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Nonreciprocal translocations and gene amplifications are commonly found in human tumors. Although little is known about the mechanisms leading to such aberrations, tissue culture models predict that they can arise from DNA breakage, followed by cycles of chromatid fusion, asymmetric mitotic breakage, and replication. Mice deficient in both a nonhomologous end joining (NHEJ) DNA repair protein and the p53 tumor suppressor develop lymphomas at an early age harboring amplification of an IgH/c-myc fusion. Here we report that these chromosomal rearrangements are initiated by a recombination activating gene (RAG)-induced DNA cleavage. Subsequent DNA repair events juxtaposing IgH and c-myc are mediated by a break-induced replication pathway. Cycles of breakage-fusion-bridge result in amplification of IgH/c-myc while chromosome stabilization occurs through telomere capture. Thus, mice deficient in NHEJ provide excellent models to study the etiology of unbalanced translocations and amplification events during tumorigenesis.

Comparative genomic hybridization (CGH) was used to screen 54 advanced colon carcinomas, i.e., 24 primary tumors and 30 metastases, for chromosomal alterations. Using a sensitive statistical method for the determination of DNA imbalances and histograms for analysis of the incidence of changes, we identified the DNA over-representation of chromosome 20q as the most common alteration being present in 100% of cases. High incidence deletions were observed on 18q21–18q23 (96%), 4q27–4q28 (96%), 4p14 (87%), 5q21 (81%), 1p21–1p22 (72%), 21q21 (74%), 6q16 (72%), 3p12 (66%), 8p24–8p21 (66%), 9p21 (64%), 11q22 (64%), and 14q13–14q21 (64%). Further frequent over-representation was found on 7q12–7q11.2 (75%), 16p11–16p12 (70%), 19p13 (70%), 9q34 (67%), 19q13 (67%), 13q34 (64%), 13q13 (64%), 17q21 (59%), 22q11 (61%), 8q24 (57%), and 1q21 (57%). Pronounced DNA gains and losses being defined as regions in which the ratio profiles exceeded the values of 1.5 and 0.5, respectively, frequently colocalized with peaks of incidence curve. The use of difference histograms for the comparison of tumor subgroups as well as case-by-case histogram for the analysis of 15 paired tumor samples identified several of the above alterations as relevant for tumor progression and metastasis formation. The study identified additional loci and delineates more precisely those that have been previously reported. For comparative purposes, we have made our primary data (ratio profiles, clinicopathological parameters, histograms) available at the interactive web site http://amba.charite.de/cgh, where the incidence of changes can be determined at individual loci and additional parameters can be applied for the analysis of our CGH results.

Loss of chromosome 10q is a critical step during the progression and metastasis formation of lung cancer. We recently defined 3 distinct regions of allelic imbalances and considered the DMBT1 gene at 10q25-q26 an interesting candidate for the most telomeric region. Therefore, we investigated DMBT1 in 25 cancer cell lines and 39 primary tumors of the respiratory tract. The analysis by RT-PCR and Northern blot hybridization revealed that the gene is expressed in all tumors and cell lines and diminished in the SCLC line H187, indicating that RT-PCR is critical when used as the single method for the evaluation of gene expression. No mutations were found by SSCP analysis of the cDNA and the partially known genomic sequence. Similarly, Southern blot hybridization was unable to detect homozygous deletions. Allelotyping of the markers D10S587, D10S1708 and D10S1723 located near or within the DMBT1 gene did not reach the peak incidence of the 3 minimally deleted regions that we recently defined. In summary, our data do not confirm previous findings reporting frequent loss of DMBT1 expression in lung cancer. However, they strengthen the notion that the responsible gene on chromosome 10q25-q26 mediating tumor progression and metastasis formation in respiratory tract cancer remains enigmatic. Int. J. Cancer 88:71–76, 2000. © 2000 Wiley-Liss, Inc.

Chromosomal imbalances in 113 primary head and neck squamous cell carcinomas (HNSCCs) determined by comparative genomic hybridization were correlated with patients survival using custom-made computer software which enabled the assessment of individual chromosomal loci. The Kaplan-Meier analysis revealed that overrepresentations of 2q12, 3q21-29, 6p21.1, 11q13, 14q23, 14q24, 14q31, 14q32, 15q24, 16q22, and deletions of 8p21-22 and 18q11.2 were significantly associated with both shorter disease-free interval and disease-specific survival in this tumor collective. Multivariate Cox proportional hazards regression models consistently identified the gains of 3q21-29, 11q13, and the loss of 8p21-22 as independent prognostic markers carrying a higher significance than the nodal status as the only clinicopathological parameter with statistical importance. In addition, these three markers allowed a molecular dissection of the patients with low clinical risk (pN0 and pT2 tumors). Thus, the genomic data being derived from the evaluation of primary HNSCC enabled a stratification of the patients into subgroups with different survival highlighting the necessity of a genetically based tumor classification for refining diagnosis and treatment of HNSCC patients.

