Jason M. Dunn

Population-Based Study Reveals New Risk-Stratification Biomarker Panel for Barrett's Esophagus

BACKGROUND & AIMS The risk of progression of Barretts esophagus (BE) to esophageal adenocarcinoma (EAC) is low and difficult to calculate. Accurate tools to determine risk are needed to optimize surveillance and intervention. We assessed the ability of candidate biomarkers to predict which cases of BE will progress to EAC or high-grade dysplasia and identified those that can be measured in formalin-fixed tissues. METHODS We analyzed data from a nested case-control study performed using the population-based Northern Ireland BE Register (1993-2005). Cases who progressed to EAC (n = 89) or high-grade dysplasia ≥ 6 months after diagnosis with BE were matched to controls (nonprogressors, n = 291), for age, sex, and year of BE diagnosis. Established biomarkers (abnormal DNA content, p53, and cyclin A expression) and new biomarkers (levels of sialyl Lewis(a), Lewis(x), and Aspergillus oryzae lectin [AOL] and binding of wheat germ agglutinin) were assessed in paraffin-embedded tissue samples from patients with a first diagnosis of BE. Conditional logistic regression analysis was applied to assess odds of progression for patients with dysplastic and nondysplastic BE, based on biomarker status. RESULTS Low-grade dysplasia and all biomarkers tested, other than Lewis(x), were associated with risk of EAC or high-grade dysplasia. In backward selection, a panel comprising low-grade dysplasia, abnormal DNA ploidy, and AOL most accurately identified progressors and nonprogressors. The adjusted odds ratio for progression of patients with BE with low-grade dysplasia was 3.74 (95% confidence interval, 2.43-5.79) for each additional biomarker and the risk increased by 2.99 for each additional factor (95% confidence interval, 1.72-5.20) in patients without dysplasia. CONCLUSIONS Low-grade dysplasia, abnormal DNA ploidy, and AOL can be used to identify patients with BE most likely to develop EAC or high-grade dysplasia.

Image cytometry accurately detects DNA ploidy abnormalities and predicts late relapse to high-grade dysplasia and adenocarcinoma in Barrett's oesophagus following photodynamic therapy

Background and aims:DNA ploidy abnormalities (aneuploidy/tetraploidy) measured by flow cytometry (FC) are strong predictors of future cancer development in untreated Barretts oesophagus, independent of histology grade. Image cytometric DNA analysis (ICDA) is an optical technique allowing visualisation of abnormal nuclei that may be undertaken on archival tissue. Our aim was to determine the accuracy of ICDA vs FC, and evaluate DNA ploidy as a prognostic biomarker after histologically successful treatment with photodynamic therapy (PDT).Methods:Nuclei were extracted from 40 μm sections of paraffin-embedded biopsies and processed for ICDA at UCL and FC at UW using standardised protocols. Subsequently, DNA ploidy was evaluated by ICDA on a cohort of 30 patients clear of dysplasia 1 year after aminolaevulinic acid PDT for high-grade dysplasia (HGD). The results were correlated with long-term outcome.Results:In the comparative study, 93% (41 out of 44) of cases were classified identically. Errors occurred in the near-diploid region by ICDA and the tetraploid region by FC. In the cohort study, there were 13 cases of late relapse (7 cancer, 6 HGD) and 17 patients who remained free of dysplasia after a mean follow-up of 44 months. Aneuploidy post-PDT was highly predictive for recurrent HGD or cancer with a hazard ratio of 8.2 (1.8–37.8) (log-rank P=0.001).Conclusions:ICDA is accurate for the detection of DNA ploidy abnormalities when compared with FC. After histologically successful PDT, patients with residual aneuploidy are significantly more likely to develop HGD or cancer than those who become diploid. DNA ploidy by ICDA is a valuable prognostic biomarker after ablative therapy.

