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Supplementary Tables S1 & S2 from Reprimo Methylation Is a Potential Biomarker of Barrett's-Associated Esophageal Neoplastic Progression

ImportanceFor many types of epithelial malignant neoplasms that are treated with definitive radiotherapy (RT), treatment prolongation and interruptions have an adverse effect on outcomes.ObjectiveTo analyze the association between RT duration and outcomes in patients with esophageal cancer who were treated with definitive chemoradiotherapy (CRT).Design, Setting, and ParticipantsThis study was an unplanned, post hoc secondary analysis of 3 prospective, multi-institutional phase 3 randomized clinical trials (Radiation Therapy Oncology Group [RTOG] 8501, RTOG 9405, and RTOG 0436) of the National Cancer Institute–sponsored NRG Oncology (formerly the National Surgical Adjuvant Breast and Bowel Project, RTOG, and Gynecologic Oncology Group). Enrolled patients with nonmetastatic esophageal cancer underwent definitive CRT in the trials between 1986 and 2013, with follow-up occurring through 2014. Data analyses were conducted between March 2022 to February 2023.ExposuresTreatment groups in the trials used standard-dose RT (50 Gy) and concurrent chemotherapy.Main Outcomes and MeasuresThe outcomes were local-regional failure (LRF), distant failure, disease-free survival (DFS), and overall survival (OS). Multivariable models were used to examine the associations between these outcomes and both RT duration and interruptions. Radiotherapy duration was analyzed as a dichotomized variable using an X-Tile software to choose a cut point and its median value as a cut point, as well as a continuous variable.ResultsThe analysis included 509 patients (median [IQR] age, 64 [57-70] years; 418 males [82%]; and 376 White individuals [74%]). The median (IQR) follow-up was 4.01 (2.93-4.92) years for surviving patients. The median cut point of RT duration was 39 days or less in 271 patients (53%) vs more than 39 days in 238 patients (47%), and the X-Tile software cut point was 45 days or less in 446 patients (88%) vs more than 45 days in 63 patients (12%). Radiotherapy interruptions occurred in 207 patients (41%). Female (vs male) sex and other (vs White) race and ethnicity were associated with longer RT duration and RT interruptions. In the multivariable models, RT duration longer than 45 days was associated with inferior DFS (hazard ratio [HR], 1.34; 95% CI, 1.01-1.77; P = .04). The HR for OS was 1.33, but the results were not statistically significant (95% CI, 0.99-1.77; P = .05). Radiotherapy duration longer than 39 days (vs ≤39 days) was associated with a higher risk of LRF (HR, 1.32; 95% CI, 1.06-1.65; P = .01). As a continuous variable, RT duration (per 1 week increase) was associated with DFS failure (HR, 1.14; 95% CI, 1.01-1.28; P = .03). The HR for LRF 1.13, but the result was not statistically significant (95% CI, 0.99-1.28; P = .07).Conclusions and RelevanceResults of this study indicated that in patients with esophageal cancer receiving definitive CRT, prolonged RT duration was associated with inferior outcomes; female patients and those with other (vs White) race and ethnicity were more likely to have longer RT duration and experience RT interruptions. Radiotherapy interruptions should be minimized to optimize outcomes.

MicroRNA (miR)-141-3p, which functions as an oncogene in multiple malignancies, has been shown to be highly overexpressed in esophageal cancer cells in our previous work. miR-141-3p is predicted to bind the messenger RNA (mRNA) of tuberous sclerosis complex 1 (TSC1), a tumor suppressor, with high affinity. In this study, we investigated the expression and functional interaction between miR-141-3p and TSC1 in esophageal cancer cells. Experiments were conducted in four esophageal cancer lines and in tumor cells isolated from human esophageal cancer specimens by laser capture microdissection. miR-141-3p expression was measured by real time and droplet digital PCR. Biotinylated RNA pull-down and luciferase reporter assays were used to assess binding. miR-141-3p function was tested by assessing proliferation, migration, invasion, and induction of autophagy following its silencing. We found that miR-141-3p levels were increased in TE7, OE33, and TE10 esophageal cancer cells compared to FLO-1 cells, with similar heterogeneity observed in human esophageal cancer specimens. Silencing of miR-141-3p led to increased TSC1 protein expression in these cells and was associated with increased TSC1 translation. Binding studies reveal that miR-141-3p binds to each of the predicted binding sites in the 3'-untranslated region of TSC1 mRNA. Following miR-141-3p silencing, TE7, OE33, and TE10 cells exhibited decreased proliferation, migration, and invasion, as well as enhanced autophagy. Importantly, these phenotypic effects were replicated by overexpression of TSC1 alone in these cells. Our results indicate that miR-141-3p functions in an oncogenic capacity in a subset of esophageal cancer cells, in part by suppressing TSC1 expression.

