Laura J. Hardie

Reflux, Barrett's oesophagus and adenocarcinoma: burning questions

The incidence of oesophageal adenocarcinoma is increasing rapidly in Western populations. Gastro-oesophageal reflux disease is a strong risk factor for both this tumour and the pre-cancerous lesion Barretts oesophagus, but the underlying disease mechanisms remain unclear. Developing a better understanding of the aetiology and pathogenesis of Barretts oesophagus, including the induction of DNA damage and genetic alterations, might provide opportunities for improved management of individuals with this disease. This could include a better rationale for screening and surveillance programmes, as well as targeted intervention strategies.

Chromoendoscopy with methylene blue and associated DNA damage in Barrett's oesophagus

Chromoendoscopy with methylene blue has been proposed to improve targeting of biopsies to specialised intestinal metaplasia and dysplasia in Barretts oesophagus. However, methylene blue can induce oxidative damage of DNA when photosensitised by white light. We show that damage to DNA is increased in Barretts mucosa after chromoendoscopy with methylene blue, an effect apparently dependent on presence of both methylene blue and endoscopic white light. Exposure of Barretts mucosa to DNA damage during endoscopy warrants caution since it could accelerate carcinogenesis. This risk needs to be carefully balanced against the possible benefit of improved early detection of preneoplastic lesions with methylene blue chromoendoscopy.

A systematic review and meta-analysis of the risk of increasing adiposity on Barrett's esophagus.

OBJECTIVES:Esophageal adenocarcinoma and its precursor lesion, Barretts esophagus, are increasing in incidence in western populations. Gastroesophageal reflux disease (GERD) and high body mass index (BMI) are known risk factors, but it is unclear whether BMI mediates its risk on Barretts esophagus independently. This systematic review and meta-analysis investigated whether increasing BMI is associated with Barretts esophagus as compared to general population and GERD controls.METHODS:Search strategies were conducted in MEDLINE (U.S. National Library of Medicine, Bethesda, MD) (1966–2005) and EMBASE (Reed Elsevier PLC, Amsterdam, The Netherlands) (1980–2005). Studies to be included were required to present “current” BMI data for consecutively recruited Barretts esophagus patients and appropriate comparison arms with a minimum number of 30 subjects in each.RESULTS:The literature search produced 5,501 hits from which 295 papers were extracted. Only 10 studies met the criteria for inclusion. The Statistics/Data Analysis (STATA) program was used to conduct random effects meta-analyses. Nine studies comparing the BMI of the Barretts esophagus and GERD groups produced a pooled odds ratio (OR) of 0.99 per kg/m2 (95% confidence interval [CI] 0.97–1.01, I2 = 52%), while the pooled estimate of three studies comparing Barretts esophagus with general population controls was 1.02 per kg/m2 (95% CI 1.01–1.04, I2 = 0%).CONCLUSIONS:Increasing adiposity is only an indirect risk factor of Barretts esophagus through the precursor lesion of GERD. Hence, BMI status has no predictive value with respect to GERD patients and their risk of progression to Barretts esophagus.

A genome-wide association study identifies new susceptibility loci for esophageal adenocarcinoma and Barrett's esophagus.

Esophageal adenocarcinoma is a cancer with rising incidence and poor survival. Most such cancers arise in a specialized intestinal metaplastic epithelium, which is diagnostic of Barretts esophagus. In a genome-wide association study, we compared esophageal adenocarcinoma cases (n = 2,390) and individuals with precancerous Barretts esophagus (n = 3,175) with 10,120 controls in 2 phases. For the combined case group, we identified three new associations. The first is at 19p13 (rs10419226: P = 3.6 × 10−10) in CRTC1 (encoding CREB-regulated transcription coactivator), whose aberrant activation has been associated with oncogenic activity. A second is at 9q22 (rs11789015: P = 1.0 × 10−9) in BARX1, which encodes a transcription factor important in esophageal specification. A third is at 3p14 (rs2687201: P = 5.5 × 10−9) near the transcription factor FOXP1, which regulates esophageal development. We also refine a previously reported association with Barretts esophagus near the putative tumor suppressor gene FOXF1 at 16q24 and extend our findings to now include esophageal adenocarcinoma.

