Spectral karyotyping detects chromosome damage in bronchial cells of smokers and patients with cancer

Abstract

Rationale: Lung cancer is a multistep process that is preceded and often accompanied by molecular cytogenetic lesions in benign bronchial epithelium, the precise character, extent and timing of which are not well defined.

Objectives: In this study we comprehensively defined molecular cytogenetic changes in bronchial cells that may precede lung carcinoma using spectral karyotyping (SKY).

Methods: SKY was applied to cultured benign bronchial cells from 43 high-risk smokers without carcinoma, 14 patients with concurrent lung carcinoma, and 14 never-smoker healthy volunteers.

Measurements and main results: The proportion of cells displaying numeric or structural anomalies/total number of metaphase cells was calculated for each case and was referred to as the chromosomal abnormality index. Mean chromosomal abnormality indices were 15.8, 10.1, and 0.7% for patients with cancer, high-risk smokers, and never-smokers, respectively. Clonal abnormalities were found in 17 (40%) of the high-risk smokers without carcinoma and 7 (50%) of the patients with carcinoma, but in none of 14 (0%) never-smokers. Chromosomal gains observed by SKY were confirmed in interphase cultured cells or paraffin sections of biopsy specimens by fluorescence in situ hybridization in 11 of 13 cases for which appropriate probes were available. In 6 of 57 high-risk patients or those with carcinoma, identical clonal abnormalities were dispersed at multiple bronchial sites and were admixed with nonclonal cells.

Conclusions: Clonal and single-cell chromosomal abnormalities are frequent in benign bronchial epithelium during lung carcinogenesis, indicating that chromosomal missegregation and other chromosomal rearrangements occur before overt malignancy.

Figure 1.

Mean chromosomal abnormality index (CAI) is depicted for each subject group. Significant differences were observed between patients with carcinoma and never-smokers, and between high-risk subjects and never-smokers (analysis of variance), but the difference between high-risk smokers and patients with cancer was not significant. Error bars represent variances of groups.

Figure 2.

Graph and table indicate the proportions of subjects whose biopsies contained any clonal or nonclonal spectral karyotyping (SKY) abnormality. Abnormalities were nearly as frequent in high-risk smokers as in patients with lung cancer, but were significantly less frequent in never-smokers (chi-square, P = 0.00002).

Figure 3.

Composite figure showing spectral image (A), inverted 4′,6-diamidino-2-phenylindole dihydrochloride (DAPI) band image (B), fluorescence in situ hybridization (FISH) image of interphase cells cultured from the same biopsy on which SKY was performed (C), and FISH image of paraffin section of bronchial biopsy adjacent to the biopsy used for spectral imaging (D). Spectral and inverted DAPI images indicate trisomy 18 (arrows), which is confirmed in FISH images of cells probed for CEP18 (three red signals) and CEP 7 (two green signals) per cell.

Figure 4.

Composite metaphase figure illustrating inverted DAPI band image (A), spectral image (B) and computer-generated karyotype table (C). A three-way translocation (t 1;7;18) is evident (arrows) in which a portion of the long arm of chromosome 1 is translocated to the long arm of chromosome 7, which, in turn, translocates to chromosome 18. Finally, a short segment of chromosome 18 is translocated to chromosome 1.