BACKGROUND Over 6 million esophagogastroduodenoscopy (EGD) procedures are performed in the United States each year. Patients having anesthesia for advanced EGD procedures, such as interventional procedures, are at high risk for hypoxemia. METHODS Our primary study aim was to evaluate whether high-flow nasal cannula (HFNC) oxygen reduces the incidence of hypoxemia during anesthesia for advanced EGD. Secondarily, we studied whether HFNC oxygen reduces hypercarbia or hypotension. After obtaining written informed consent, adults having anesthesia for advanced EGD, expected to last longer than 15 minutes, were randomly assigned to receive HFNC oxygen or standard nasal cannula (SNC) oxygen. The primary outcome was occurrence of one or more hypoxemia events during anesthesia, defined by arterial oxygen saturation

Background Salmonella enterica serovar Typhi (S. Typhi) is a highly invasive bacterium that infects the human intestinal mucosa and causes ~ 11.9–20.6 million infections and ~ 130,000–223,000 deaths annually worldwide. Oral typhoid vaccine Ty21a confers a moderate level of long-lived protection (5–7 years) in the field. New and improved vaccines against enteric pathogens are needed but their development is hindered by a lack of the immunological correlates of protection especially at the site of infection. Tissue resident memory T (TRM) cells provide immediate adaptive effector immune responsiveness at the infection site. However, the mechanism(s) by which S. Typhi induces TRM in the intestinal mucosa are unknown. Here, we focus on the induction of S. Typhi-specific CD4+TRM subsets by Ty21a in the human terminal ileum lamina propria and epithelial compartments. Methods Terminal ileum biopsies were obtained from consenting volunteers undergoing routine colonoscopy who were either immunized orally with 4 doses of Ty21a or not. Isolated lamina propria mononuclear cells (LPMC) and intraepithelial lymphocytes (IEL) CD4+TRM immune responses were determined using either S. Typhi-infected or non-infected autologous EBV-B cell lines as stimulator cells. T-CMI was assessed by the production of 4 cytokines [interferon (IFN)γ, interleukin (IL)-2, IL-17A and tumor necrosis factor (TNF)α] in 36 volunteers (18 vaccinees and 18 controls volunteers). Results Although the frequencies of LPMC CD103+ CD4+TRM were significant decreased, both CD103+ and CD103− CD4+TRM subsets spontaneously produced significantly higher levels of cytokines (IFNγ and IL-17A) following Ty21a-immunization. Importantly, we observed significant increases in S. Typhi-specific LPMC CD103+ CD4+TRM (IFNγ and IL-17A) and CD103− CD4+TRM (IL-2 and IL-17A) responses following Ty21a-immunization. Further, differences in S. Typhi-specific responses between these two CD4+TRM subsets were observed following multifunctional analysis. In addition, we determined the effect of Ty21a-immunization on IEL and observed significant changes in the frequencies of IEL CD103+ (decrease) and CD103− CD4+TRM (increase) following immunization. Finally, we observed that IEL CD103− CD4+TRM, but not CD103+ CD4+TRM, produced increased cytokines (IFNγ, TNFα and IL-17A) to S. Typhi-specific stimulation following Ty21a-immunization. Conclusions Oral Ty21a-immunization elicits distinct compartment specific immune responses in CD4+TRM (CD103+ and CD103−) subsets. This study provides novel insights in the generation of local vaccine-specific responses. Trial registration This study was approved by the Institutional Review Board and registered on ClinicalTrials.gov (identifier NCT03970304, Registered 29 May 2019—Retrospectively registered, http://www.ClinicalTrials.gov/NCT03970304)

