Frequent deletion of the CDKN2A locus in chordoma: analysis of chromosomal imbalances using array comparative genomic hybridisation

Abstract

The initiating somatic genetic events in chordoma development have not yet been identified. Most cytogenetically investigated chordomas have displayed near-diploid or moderately hypodiploid karyotypes, with several numerical and structural rearrangements. However, no consistent structural chromosome aberration has been reported. This is the first array-based study characterising DNA copy number changes in chordoma. Array comparative genomic hybridisation (aCGH) identified copy number alterations in all samples and imbalances affecting 5 or more out of the 21 investigated tumours were seen on all chromosomes. In general, deletions were more common than gains and no high-level amplification was found, supporting previous findings of primarily losses of large chromosomal regions as an important mechanism in chordoma development. Although small imbalances were commonly found, the vast majority of these were detected in single cases; no small deletion affecting all tumours could be discerned. However, the CDKN2A and CDKN2B loci in 9p21 were homo- or heterozygously lost in 70% of the tumours, a finding corroborated by fluorescence in situ hybridisation, suggesting that inactivation of these genes constitute an important step in chordoma development.

Figure 1

Genomic imbalances detected in individual samples. Gains (red) and losses (green) of genomic material were detected in all samples investigated by array comparative genomic hybridisation (aCGH). Each row corresponds to a separate sample and each column represents an individual chromosome. The respective relapse was investigated in four cases, and samples from the same tumour showed very similar patterns of aberrations. However, although the pattern of aberrations was almost identical, a few aberrations escaped detection in one of the samples from the same tumour. This was primarily found for low copy number changes and can most probably be explained by normal cell contamination (See online version for colour figure.).

Figure 2

Frequency of DNA copy number changes detected by array comparative genomic hybridisation (aCGH) in 21 chordomas. Copy number alterations present in five or more of the samples were identified on all chromosomes. The number of deletions was larger than the number of gains, and the size of the deleted regions was significantly larger than the gained regions. The genomic positions of the imbalances are presented in Table 2.

Figure 3

DNA copy number changes in a representative chordoma. Genomic profile of case 8 analysed using 32k array comparative genomic hybridisation (aCGH; top left). Tumour/reference log 2 ratios are displayed as the moving average of three consecutive bacterial artificial chromosome (BAC) clones, and individual chromosomes are separated by vertical bars. The profile demonstrates multiple imbalances, e.g., loss of chromosome 13 and homozygous deletion of CDKN2A (p16) on chromosome 9 (bottom left). Fluorescence in situ hybridisation (FISH) analysis of the same case displays loss of chromosome 13 (blue) and homozygous deletion of CDKN2A (p16) (top right). For comparison, a normal cell shows two chromosomes 13 and two normal chromosomes 9, with centromere of chromosome 9 (cep 9) and LSI p16 indicated in green and red, respectively (bottom right) (See online version for colour figure.).