Glutathione S-transferase M1 null genotype is associated with a decreased risk of myocardial infarction

Tobacco smoke is a major cause of both cancer and vascular disease. Although its carcinogenic role via induction of DNA damage and mutation is well established, the mechanisms involved in vascular disease remain unclear. One possibility is that DNA damage causes smooth muscle cell proliferation in the intima of arteries, thereby contributing to atherothrombotic processes. The binding of chemicals to DNA is modulated by detoxification enzymes, including glutathione S‐trans‐ferases (GST) and microsomal epoxide hydrolase (EPXH). We therefore examined whether polymorphisms in these genes influence risk of cardiovascular disease. Blood was obtained from 398 patients admitted for angiographic investigation of chest pain and 196 age‐ and sex‐matched controls. Patients were subdivided into those with and without previous acute myocardial infarction (AMI). DNA was analyzed for deletions in the GSTM1 and T1 genes and for substitutions in EPXH and GSTP1 genes. The GSTM1 null genotype occurred at a significantly lower frequency in the AMI patient group (48%) compared both to patients with no history of AMI (59%) and to the control group (57.2%). When subjects were stratified for smoking status, a significant association was observed only in smokers, suggesting the polymorphism is more important in the presence of tobacco smoke exposure. The association remained significant after adjusting for age, sex, and stenosis (presence or absence). No significant associations were observed between the other genotypes and cardiovascular disease (X2 test; P>0.1). The results of this study indicate that the GSTM1 null genotype is protective against AMI, an effect that is more marked in smokers. However, further study is required in order to elucidate the as yet unexplained, mechanisms underlying this association.—Wilson, M. H., Grant, P. J., Hardie, L. J., Wild, C. P. Glutathione S‐transferase M1 null genotype is associated with a decreased risk of myocardial infarction. FASEB J. 14, 791–796 (2000)

TP53 and progression from Barrett's metaplasia to oesophageal adenocarcinoma in a UK population cohort

Background and aims: Oesophageal adenocarcinoma frequently develops on a background of metaplastic Barrett’s epithelium. The development of malignancy is accompanied by genetic alterations, which may be promising biomarkers of disease progression. Methods: A case control study was conducted nested within a large unselected population based cohort of Barrett’s patients. Incident oesophageal malignancies and high grade dysplasias were identified. For each case up to five controls were matched on age, sex, and year of diagnosis. Biopsies from the time of diagnosis of Barrett’s epithelium were stained immunohistochemically for TP53, cyclin D1, cyclooxygenase 2 (COX-2), and β-catenin proteins. Results: Twenty nine incident oesophageal malignancies and six cases of high grade dysplasia were identified. The odds of diffuse or intense TP53 staining were substantially elevated in biopsies from patients who developed oesophageal adenocarcinoma compared with controls (odds ratio (OR) 11.7 (95% confidence interval (CI) 1.93, 71.4)). This difference was also present when all cases were considered (OR 8.42 (95% CI 2.37, 30.0). Despite the association with TP53 staining, only 32.4% of cases had an initial biopsy showing diffuse/intense TP53 staining. There were no significant associations between cyclin D1, COX-2, or β-catenin staining and case control status. The OR for positive staining for both TP53 and COX-2 was markedly increased in cases compared with controls (OR 27.3 (95% CI 2.89, 257.0)) although only 15% of cases had positive staining for both markers. Conclusions: Immunohistochemical detection of TP53 expression is a biomarker of malignant progression in Barrett’s oesophagus but sensitivity is too low to act as a criterion to inform endoscopic surveillance strategies. Additional biomarkers are required which when combined with TP53 will identify, with adequate sensitivity and specificity, Barrett’s patients who are at risk of developing cancer.

Risk Factors, DNA Damage, and Disease Progression in Barrett's Esophagus

Esophageal adenocarcinoma develops on a background of Barretts esophagus. A number of risk factors have been linked to both conditions, including gastroesophageal reflux and smoking. However, the molecular mechanisms by which these factors influence disease progression remain unclear. One possibility is that risk factors generate promutagenic DNA damage in the esophagus. The comet assay was used to measure DNA damage in esophageal (Barretts and squamous) and gastric mucosa of Barretts patients with (n = 24) or without (n = 50) associated adenocarcinoma or high-grade dysplasia in comparison with control patients (squamous mucosa) without Barretts esophagus (n = 64). Patients completed a questionnaire detailing exposure to some of the known risk factors for Barretts esophagus and adenocarcinoma. In Barretts esophagus patients, DNA damage was higher in Barretts mucosa compared with normal esophageal and gastric mucosa (P < 0.001). In addition, the highest quartile of DNA damage in Barretts mucosa was associated with an increased risk (odds ratio, 9.4; 95% confidence interval, 1.1-83.4; P = 0.044) of developing adenocarcinoma or high-grade dysplasia compared with DNA damage levels in the lowest quartile. Smoking was associated with higher DNA damage in squamous epithelium in all patient groups (P < 0.01) and in Barretts mucosa (P < 0.05) in Barretts esophagus patients only. In controls only, current reflux was associated with higher DNA damage, whereas anti-inflammatory drug use resulted in lower levels. Collectively, these data imply a genotoxic insult to the premalignant Barretts mucosa that may explain the genetic instability in this tissue and the progression to adenocarcinoma. There is an indication for a role for smoking in inducing DNA damage in esophageal mucosa but an understanding of the role of reflux requires further investigation.