Esophageal adenocarcinoma (EAC) develops as a consequence of gastroesophageal reflux disease and Barrett’s esophagus (BE). While combination therapy with chemotherapy or concurrent chemoradiotherapy followed by esophagectomy improves survival in more advanced tumors, the optimal treatment strategy for early-stage EAC is undefined. Endoscopic eradication therapy, consisting of endoscopic resection and mucosal ablation, has revolutionized therapy for superficial (T1a) EAC in BE and allows for esophageal preservation in appropriate patients at low risk for lymph node metastasis (LNM). This review critically examines the literature regarding evaluation, treatment, and outcomes in patients with T1 EAC. The literature was queried via the PubMed database to include articles published between 1990 and 2017. Search terms were generated from the key statements “Endoscopic eradication therapy results in equivalent overall survival when compared to esophagectomy for clinical T1aN0 EAC” and “Esophagectomy provides better overall survival than endoscopic eradication therapy for cT1b EAC”. Abstracts were reviewed and included according to predefined selection and exclusion criteria, and were then assessed according to the GRADE system. In patients with T1aN0 EAC, overall survival with endoscopic eradication therapy is equal to esophagectomy. Given the substantial risk of LNM in patients with submucosal (T1b) EAC, esophagectomy remains the standard of care for surgical candidates. In the case of inoperability or low-risk lesions, endoscopic resection may be considered adequate therapy. Chemotherapy and radiation can be offered as primary therapy for non-surgical candidates with lesions not amenable to endoscopic therapy, but does not have a clear role in the adjuvant setting after either endoscopic or surgical resection.

Purpose/Objective(s) Standard of care for surgical candidates with locally advanced esophageal/gastroesophageal junction (E/GEJ) cancers is trimodality therapy. Similar to other groups, we have previously demonstrated the strong prognostic significance of pathological complete response (pCR) at the time of esophagectomy. Little is known about the prognostic significance of nodal clearance alone, however. We sought to determine the prognostic significance of nodal clearance in the subgroup of node positive E/GEJ (N+ E/GEJ), hypothesizing that nodal clearance drives the improved outcomes seen in patients with pCR as the primary tumor is more completely removed at esophagectomy. Materials/Methods Patients undergoing TMT for a primary N+ E/GEJ cancer from 1993-2020 within a single institution were retrospectively analyzed. All patients had either clinically involved nodal disease prior to nCRT and/or pathologically confirmed residual nodal disease at time of esophagectomy. Cancer and demographic features were collected. Direct comparisons were made between pCR versus no pCR and nodal clearance (ypN0) versus persistent nodal disease (ypN+) using the Kaplan-Meier method to estimate progression free survival (PFS) and overall survival (OS). Results One hundred eighty-one patients with median follow up of 140 months were included for analysis. Median age at diagnosis was 60.1 years (30.1-80.9), 142 (78.5%) were adenocarcinomas, 102 (56.4%) were AJCC stage IIIA. Fifty-one (28.2%) had pCR, 103 (56.9%) had ypN0 (including 52 [28.7%] with nodal clearance and residual tumor) and 78 (43.1%) had ypN+. Median OS and PFS for the entire cohort were 26.4 (95% CI, 20.5-32.3) and 18.0 months (95% CI, 12.7-23.4), respectively; 2-year local control rate was 87.4% (95% CI, 84.6-90.2). When comparing ypN0 to ypN+, there were statistically significant improvements in OS and PFS. In contrast, there were no significant differences in OS and PFS when comparing patients with pCR to no pCR (Table 1). Conclusion While pCR is a known prognostic factor in E/GEJ regardless of nodal status, in the high-risk subset of N+ E/GEJ patients, nodal clearance was associated with significantly improved OS and PFS regardless of primary tumor response. This is a novel observation and warrants similar hypothesis testing in other data sets and may have implications for therapy in node positive patients.

