Marja Steenman

Parental imprinting of human chromosome region 11p15.3-pter involved in the Beckwith-Wiedemann syndrome and various human neoplasia.

Cytogenetic and DNA analyses of patients with the Beckwith-Wiedemann syndrome (BWS) enabled us to refine the localization of the syndrome at 11p15.3-pter to two distinct regions. One chromosome region (BWSCR1) is near the insulin (INS) and insulin-like growth factor 2 (IGF2) genes. The other region (BWSCR2) is more proximal near two sequences with zinc-binding finger motifs and a number of known and putative genes. This latter region, at least, seems to be associated with the development of childhood tumors. Our results strongly support the proposed involvement of parental imprinting in the etiology of BWS since all balanced chromosomal abnormalities in these patients were maternally transmitted while the mothers were phenotypically normal. We demonstrate that such an autosomal balanced rearrangement can lead to a specific maternal hypomethylation of the INS/IGF2 genes localized distal to the breakpoint. This underlines the role of these genes in the etiology of the syndrome.

Comparative genomic hybridization analysis of Wilms tumors

In this study we have applied the technique of comparative genomic hybridization (CGH) to a large series of sporadic Wilms tumors, including six samples of the associated nephroblastomatosis. The data obtained were compared with the findings of molecular studies carried out on the same material. The aims of the study were (1) to characterize the range of genetic variation in sporadic Wilms tumor and nephroblastomatosis, (2) to determine whether changes could be found that have not been detected by commonly used techniques, and (3) to compare the sensitivity of CGH with that of conventional molecular analysis. The chromosomes that showed gains and losses by CGH were similar to those previously found in molecular and cytogenetic studies, however loss of 4q was a new event identified in 2 out of 46 tumors. We did not detect amplified genetic material. Comparison of the data from the nephroblastomatosis and tumor samples from the same patient showed that loss of 7p may be associated with malignant transformation, and that losses in 1p, 11p, 4q and gains in 1q and 12q can be early events; whilst loss in 9p and gain of 8, 10q and 18 are possible secondary changes in tumor development. The combined CGH and molecular techniques used demonstrated involvement of two specific 1p regions in the etiology of Wilms tumor.

Comparative genomic hybridization analysis of hepatoblastomas : additional evidence for a genetic link with Wilms tumor and rhabdomyosarcoma

We applied the technique of comparative genomic hybridization (CGH) to a series of 16 hepatoblastomas. Our goals were (1) to identify all quantitative chromosome abnormalities that appear in this type of tumor and (2) to compare the results with data from similar studies on other tumors associated with the Beckwith-Wiedemann syndrome (BWS). We found that the most commonly detected (> 30%) chromosome abnormalities were gains of chromosomes 1, 2, 7, 8, and 17. Losses of chromosomes were found in only a few cases. On comparing our results with those from studies on the BWS-associated tumors, Wilms tumor and rhabdomyosarcoma, it became clear that three chromosome regions, namely, 7q, 8q, and 17q, were the ones most commonly involved in all three types of tumors. These regions, therefore, may harbor genes that play a role in the etiology of BWS-associated tumors in general.

The application of microwave denaturation in comparative genomic hybridization

Comparative genomic hybridization (CGH) is a powerful tool for analyzing unbalanced chromosomal rearrangements in a variety of tissues. However, reproducibility of the technique is poor. We have developed an alternative protocol involving microwave denaturation of the metaphase chromosome preparations prior to the hybridization step. The advantage of this method for CGH is the retention of the morphology of the chromosomes and hence an improved chromosome banding pattern. Furthermore, it results in a consistently strong hybridization which is not dependent on the batch of lymphocytes used to obtain the metaphase chromosome spreads. This procedure has also proved to be applicable to nucleic acid hybridizations in general. The protocol, its application and the results of this method in CGH is discussed. Furthermore preliminary results of this method in paint and DNA probe hybridizations to chromosome spreads and to RNA in tissue sections are presented.

Delineation and physical separation of novel translocation breakpoints on chromosome 1p in two genetically closely associated childhood tumors.

Sporadic childhood tumors associated with Beckwith-Wiedemann syndrome (BWS) all show abnormalities of the same region on chromosome 11. In addition to chromosome 11, other chromosome regions are affected in some of these tumor types. In this study we analyzed the region on chromosome 1p involved in the etiology of BWS-associated tumors, Wilms tumor, rhabdomyosarcoma, and hepatoblastoma. For this purpose we determined the location of two novel translocation breakpoints in this chromosome region in cells from a Wilms tumor and cells from a rhabdomyosarcoma. We constructed a map of the region and found that both breakpoints are separated by at least 875 kb. We identified a PAC clone which crosses the rhabdomyosarcoma breakpoint and found several exons within this clone. We established that this breakpoint is located proximal to the PAX7 gene and, therefore, identified a new region involved in the etiology of rhabdomyosarcomas.

Comparative genomic hybridization analysis of wilms tumors

In this study we have applied the technique of comparative genomic hybridization (CGH) to a large series of sporadic Wilms tumors, including six samples of the associated nephroblastomatosis. The data obtained were compared with the findings of molecular studies carried out on the same material. The aims of the study were (1) to characterize the range of genetic variation in sporadic Wilms tumor and nephroblastomatosis, (2) to determine whether changes could be found that have not been detected by commonly used techniques, and (3) to compare the sensitivity of CGH with that of conventional molecular analysis. The chromosomes that showed gains and losses by CGH were similar to those previously found in molecular and cytogenetic studies, however loss of 4q was a new event identified in 2 out of 46 tumors. We did not detect amplified genetic material. Comparison of the data from the nephroblastomatosis and tumor samples from the same patient showed that loss of 7p may be associated with malignant transformation, and that losses in 1p, 11p, 4q and gains in 1q and 12q can be early events; whilst loss in 9p and gain of 8, 10q and 18 are possible secondary changes in tumor development. The combined CGH and molecular techniques used demonstrated involvement of two specific 1p regions in the etiology of Wilms tumor.