay, most likely because of very low levels of mosaicism and/or their small size and lack of detectable euchromatin. The expanded coverage of the pericentromeric regions represented on the array was adequate to refine the breakpoints in two-thirds of all cases in which a marker chromosome was identified by this assay. CONCLUSIONS: This study demonstrates the utility of array CGH in the detection and characterization of mosaic marker chromosomes. Because approximately one-third of the markers characterized in this study involved more unique sequence than that represented on this array, additional pericentromeric coverage may be even more valuable. We anticipate that this will allow detailed characterization of small supernumerary marker chromosomes that will greatly facilitate phenotype/genotype correlations and play a valuable role in the diagnosis and medical management of both pre- and postnatal cases in which marker chromosomes have been identified.     

Comprehensive whole genome array CGH profiling of mantle cell lymphoma model genomes

Mantle cell lymphoma (MCL) is an aggressive non-Hodgkin's lymphoma with median patient survival times of approximately 3 years. Although the characteristic t(11;14)(q13;q32) is found in virtually all cases, experimental evidence suggests that this event alone is insufficient to result in lymphoma and secondary genomic alterations are required. Using a newly developed DNA microarray of 32 433 overlapping genomic segments spanning the entire human genome, we can for the first time move beyond marker based analysis and comprehensively search for secondary genomic alterations concomitant with the t(11;14) in eight commonly used cell models of MCL (Granta-519, HBL-2, NCEB-1, Rec-1, SP49, UPN-1, Z138C and JVM-2). Examining these genomes at tiling resolution identified an unexpected average of 35 genetic alterations per cell line, with equal numbers of amplifications and deletions. Recurrent high-level amplifications were identified at 18q21 containing BCL2, and at 13q31 containing GPC5. In addition, a recurrent homozygous deletion was identified at 9p21 containing p15 and p16. Alignment of these profiles revealed 14 recurrent losses and 21 recurrent gains as small as 130 kb. Remarkably, even the intra immunoglobulin gene deletions at 2p11 and 22q11 were detected, demonstrating the power of combining the detection sensitivity of array comparative genomic hybridization (CGH) with the resolution of an overlapping whole genome tiling-set. These alterations not only coincided with previously described aberrations in MCL, but also defined 13 novel regions. Further characterization of such minimally altered genomic regions identified using whole genome array CGH will define novel dominant oncogenes and tumor suppressor genes that play important roles in the pathogenesis of MCL.     

Chromosome 8 genetic analysis and phenotypic characterization of 21 ovarian cancer cell lines

The short arm of chromosome 8 undergoes frequent loss of heterozygosity (LOH) in ovarian adenocarcinomas. Fine mapping has identified several distinct critical regions within 8p which undergo rates of LOH of 50% or greater, suggesting that there may be more than one tumor suppressor gene located on this chromosome arm. In an effort to refine the location of these putative tumor suppressor genes by homozygosity-mapping-of-deletion analysis, we have analyzed 21 ovarian cancer cell lines with 19 polymorphic microsatellite markers from 8p. Eleven of the cell lines (55%) were homozygous at every marker, indicating loss of an entire 8p arm. No smaller extended regions of hemizygosity were identified. Refinement of these 8p target regions was therefore not possible, but this analysis did identify the ovarian cancer cell lines that would be most appropriate for microcell-mediated chromosome transfer to complement the hypothesized mutation in the target tumor suppressor gene(s) on 8p. The 11 cell lines that had undergone 8p LOH were therefore characterized for colony formation in soft agar and tumor formation in nude mice. We identified four cell lines (JAM, OVCA4, OVCA5, and OVCA8) that were hemizygous for 8p and that formed colonies in soft agar and tumors in nude mice, making them ideal cell lines for chromosome 8 or candidate gene transfer.     

A nonrandom chromosomal abnormality, del 3p(14-23), in human small cell lung cancer (SCLC)

In order to determine whether or not there are specific chromosomal changes in small cell lung cancer (SCLC), karyotypic analyses of 16 continuous SCLC tissue culture lines, three fresh tumor specimens (bone marrow), one direct preparation of bone marrow involved with SCLC, and two lymphoblastoid lines derived from SCLC patients were studied. Cell lines were derived from primary tumor, or metastases to bone marrow, subcutaneous nodules, or pleural fluid; all 16 lines had biochemical and histologic properties characteristic of SCLC. Of the 15 males and 3 females, 6 patients had no prior treatment. All of the 16 cell lines, the 3 fresh specimens, and the direct bone marrow preparation had a common deletion of the short arm of chromosome #3. Use of the shortest region of overlap analysis showed the common deletion was of the short arm in the regions p(14-23). This specific chromosomal abnormality, del 3p, was not found in five non-SCLC cell lines studied and is of major potential biological and diagnostic importance.     

Y chromosome loss and other genomic alterations in hepatocellular carcinoma cell lines analyzed by CGH and CGH array

Hepatocellular carcinoma (HCC) is one of the most frequently occurring malignant tumors worldwide. The incidence of HCC is much higher in males than in females. In order to clarify the molecular basis of the male predominance in HCC, we have characterized the detailed genomic alterations in 5 hepatitis B virus integrated Korean HCC cell lines using G-banding, comparative genomic hybridization (CGH), fluorescence in situ hybridization (FISH), PCR, and CGH array. The commonest alterations were observed in chromosome 7 and Y, as well as chromosomal regions 1q, 8q, 4q, and 16q. The most frequent aberration of genomic material was gain of 1q and loss of chromosome Y. Significant loss of DNA copy number of the cancer related genes that are located on chromosome Y was detected by CGH array. By investigating the karyotypes of the previously reported 21 male HCC cell lines, we found 18 HCC cell lines with Y chromosome loss, indicating that this loss is a significant feature of HCC cell lines. We propose that Y chromosome loss in HCC cell lines may be responsible for the preponderance of males in HCC and its significance may lead to further studies for better understanding of carcinogenesis in HCC.     

Determination of amplicon boundaries at 20q13.2 in tissue samples of human gastric adenocarcinomas by high-resolution microarray comparative genomic hybridization

Comparative genomic hybridization (CGH) of gastric adenocarcinomas frequently shows gains and amplifications of chromosome 20. However, the underlying genetic lesion is unknown and conventional CGH results do not allow specification of the target region. In order to investigate this chromosomal aberration with a higher resolution and sensitivity, microarray-based CGH was performed with both scanning and high-resolution arrays of chromosome 20 in a series of 27 gastric adenocarcinomas. Locus-specific fragments of genomic DNA from bacterial artificial chromosome (BAC) clones were spotted as microarrays. A scanning array contained a set of 27 BAC clones covering chromosome 20q. A high-resolution array contained 27 overlapping BAC clones at 20q13.2. This high-resolution array was used to narrow down the amplicon at 20q13.2 in tumours showing amplification of this chromosomal region with the scanning array. Positive copy number changes on chromosome 20q were detected in 12 of 27 cases (44%). These changes included gain of the whole arm of chromosome 20q in 8 of 27 (30%) cases, amplification restricted to 20q12.1 in one case, and amplifications restricted to 20q13 in three cases (11%). The three tumours showing amplification restricted to 20q13 were analysed further using the high-resolution array. In one tumour, the whole contig was amplified at a constant level. One of the other two tumours had a clear proximal breakpoint, while the other tumour had a clear distal breakpoint within the 20q13.2 region. The proximal and the distal breakpoint were approximately 800 kb apart. In the present study, an amplicon at 20q13.2 has been narrowed down to 800 kb which is likely to harbour one or more putative oncogenes relevant to gastric carcinogenesis, for which ZNF217 and CYP24 are good candidates.