Mucosal CD4+ T memory responses after typhoid immunization, Our current understanding of CD4+ T-cell-mediated immunity (CMI) elicited by the oral live attenuated typhoid vaccine Ty21a is primarily derived from studies using peripheral blood. Very limited data are available in humans regarding mucosal immunity (especially CD4+ T) at the site of infection (e.g. terminal ileum; TI). Here using multiparametric flow cytometry, we examined the effect of Ty21a immunization on TI-lamina propria mononuclear cells (LPMC) and peripheral blood CD4+ T memory (TM) subsets in volunteers undergoing routine colonoscopy. Interestingly, we observed significant increases in the frequencies of LPMC CD4+ T cells following Ty21a immunization, restricted to the T effector/memory (TEM)-CD45RA+ (TEMRA) subset. Importantly, Ty21a immunization elicited Salmonella Typhi-responsive LPMC CD4+ T cells in all major TM subsets [interferon (IFN)γ and interleukin (IL)-17A in TEM; IFNγ and macrophage inflammatory protein (MIP)1β in T central/memory (TCM); and IL-2 in TEMRA]. Subsequently, we analyzed LPMC S. Typhi-responsive CD4+ T cells in depth for multifunctional (MF) effectors. We found that LPMC CD4+ TEM responses were mostly MF, except for those cells exhibiting the characteristics associated with IL-17A responses. Finally, we compared mucosal to systemic responses and observed that LPMC CD4+S. Typhi-specific responses were unique and distinct from their systemic counterparts. This study provides the first demonstration of S. Typhi-specific CD4+ TM responses in the human TI mucosa and provides valuable information about the generation of mucosal immune responses following oral Ty21a immunization.

Re-irradiation (re-RT) for locoregionally recurrent esophageal and gastroesophageal junction (GEJ) cancer and de novo esophageal + GEJ cancer arising in-field after a course of prior radiation poses considerable treatment challenges given the sensitivity of surrounding organs at risk (OARs). Guidelines for treatment of this presentation are not well established. Pencil-beam scanning (PBS) proton therapy has the ability to decrease radiation dose to OARs relative to photon plans. We present the first published series to date of re-RT with PBS for esophageal + GEJ malignancies and hypothesize that re-RT with proton PBS will be feasible and improve the safety profile of re-RT for this cohort of patients.Consecutive esophageal + GEJ cancers treated with PBS re-RT within a single institution were analyzed. Comparative volumetric-modulated arc therapy photon plans were generated. A total of 17 patients were included for analysis.At a median follow-up of 11.6 months, 1-year local control was 75.3% and overall survival was 68.9%. There were five (27.8%) grade 3 or higher late toxicities. When matched for clinical target volume coverage, proton PBS plans delivered significantly lower doses to the spinal cord, lungs, liver, and heart (all p0.05); five volumetric-modulated arc therapy plans would have been undeliverable on the basis of physician-specified OAR constraints.Re-RT for de novo or recurrent malignancies of the esophagus + GEJ, when delivered with PBS proton therapy, yields high rates of local control with acceptable acute and late toxicities in a high-risk population and decreased radiation dose to OARs relative to comparative photon plans. This is the largest series of proton re-RT for esophageal malignancies and the first that exclusively used PBS.

Background Salmonella enterica serovar Typhi (S. Typhi) is a highly invasive bacterium that infects the human intestinal mucosa and causes ~ 11.9–20.6 million infections and ~ 130,000–223,000 deaths annually worldwide. Oral typhoid vaccine Ty21a confers a moderate level of long-lived protection (5–7 years) in the field. New and improved vaccines against enteric pathogens are needed but their development is hindered by a lack of the immunological correlates of protection especially at the site of infection. Tissue resident memory T (TRM) cells provide immediate adaptive effector immune responsiveness at the infection site. However, the mechanism(s) by which S. Typhi induces TRM in the intestinal mucosa are unknown. Here, we focus on the induction of S. Typhi-specific CD4+TRM subsets by Ty21a in the human terminal ileum lamina propria and epithelial compartments. Methods Terminal ileum biopsies were obtained from consenting volunteers undergoing routine colonoscopy who were either immunized orally with 4 doses of Ty21a or not. Isolated lamina propria mononuclear cells (LPMC) and intraepithelial lymphocytes (IEL) CD4+TRM immune responses were determined using either S. Typhi-infected or non-infected autologous EBV-B cell lines as stimulator cells. T-CMI was assessed by the production of 4 cytokines [interferon (IFN)γ, interleukin (IL)-2, IL-17A and tumor necrosis factor (TNF)α] in 36 volunteers (18 vaccinees and 18 controls volunteers). Results Although the frequencies of LPMC CD103+ CD4+TRM were significant decreased, both CD103+ and CD103− CD4+TRM subsets spontaneously produced significantly higher levels of cytokines (IFNγ and IL-17A) following Ty21a-immunization. Importantly, we observed significant increases in S. Typhi-specific LPMC CD103+ CD4+TRM (IFNγ and IL-17A) and CD103− CD4+TRM (IL-2 and IL-17A) responses following Ty21a-immunization. Further, differences in S. Typhi-specific responses between these two CD4+TRM subsets were observed following multifunctional analysis. In addition, we determined the effect of Ty21a-immunization on IEL and observed significant changes in the frequencies of IEL CD103+ (decrease) and CD103− CD4+TRM (increase) following immunization. Finally, we observed that IEL CD103− CD4+TRM, but not CD103+ CD4+TRM, produced increased cytokines (IFNγ, TNFα and IL-17A) to S. Typhi-specific stimulation following Ty21a-immunization. Conclusions Oral Ty21a-immunization elicits distinct compartment specific immune responses in CD4+TRM (CD103+ and CD103−) subsets. This study provides novel insights in the generation of local vaccine-specific responses. Trial registration This study was approved by the Institutional Review Board and registered on ClinicalTrials.gov (identifier NCT03970304, Registered 29 May 2019—Retrospectively registered, http://www.ClinicalTrials.gov/NCT03970304)

