Detection of amplified DNA sequences by reverse chromosome painting using genomic tumor DNA as probe

A modification of reverse chromosome painting was carried out using genomic DNA from tumor cells as a complex probe for chromosomal in situ suppression hybridization to normal metaphase chromsome spreads. Amplified DNA sequences contained in such probes showed specific signals, revealing the normal chromosome positions from which these sequences were derived. As a model system, genomic DNAs were analyzed from three tumor cell lines with amplification units including the proto-oncogene c-myc. The smallest amplification unit was about 90 kb and was present in 16–24 copies; the largest unit was bigger than 600 kb and was present in 16–32 copies. Specific signals that co-localized with a differently labeled c-myc probe on chromosome band 8q24 were obtained with genomic DNA from each cell line. In further experiments, genomic DNA derived from primary tumor material was used in the case of a male patient with glioblastoma multiforme (GBM). Southern blot analysis using an epidermal growth factor receptor gene (EGFR) probe that maps to 7p13 indicated the amplification of sequences from this gene. Using reverse chromosome painting, signals were found both on band 7p13 and bands 12q13–q15. Notably, the signal on 12q13–q15 was consistently stronger. The weaker 7p13 signal showed co-localization with the major signal of the differently labeled EGFR probe. A minor signal of this probe was seen on 12q13, suggesting cross-hybridization to ERB3 sequences homologous to EGFR. The results indicate co-amplification of sequences from bands 12q13–q15, in addition to sequences from band 7p13. Several oncogenes map to 12q13–q15 providing candidate genes for a tumor-associated proto-oncogene amplification. Although the nature of the amplified sequences needs to be clarified, this experiment demonstrates the potential of reverse chromosome painting with genomic tumor DNA for rapidly mapping the normal chromosomal localization of the DNA from which the amplified sequences were derived. In addition, a weaker staining of chromosomes 10 and X was consistently observed indicating that these chromosomes were present in only one copy in the GBM genome. This rapid approach can be used to analyze cases where no metaphase spreads from the tumor material are available. It does not require any preknowledge of amplified sequences and can be applied to screen large numbers of tumors

Idiopathic Colonic Varices: A Case Report and Review of the Literature

Colonic varices appreciated on colonoscopy are extremely rare and typically indicative of portal hypertension or chronic hepatopathology. Even more rare are those cases with no underlying pathology, idiopathic colonic varices. Here we report a case of these unexplained varices

Mapping of Multiple DNA Gains and Losses in Primary Small Cell Lung Carcinomas by Comparative Genomic Hybridization

Comparative genomic hybridization was applied for a comprehensive screening of under- and overrepresentation of genetic material in 13 autoptic small cell lung cancer specimens. The most abundant genetic changes include DNA losses of chromosome arms 3p, 5q, 10q, 13q, and 17p and DNA gains of 3q, 5p, 8q, and 17q. Amplification sites in these tumors were mapped to 22 chromosome bands. The most frequently involved band was 19q13.1 (4 cases). Bands 1p32, 2p23, 7q11.2, 8q24, and 13q33–34 were involved in two cases each

Chromosomal Gains and Losses in Uveal Melanomas Detected by Comparative Genomic Hybridization

Eleven uveal melanomas were analyzed using comparative genomic hybridization (CGH). The most abundant genetic changes were loss of chromosome 3, overrepresentation of 6p, loss of 6q, and multiplication of 8q. The smallest overrepresented regions on 6p and 8q were 6pterp21 and 8q24qter, respectively. Several additional gains and losses of chromosome segments were repeatedly observed, the most frequent one being loss of 9p (three cases). Monosomy 3 appeared to be a marker for ciliary body involvement. CGH data were compared with the results of chromosome banding. Some alterations, e.g., gains of 6p and losses of 6q, were observed with higher frequencies after CGH, while others, e.g., 9p deletions, were detected only by CGH. The data suggest some similarities of cytogenetic alterations between cutaneous and uveal melanoma. In particular, the 9p deletions are of interest due to recent reports about the location of a putative tumor-suppressor gene for cutaneous malignant melanoma in this region

Molecular cytogenetic analysis of formalin-fixed, paraffin-embedded solid tumors by comparative genomic hybridization after universal DNA-amplification

We present a technique which allows the detection and chromosomal localization of DNA sequence copy number changes in solid tumor genomes from frozen sections and paraffin embedded, formalin fixed specimens. Based on comparative genomic hybridization and on universal DNA amplification procedures this technique is possible even if only a few tumor cells are available. We demonstrate the feasibility of this method to visualize complete and partial chromosome gains and losses and gene amplifications In archived solid tumor samples