Radiofrequency ablation for early oesophageal squamous neoplasia: Outcomes form United Kingdom registry

AIM To report outcomes on patients undergoing radiofrequency ablation (RFA) for early oesophageal squamous neoplasia from a National Registry. METHODS A Prospective cohort study from 8 tertiary referral centres in the United Kingdom. Patients with squamous high grade dysplasia (HGD) and early squamous cell carcinoma (ESCC) confined to the mucosa were treated. Visible lesions were removed by endoscopic mucosal resection (EMR) before RFA. Following initial RFA treatment, patients were followed up 3 monthly. Residual flat dysplasia was treated with RFA until complete reversal dysplasia (CR-D) was achieved or progression to invasive Squamous cell cancer defined as infiltration into the submucosa layer or beyond. The main outcome measures were CR-D at 12 mo from start of treatment, long term durability, progression to cancer and adverse events. RESULTS Twenty patients with squamous HGD/ESCC completed treatment protocol. Five patients (25%) had EMR before starting RFA treatment. CR-D was 50% at 12 mo with a median of 1 RFA treatment, mean 1.5 (range 1-3). Two further patients achieved CR-D with repeat RFA after this time. Eighty per cent with CR-D remain dysplasia free at latest biopsy, with median follow up 24 mo (IQR 17-54). Six of 20 patients (30%) progressed to invasive cancer at 1 year. Four patients (20%) required endoscopic dilatations for symptomatic structuring after treatment. Two of these patients have required serial dilatations thereafter for symptomatic dysphagia with a median of 4 dilatations per patient. The other 2 patients required only a single dilatation to achieve an adequate symptomatic response. One patient developed cancer during follow up after end of treatment protocol. CONCLUSION The role of RFA in these patients remains unclear. In our series 50% patients responded at 12 mo. These figures are lower than limited published data.

Elastic scattering spectroscopy for detection of cancer risk in Barrett's esophagus: experimental and clinical validation of error removal by orthogonal subtraction for increasing accuracy

Elastic scattering spectroscopy (ESS) may be used to detect high-grade dysplasia (HGD) or cancer in Barretts esophagus (BE). When spectra are measured in vivo by a hand-held optical probe, variability among replicated spectra from the same site can hinder the development of a diagnostic model for cancer risk. An experiment was carried out on excised tissue to investigate how two potential sources of this variability, pressure and angle, influence spectral variability, and the results were compared with the variations observed in spectra collected in vivo from patients with Barretts esophagus. A statistical method called error removal by orthogonal subtraction (EROS) was applied to model and remove this measurement variability, which accounted for 96.6% of the variation in the spectra, from the in vivo data. Its removal allowed the construction of a diagnostic model with specificity improved from 67% to 82% (with sensitivity fixed at 90%). The improvement was maintained in predictions on an independent in vivo data set. EROS works well as an effective pretreatment for Barretts in vivo data by identifying measurement variability and ameliorating its effect. The procedure reduces the complexity and increases the accuracy and interpretability of the model for classification and detection of cancer risk in Barretts esophagus.

Clonal Selection and Persistence in Dysplastic Barrett's Esophagus and Intramucosal Cancers After Failed Radiofrequency Ablation

OBJECTIVES:Radiofrequency ablation (RFA) is used to successfully eliminate Barretts esophagus (BE)-related dysplasia or intramucosal carcinoma and aims to cause reversion to squamous epithelium. However, in 20% of cases RFA fails to return the epithelium to squamous phenotype. Follow-up studies show a similar dysplasia recurrence rate. We hypothesize that failed RFA is due to clonally mutated epithelial populations harbored in RFA-privileged sites and that RFA can select for the mutant clonal expansion.METHODS:A longitudinal case series of 19 patients with BE and high-grade dysplasia or intramucosal carcinoma were studied. DNA was extracted from individual Barretts glands, deep esophageal glands within mucosal resections and biopsy specimens before and after RFA. Mutations were identified by targeted sequencing of genes commonly mutated in Barretts adenocarcinoma.RESULTS:Five patients demonstrated persistent post-RFA pathology with persistent mutations, sometimes detected in deep esophageal glands or neighboring squamous epithelium after several rounds of RFA preceded by mucosal resection. Recurrence of pathology in three other patients was characterized by de novo mutations.CONCLUSIONS:Protumorigenic mutations can be found in post-ablation squamous mucosa as well as in mutant deep esophageal glands; both are associated with dysplasia recurrence. Following RFA, non-dysplastic Barretts epithelium can contain mutant clones that are found in a subsequent adenocarcinoma. Ablation may also drive the clonal expansion of pre-existing clones after a “bottleneck” created by the RFA. Overall, recurrence of dysplasia post RFA reflects the multicentric origins of Barretts clones and highlights the role of clonal selection in carcinogenesis.