Background and Aims Liquid nitrogen spray cryotherapy (LNSCT) has been shown to be a safe, well-tolerated, and effective therapy for Barrett's esophagus (BE)–associated high-grade dysplasia (BE-HGD) and intramucosal adenocarcinoma (IMC). Long-term follow-up is lacking. Aims The aim of this study was to assess the efficacy, durability, and rate of neoplastic progression after LNSCT in BE-HGD/IMC at 3 and 5 years. Methods In this single-center, retrospective study drawn from a prospective database, patients with BE-HGD/IMC of any length treated with LNSCT were followed with surveillance endoscopy with biopsy for 3 to 5 years. Patients with IMC completely removed by endoscopic resection were included. Outcome measures included complete eradication of HGD (CE-HGD), dysplasia, and intestinal metaplasia; incidence rates; durability of response; location of recurrent intestinal metaplasia and dysplasia; and rate of disease progression. Results A total of 50 and 40 patients were included in 3-year and 5-year analyses. Initial CE-HGD, dysplasia, and intestinal metaplasia achieved in 98%, 90%, and 60%, respectively. Overall CE-HGD, dysplasia, and intestinal metaplasia at 3 years were 96% (48/50), 94% (47/50), and 82% (41/50), and at 5 years were 93% (37/40), 88% (35/40), and 75% (30/40). Incidence rates of recurrent intestinal metaplasia, dysplasia, and HGD/esophageal adenocarcinoma per person-year of follow-up after initial complete eradication of intestinal metaplasia (CE-IM) were 12.2%, 4.0%, and 1.4% per person-year for the 5-year cohort. Most recurrences were found immediately below the neosquamocolumnar junction. Two of 7 HGD recurrences occurred later than 4 years after initial eradication, and 2 patients (4%) progressed to adenocarcinoma despite treatment. Conclusions In patients with BE-HGD/IMC, LNSCT is effective in eliminating dysplasia and intestinal metaplasia. Progression to adenocarcinoma was uncommon, and recurrence of dysplasia was successfully treated in most cases. Long-term surveillance is necessary to detect late recurrence of dysplasia.

Background & Aims Systemic cellular immunity elicited by the Ty21a oral typhoid vaccine has been extensively characterized. However, very limited data are available in humans regarding mucosal immunity at the site of infection (terminal ileum [TI]). Here we investigated the host immunity elicited by Ty21a immunization on terminal ileum–lamina propria mononuclear cells (LPMC) and peripheral blood in volunteers undergoing routine colonoscopy. Methods We characterized LPMC-T memory (TM) subsets and assessed Salmonella enterica serovar Typhi (S Typhi)–specific responses by multichromatic flow cytometry. Results No differences were observed in cell yields and phenotypes in LPMC CD8+-TM subsets following Ty21a immunization. However, Ty21a immunization elicited LPMC CD8+ T cells exhibiting significant S Typhi–specific responses (interferon-γ, tumor necrosis factor-α, interleukin-17A, and/or CD107a) in all major TM subsets (T-effector/memory [TEM], T-central/memory, and TEM-CD45RA+), although each TM subset exhibited unique characteristics. We also investigated whether Ty21a immunization elicited S Typhi–specific multifunctional effectors in LPMC CD8+ TEM. We observed that LPMC CD8+ TEM responses were mostly multifunctional, except for those cells exhibiting the characteristics associated with cytotoxic responses. Finally, we compared mucosal with systemic responses and made the important observation that LPMC CD8+S Typhi–specific responses were unique and distinct from their systemic counterparts. Conclusions This study provides the first demonstration of S Typhi–specific responses in the human terminal ileum mucosa and provides novel insights into the generation of mucosal immune responses following oral Ty21a immunization., Graphical abstract

