Chromosome 4 hyperploidy represents an early genetic aberration in premalignant Barrett's oesophagus

Abstract

Background and aims: Characterisation of the underlying molecular mechanisms that promote Barrett's progression may ultimately lead to identification of potential predictive genetic markers that classify patients' malignant risk. In an attempt to understand these causative pathways, fluorescence in situ hybridisation (FISH) was used in this study to determine when specific genetic alterations arise during Barrett's associated neoplastic progression.

Methods: Endoscopic cytology brushings were obtained from 28 patients with Barrett's metaplasia, 28 with dysplasia (20 low grade dysplasia (LGD) and eight with high grade dysplasia (HGD)), and seven with adenocarcinoma, together with paired control brushings from regions of normal proximal squamous cell epithelium. The exfoliated epithelial cells were washed and deposited onto slides. Probes specific for the centromeres of chromosomes 4, 8, 20, and Y, and locus specific probes for the tumour suppressor genes p16, p53, and Rb were subsequently hybridised.

Results: Aneuploidy was found early in progression, with metaplastic tissues displaying increased copy numbers of chromosomes 4 and 8. Chromosome 4 hyperploidy was found in 89%, 90%, 88%, and 100% of metaplasias, LGD, HGD, adenocarcinomas, respectively, while chromosome 8 hyperploidy occurred in 71%, 75%, 100%, and 100% of patients with the respective staging. Loss of the p16 tumour suppressor gene also presented in metaplastic epithelium (7%) but most other genetic aberrations were only seen in HGD.

Conclusions: Genetic instability arises well before dysplasia in Barrett's oesophagus, with chromosome 4 and 8 hyperploidy representing the earliest and most common alterations identified. As these aberrations are widespread at all the premalignant stages, there may be genes on chromosomes 4 and 8 that are involved in both the initiation and progression of Barrett's oesophagus.

Figure 1

Examples of a variety of fluorescence in situ hybridisation (FISH) images obtained from Barrett’s interphase cell preparations. (A) Two of three nuclei exhibiting chromosome 4 trisomy (cells originating from a female patient). (B) Male nucleus displaying chromosome 4 hyperploidy and loss of chromosome Y. (C) Nuclei all monoallelic for p16 (red) despite retaining two copies of chromosome 9 (green). (D) Nuclei containing the normal compliment of the Rb gene (green signal) but were monoallelic for p53 (red). (E) Nucleus that was trisomic for chromosome 20 (red) and tetrasomic for chromosome 8 (green signals). (F) Chromosome 8 hyperploidy (green) and chromosome 20 monosomy (red) are exhibited in this nucleus.

Figure 2

Percentage of metaplastic, dysplastic, and oesophageal adenocarcinoma samples that had chromosome 4/8 hyperploidy, monoallelic p16, or lacked their Y chromosome. The results are derived from analysis of 28 patients with metaplasia (BM), 20 with low grade dysplasia (LGD), eight with high grade dysplasia (HGD), and seven with oesophageal adenocarcinoma (OA).