Comparison of nuclear texture analysis and image cytometric DNA analysis for the assessment of dysplasia in Barrett's oesophagus

Background:Dysplasia is a marker of cancer risk in Barretts oesophagus (BO), but this risk is variable and diagnosis is subject to inter-observer variability. Cancer risk in BO is increased when chromosomal instability is present. Nucleotyping (NT) is a new method that uses high-resolution digital images of nuclei to assess chromatin organisation both quantitatively and qualitatively. We aimed to evaluate NT as a marker of dysplasia in BO and compare with image cytometric DNA analysis (ICM).Methods:In all, 120 patients with BO were studied. The non-dysplastic group (n=60) had specialised intestinal metaplasia only on two consecutive endoscopies after 51 months median follow-up (IQR=25–120 months). The dysplastic group (n=60) had high-grade dysplasia or carcinoma in situ. The two groups were then randomly assigned to a training set and a blinded test set in a 1 : 1 ratio. Image cytometric DNA analysis and NT was then carried out on Feulgen-stained nuclear monolayers.Results:The best-fit model for NT gave a correct classification rate (CCR) for the training set of 83%. The test set was then analysed using the same textural features and yielded a CCR of 78%. Image cytometric DNA analysis alone yielded a CCR of 73%. The combination of ICM and NT yielded a CCR of 84%.Conclusion:Nucleotyping differentiates dysplastic and non-dysplastic BO, with a greater sensitivity than ICM. A combination score based on both techniques performed better than either test in isolation. These data demonstrate that NT/ICM on nuclear monolayers is a very promising single platform test of genomic instability, which may aid pathologists in the diagnosis of dysplasia and has potential as a biomarker in BO.

Radiofrequency ablation is effective for the treatment of high-grade dysplasia in Barrett's esophagus after failed photodynamic therapy.

Endoscopic radiofrequency ablation (RFA) is an effective treatment for high-grade dysplasia in Barretts esophagus in ablation-naïve patients, but no studies have evaluated its use in patients in whom ablative therapy has previously failed. We describe 14 patients with residual high-grade dysplasia following aminolevulinic acid or Photofrin (porfimer sodium) photodynamic therapy (PDT). An overall complete reversal of dysplasia was achieved in 86 % with a combination of RFA and rescue endoscopic mucosal resection. The median total follow-up is 19 months. The rate of strictures was 7 % (1/14) and there was a low rate of buried glands (0.5 % follow-up biopsies). These data suggest RFA is both safe and effective for eradication of high-grade dysplasia in patients in whom PDT has failed.

Comparing outcome of radiofrequency ablation in Barrett’s with high grade dysplasia and intramucosal carcinoma: a prospective multicenter UK registry

BACKGROUND AND STUDY AIM Mucosal neoplasia arising in Barretts esophagus can be successfully treated with endoscopic mucosal resection (EMR) followed by radiofrequency ablation (RFA). The aim of the study was to compare clinical outcomes of patients with high grade dysplasia (HGD) or intramucosal cancer (IMC) at baseline from the United Kingdom RFA registry. PATIENTS AND METHODS Prior to RFA, visible lesions and nodularity were removed entirely by EMR. Thereafter, patients underwent RFA every 3 months until all visible Barretts mucosa was ablated or cancer developed (end points). Biopsies were taken at 12 months or when end points were reached. RESULTS A total of 515 patients, 384 with HGD and 131 with IMC, completed treatment. Prior to RFA, EMR was performed for visible lesions more frequently in the IMC cohort than in HGD patients (77 % vs. 47 %; P < 0.0001). The 12-month complete response for dysplasia and intestinal metaplasia were almost identical in the two cohorts (HGD 88 % and 76 %, respectively; IMC 87 % and 75 %, respectively; P = 0.7). Progression to invasive cancer was not significantly different at 12 months (HGD 1.8 %, IMC 3.8 %; P = 0.19). A trend towards slightly worse medium-term durability may be emerging in IMC patients (P = 0.08). In IMC, EMR followed by RFA was definitely associated with superior durability compared with RFA alone (P = 0.01). CONCLUSION The Registry reports on endoscopic therapy for Barretts neoplasia, representing real-life outcomes. Patients with IMC were more likely to have visible lesions requiring initial EMR than those with HGD, and may carry a higher risk of cancer progression in the medium term. The data consolidate the approach to ensuring that these patients undergo thorough endoscopic work-up, including EMR prior to RFA when necessary.