CD4+ and CD8+ T subsets are essential components of the adaptive immune system which act in concert at the site of infections to effectively protect against pathogens. Very limited data is available in humans regarding the relationship between CD4+ and CD8+ S. Typhi responsive cells in the terminal ileum mucosa (TI) and peripheral blood following Ty21a oral typhoid immunization. Here, we compared TI lamina propria mononuclear cells (LPMC) and peripheral blood CD4+ and CD8+ T memory (TM) subsets responses and their relationship by Spearman’s correlation following Ty21a immunization in volunteers undergoing routine colonoscopy. We observed that Ty21a immunization (i) influences the homing and accumulation of both CD4+ and CD8+ T cells in the TI, particularly integrin α4β7+ CCR9+ CD8+ T cells, (ii) elicits significantly higher frequencies of LPMC S. Typhi-responsive CD8+ T multifunctional (CD107a, IFNγ, IL-17A and/or MIP1β) cells than their CD4+ T counterparts, and (iii) results in the correlation of LPMC CD4+ Teffector/memory (TEM) S. Typhi responses (CD107a, IFNγ, TNFα, IL-17A and/or MIP1β) to their LPMC CD8+ TEM counterparts. Moreover, we demonstrated that these positive correlations between CD4+ and CD8+ TEM occur primarily in TI LPMC but not in PBMC, suggesting important differences in responses between the mucosal and systemic compartments following oral Ty21a immunization. This study provides the first demonstration of the correlation of S. Typhi-specific CD4+ and CD8+ TM responses in the human terminal ileum mucosa and provides valuable information regarding the generation of mucosal and systemic immune responses following oral Ty21a-immunization which might impact future vaccine design and development.

Tissue-resident memory T cells (TRM) are newly defined memory T cells (TM) distinct from circulating TM subsets which have the potential to mount rapid protective immune responses at the site of infection. However, very limited information is available regarding the role and contribution of TRM in vaccine-mediated immune responses in humans at the site of infection. Here, we studied the role and contribution of tissue resident memory T cells (TRM) located in the terminal ileum (TI) (favored site of infection for S. Typhi) following oral Ty21a immunization in humans. We examined TI-lamina propria mononuclear cells (LPMC) and intra-epithelial lymphocytes (IEL) CD8+ TRM subsets obtained from healthy volunteers undergoing medically-indicated colonoscopies who were either immunized with Ty21a or unvaccinated. No significant differences in the frequencies of LPMC CD8+ TRM and CD8+CD69+CD103– T cells subsets were observed following Ty21a-immunization. However, LPMC CD8+ TRM exhibited significantly higher levels of cytokines (IFN-γ, IL-17A, and TNF-α) ex-vivo in Ty21a-vaccinated than in unvaccinated volunteers. LPMC CD8+ TRM S. Typhi-specific responses were evaluated using S. Typhi-infected targets and found to produce significantly higher levels of S. Typhi-specific IL-17A. In contrast, LPMC CD8+CD69+CD103- T cells produced significantly increased S. Typhi-specific levels of IFN-γ, IL-2, and IL-17A. Finally, we assessed CD8+ TRM in IEL and observed that the frequency of IEL CD8+ TRM is significantly lower following Ty21a immunization. However, ex-vivo IEL CD8+ TRM elicited by Ty21a immunization spontaneously produced significantly higher levels of cytokines (IFN-γ, IL-17A, IL-2, and TNF-α). This study provides the first demonstration of the effect of oral Ty21a vaccination on CD8+ TRM subsets (spontaneous and S. Typhi-specific) responses in the LPMC and IEL compartment of the human terminal ileum mucosa, contributing novel information to our understanding of the generation of mucosal immune responses following oral Ty21a-immunization.