Inter-laboratory validation of procedures for measuring 8-oxo-7,8-dihydroguanine/8-oxo-7,8-dihydro-2 '-deoxyguanosine in DNA

The aim of ESCODD, a European Commission funded Concerted Action, is to improve the precision and accuracy of methods for measuring 8-oxo-7,8-dihydroguanine (8-oxoGua) or the nucleoside (8-oxodG). On two occasions, participating laboratories received samples of different concentrations of 8-oxodG for analysis. About half the results returned (for 8-oxodG) were within 20% of the median values. Coefficients of variation (for three identical samples) were commonly around 10%. A sample of calf thymus DNA was sent, dry, to all laboratories. Analysis of 8-oxoGua/8-oxodG in this sample was a test of hydrolysis methods. Almost half the reported results were within 20% of the median value, and half obtained a CV of less than 10%. In order to test sensitivity, as well as precision, DNA was treated with photosensitiser and light to introduce increasing amounts of 8-oxoGua and samples were sent to members. Median values calculated from all returned results were 45.6 (untreated), 53.9, 60.4 and 65.6 8-oxoGua/10 6 Gua; only seven laboratories detected the increase over the whole range, while all but one detected a dose response over two concentration intervals. Results in this trial reflect a continuing improvement in precision and accuracy. The next challenge will be the analysis of 8-oxodG in DNA isolated from cells or tissue, where the concentration is much lower than in calf thymus DNA.

Methylene blue but not indigo carmine causes DNA damage to colonocytes in vitro and in vivo at concentrations used in clinical chromoendoscopy

Identification of mucosal abnormalities is aided by the use of dyes during colonoscopy (chromoendoscopy).1 Two dyes that have found particular favour are methylene blue and indigo carmine.2,3 Methylene blue, which, unlike indigo carmine, is taken up by cells, induces cellular DNA damage in vitro via the generation of singlet oxygen when photoexcited by white light.4 In contrast, indigo carmine appears to be photostable and to possess little potential to damage genetic material in vitro.5,6 A recent clinical study has shown that the extent of DNA damage (particularly oxidative DNA damage) in human oesophageal cells is increased after methylene blue chromoendoscopy.7 Additional iatrogenic oxidative DNA damage to epithelial cells is of particular concern in such precancerous tissue because of the association between oxidative DNA damage, mutagenesis and the development of malignancy.8 We hypothesised that indigo carmine would induce less DNA damage than methylene blue both in vitro in cultured colon cells during simulated chromoendoscopy conditions and in vivo in colonic biopsy samples collected at chromoendoscopy. We used the alkaline comet assay …

Risk of mortality and cancer incidence in Barrett's esophagus.

Background: There are very few prospective follow-up studies of Barrett esophagus (BE) cohorts assessing the risk of extraesophageal cancer incidence or mortality. Such studies are necessary in order to understand the overall risks of cancer and death experienced by patients with BE. Methods: A cohort of 502 patients with BE were identified at Leeds General Infirmary, England. Mortality and cancer incidence information were provided by the Office for National Statistics. Standardized mortality ratios (SMR) and standardized incidence ratios (SIR) were calculated using indirect standardization. Results: All-cause mortality was found to be elevated in patients with BE [SMR, 1.21; 95% confidence interval (95% CI), 1.06, 1.37] and remained so after esophageal cancers were excluded (SMR, 1.16; 95% CI, 1.01-1.32). Increased mortality risks were also found for malignant neoplasms of the esophagus (SMR, 7.26; 95% CI, 3.87-12.42) and diseases of the digestive system (SMR, 2.03; 95% CI, 1.11-3.40). The remaining disease categories produced no altered risk estimates. Circulatory disease mortality was borderline statistically significant (SMR, 1.24; 95% CI, 1.00-1.52; P = 0.053) for those with a specialized intestinal metaplasia diagnosis of BE. In the cancer incidence analyses, esophageal malignancies (SIR, 8.66; 95% CI, 4.73-14.53) and esophageal adenocarcinomas (SIR, 14.29; 95% CI, 7.13-22.56) were found to be increased in BE. All remaining analyses provided unaltered risks, including that of colorectal cancer. Conclusions: This study has shown evidence of an increased risk of esophageal cancer incidence and mortality in BE. It has also shown that those who have a histologic BE diagnosis may also have an increased risk of circulatory disease mortality. (Cancer Epidemiol Biomarkers Prev 2007;16(10):2090–6)