53 Six Year Disease Durability Outcomes on Patients Treated With Endoscopic Therapy for Barrett's Related Neoplasia From the UK Registry

Introduction Endoscopic therapy with combined Endoscopic mucosal resection (EMR) followed by Radiofrequency ablation (RFA) is now the recommended first line treatment for patients with Barrett’s (BE) related neoplasia confined to the oesophageal mucosa. Method We examine prospective data from the United Kingdom registry of patients undergoing RFA/EMR for BE neoplasia since 2008. Before RFA, visible lesions and nodularity were entirely removed by EMR. Thereafter patients underwent RFA 3 monthly until all visible BE was ablated or cancer developed (endpoints). Biopsies were taken at 12 months or when endpoints reached. Follow up endoscopies were performed periodically in all patients to check for recurrences thereafter. All patients who had completed at least 12 months of follow up after successful treatment were included in the analysis to examine durability of disease reversal long term. Results 282 patients (81% male, mean age 70 years) have completed the 12 month treatment protocol with a minimum of 12 months follow up thereafter. At median follow up of 37 months (IQR 29–49), 93% of patients with successful disease reversal were still free of neoplasia and 88% free of intestinal metaplasia recurrence. Cancer progression at this same time was seen in 1.4% of patients. Kaplan Meier (KM) statistics demonstrated a predicted 3 year neoplasia free survival in 88% of patients. At 5 and 6 years this was 86%. Similarly KM analysis showed that at 3 years 81% of patients would be free form BE and at 5 and 6 years this figure was 73%. Conclusion We report long term outcomes of a large cohort of patients with BE neoplasia who have had successful endoscopic therapy with RFA/EMR. This approach appears to have a lasting disease free benefit in the majority of patients. Recurrences do occur in a minority of patients and highlights the need for follow up in those fit for endoscopy. All collaborators of the UK RFA registry are acknowledged for their contributions to data collection for this work. Disclosure of interest None Declared.

Tu1097 HALO Radiofrequency Ablation for High Grade Dysplasia and Early Mucosal Neoplasia Arising in Barrett's Oesophagus: Interim Results Form the UK HALO Radiofrequency Ablation Registry

Introduction Barrett9s oesophagus (BE) is the pre-cursor to oesophageal adenocarcinmoa (OAC). High grade dysplasia (HGD) and early mucosal neoplasia in BE has historically been treated with surgery. Recently there is a shift towards minimally invasive endotherapy with endoscopic mucosal resection (EMR) and Radiofrequency ablation (RFA). Methods Prospective registry from 14 UK centers to audit RFA outcomes in patients with HGD and early neoplasia in BE. Prior to RFA, any visible lesions were first removed by EMR. Patients then underwent RFA 3 monthly until all visible BE was ablated or cancer developed. Biopsies were taken at the end of this protocol. Results 216 patients have completed protocol, mean age 68.6 years (40–90), 81% male. Mean time to protocol end 11.3 months (IQR 8–14.3), median 2 ablations and mean of 2.4 (2–6) during protocol with mean 1.4 circumferential ablations and 1.2 focal ablations performed during protocol. Mean length BE segment ablated is 5.8 cm (1–20). CR-HGD was achieved in 83% patients at protocol end biopsy. CR-D was 76% and CR-BE 50% at this point. CR-D was more likely in short segment BE ( Conclusion This is the largest series to date of patients undergoing RFA from 14 UK centers. End of protocol CR-D is satisfactory at 76% and successful eradication appears to be durable. Patients with short segment BE are likely to respond better. Our data represent real life outcomes of integrating minimally invasive endotherapy into demanding endoscopy service commitments. Competing interests None declared.