Molecular damage in the bronchial epithelium of current and former smokers

Abstract

Background: Most lung cancers are attributed to smoking. These cancers have been associated with multiple genetic alterations and with the presence of preneoplastic bronchial lesions. In view of such associations, we evaluated the status of specific chromosomal loci in histologically normal and abnormal bronchial biopsy specimens from current and former smokers and specimens from nonsmokers.

Methods: Multiple biopsy specimens were obtained from 18 current smokers, 24 former smokers, and 21 nonsmokers. Polymerase chain reaction-based assays involving 15 polymorphic microsatellite DNA markers were used to examine eight chromosomal regions for genetic changes (loss of heterozygosity [LOH] and microsatellite alterations).

Results: LOH and microsatellite alterations were observed in biopsy specimens from both current and former smokers, but no statistically significant differences were observed between the two groups. Among individuals with a history of smoking, 86% demonstrated LOH in one or more biopsy specimens, and 24% showed LOH in all biopsy specimens. About half of the histologically normal specimens from smokers showed LOH, but the frequency of LOH and the severity of histologic change did not correspond until the carcinoma in situ stage. A subset of biopsy specimens from smokers that exhibited either normal or preneoplastic histology showed LOH at multiple chromosomal sites, a phenomenon frequently observed in carcinoma in situ and invasive cancer. LOH on chromosomes 3p and 9p was more frequent than LOH on chromosomes 5q, 17p (17p13; TP53 gene), and 13q (13q14; retinoblastoma gene). Microsatellite alterations were detected in 64% of the smokers. No genetic alterations were detected in nonsmokers.

Conclusions: Genetic changes similar to those found in lung cancers can be detected in the nonmalignant bronchial epithelium of current and former smokers and may persist for many years after smoking cessation.

Fig. 1

Representative autoradiographs of microsatellite DNA analyses involving biopsy specimens from four smoker subjects (a–d), showing loss of heterozygosity at chromosomal regions 3p22-24.2 (a), 3p14.2 (FHIT gene) (b), 17p (TP53 gene) (c), and 9p21 (d). S = normal stromal cells; 1 = histologic category 1 (normal bronchial epithelium); 2 = histologic category 2 (hyperplasia or simple squamous metaplasia without dysplasia); 3 = histologic category 3 (mild dysplasia); 4 = histologic category 4 (moderate or severe dysplasia); 5 = histologic category 5 (carcinoma in situ). In panel c, in the TP53 pentanucleotide repeat marker, a microsatellite alteration is evident in normal epithelium (category 1, lane 2). See text for more details.

Fig. 2

A) Loss of heterozygosity (LOH) in individual biopsy specimens according to smoking status and histologic categories. LOH is expressed in terms of the fractional regional loss biopsy (FRL-biopsy) index (i.e., the fraction of chromosome regions showing LOH in each biopsy specimen) (range, 0–1). Horizontal bars indicate the mean for each histologic category. B) The FRL-subject (i.e., fractional regional loss for all biopsy specimens from an individual subject) index distribution in current and former smoker subjects. Horizontal bars represent the mean for each group of subjects. See text for more details.

Fig. 3

A) Relationship between loss of heterozygosity (LOH) at individual chromosomal regions according to histologic categories in smokers. B) LOH at specific regions of chromosome 3p according to histologic categories in smokers. See legend to Fig. 1 for histologic category definitions.