Metastases account for approximately 50% of the malignant tumors in the brain. In order to identify structural alterations that are associated with tumor dissemination into the central nervous system we used Comparative Genomic Hybridization (CGH) to investigate 42 brain metastases and 3 primary tumors of 40 patients. The metastases originated from lung cancer (14 cases), melanomas (7), carcinomas of breast (5), colon (5), kidney (5), adrenal gland (1) and thyroid (1). In addition, tumors of initially unknown primaries were assessed in 3 cases. The highest incidence of DNA gains were observed for the chromosomal regions 1q23, 8q24, 17q24‐q25, 20q13 (>80% of cases) followed by the gain on 7p12 (77%). DNA losses were slightly less frequent with 4q22, 4q26, 5q21, 9p21 being affected in at least 70% of the cases followed by deletions at 17p12, 4q32‐q34, 10q21, 10q23‐q24 and 18q21‐q22 in 67.5% of cases. Two unusual narrow regional peaks were observed for the gain on 17q24‐q25 and loss on 17p12.The incidence at individual loci can be viewed at our CGH online tumor database at . The metastases of each tumor type showed a recurrent pattern of changes. In those cases with primary tumor and metastases available, the CGH pattern exhibited a high degree of conformity. In conclusion, our data suggests that specific genetic lesions are associated with tumor dissemination into the nervous system and that CGH analysis may be a useful supplementary tool for classification of metastases with unknown origin.

Lung cancer is a highly aggressive neoplasm with 85% mortality. To identify new tumor-associated genes, we compared the expression profile of a primary metastasizing adenocarcinoma with normal airway epithelial cells. Two cDNA libraries of up- and downregulated genes were generated, comprising 253 and 299 clones, respectively. The sequence analysis revealed 205 different known genes and 314 cDNA fragments of unknown functions. Northern-blot analysis of 167 clones confirmed differential expression in 58%, and indicated a similar expression pattern in additional lung-cancer cell lines for selected clones, strengthening the value of this model for the identification of new candidate genes in lung carcinogenesis. Int. J. Cancer 86:512–517, 2000. © 2000 Wiley-Liss, Inc.

The present study examined the mRNA levels of glucose transporter Glut4 and G-protein Gsα-subunit in isolated ventricular myocytes from lean and genetically obese (fa/fa) Zucker rats and streptozotocin-diabetic rats. In obese animals the amount of transcripts coding for Glut4 increased to 122±6% of lean controls, whereas the mRNA coding for Gsα-subunit decreased by 42±12%. An unaltered level of GsmRNA was observed in insulin deficient rats. When cardiomyocytes from normal rats were treated with insulin, the Glut4 transcript level increased by 48±5%, whereas the GsmRNA level decreased by 55±8%. The findings suggest that insulin may act as a potential regulator of Glut4 and GsmRNA expression in the cardiac cell.

The present study examined the mRNA levels of glucose transporter Glut4 and G‐protein Gsα‐subunit in isolated ventricular myocytes from lean and genetically obese (fa/fa) Zucker rats and streptozotocin‐diabetic rats. In obese animals the amount of transcripts coding for Glut4 increased to 122±6% of lean controls, whereas the mRNA coding for Gsα‐subunit decreased by 42±12%. An unaltered level of GsmRNA was observed in insulin deficient rats. When cardiomyocytes from normal rats were treated with insulin, the Glut4 transcript level increased by 48±5%, whereas the GsmRNA level decreased by 55±8%. The findings suggest that insulin may act as a potential regulator of Glut4 and GsmRNA expression in the cardiac cell.

We have demonstrated recently that chronic hyperoxic treatment accelerates the rate of aging of fibroblasts and the rate of telomere shortening in parallel. It was hypothesized that accelerated telomere shortening is due to preferential accumulation of oxidative damage in telomeres. To test this hypothesis, we measured the accumulation of sites sensitive to S1 nuclease treatment in telomeres, in minisatellites, and in the bulk of the genome of fibroblasts under different models of oxidative stress as well as after treatment with the alkylating agent,N-methyl-N′-nitro-N-nitrosoguanidine. A comparison with qualitative data obtained by alkaline electrophoresis reveals that the sites transferred to double-strand breaks by treatment with low concentrations of S1 nuclease are, in fact, single-stranded regions in the DNA. These regions may resemble single-stranded overhangs, gaps, or conventional single-strand breaks. The frequency of single-stranded regions is significantly higher in telomeres than in minisatellites or in the bulk of the genome under all conditions tested. Those regions induced in minisatellites or in the overall genome by a bolus dose of hydrogen peroxide are completely repaired within 24 h. On the other hand, 50 ± 12% of H2O2-induced single-stranded regions remain unrepaired for at least 19 days in telomeres of human fibroblasts, leading to a significant increase of the telomeric steady-state level of these lesions. This preferential accumulation might significantly contribute to telomere shortening.