Background: Intestinal exposure to gliadin leads to zonulin upregulation and consequent disassembly of intercellular tight junctions and increased intestinal permeability. We aimed to study response to gliadin exposure, in terms of barrier function and cytokine secretion, using intestinal biopsies obtained from four groups: celiac patients with active disease (ACD), celiac patients in remission (RCD), non-celiac patients with gluten sensitivity (GS) and non-celiac controls (NC). Methods: Ex-vivo human duodenal biopsies were mounted in microsnapwells and luminally incubated with either gliadin or media alone. Changes in transepithelial electrical resistance were monitored over 120 min. Media was subsequently collected and cytokines quantified. Results: Intestinal explants from all groups (ACD (n = 6), RCD (n = 6), GS (n = 6), and NC (n = 5)) demonstrated a greater increase in permeability when exposed to gliadin vs. media alone. The increase in permeability in the ACD group was greater than in the RCD and NC groups. There was a greater increase in permeability in the GS group compared to the RCD group. There was no difference in permeability between the ACD and GS groups, between the RCD and NC groups, or between the NC and GS groups. IL-10 was significantly greater in the media of the NC group compared to the RCD and GS groups. Conclusions: Increased intestinal permeability after gliadin exposure occurs in all individuals. Following gliadin exposure, both patients with gluten sensitivity and those with active celiac disease demonstrate a greater increase in intestinal permeability than celiacs in disease remission. A higher concentration of IL-10 was measured in the media exposed to control explants compared to celiac disease in remission or gluten sensitivity.

Background and Aims The obesity epidemic has led to increased use of Roux-en-Y gastric bypass (RYGB). These patients have an increased incidence of pancreaticobiliary diseases, yet standard ERCP is not possible because of surgically altered gastroduodenal anatomy. Laparoscopy-assisted ERCP (LA-ERCP) has been proposed as an option, but supporting data are derived from single-center small case series. Therefore, we conducted a large multicenter study to evaluate the feasibility, safety, and outcomes of LA-ERCP. Methods This is a retrospective cohort study of adult patients with RYGB who underwent LA-ERCP in 34 centers. Data on demographics, indications, procedure success, and adverse events were collected. Procedure success was defined when all the following were achieved: reaching the papilla, cannulating the desired duct, and providing endoscopic therapy as clinically indicated. Results A total of 579 patients (median age, 51; 84% women) were included. Indication for LA-ERCP was biliary in 89%, pancreatic in 8%, and both in 3%. Procedure success was achieved in 98%. Median total procedure time was 152 minutes (interquartile range [IQR], 109-210), with a median ERCP time of 40 minutes (IQR, 28-56). Median hospital stay was 2 days (IQR, 1-3). Adverse events were 18% (laparoscopy related, 10%; ERCP related, 7%; both, 1%) with the clear majority (92%) classified as mild/moderate, whereas 8% were severe and 1 death occurred. Conclusions Our large multicenter study indicates that LA-ERCP in patients with RYGB is feasible with a high procedure success rate comparable with that of standard ERCP in patients with normal anatomy. The ERCP-related adverse events rate is comparable with conventional ERCP, but the overall adverse event rate was higher because of the added laparoscopy-related events.

Background and Objective: Accessory spleen (AS) may be encountered as an intrapancreatic lesion on EUS. This can look similar to other pancreatic pathologies and may lead to unnecessary interventions. The goal of this study was to evaluate the accuracy of EUS in distinguishing intrapancreatic AS (IPAS) from other pancreatic lesions. Materials and Methods: Twelve sets of endoscopic images of the spleen and various pancreatic lesions confirmed on histology or cytology were gathered. Ten endosonographers were asked to characterize and identify the lesions. The responses were analyzed via Excel and the interobserver agreement was analyzed using Gwet's agreement coefficient statistic via Stata I/C v15. Results: In our sample, the interobserver agreement was 0.37 (−+1–1; 0–0.2 poor, 0.2–0.4 fair, 0.4–0.6 moderate, 0.6–0.8 substantial, and 0.8–1.0 almost perfect) for determining whether or not the pancreatic lesion is IPAS. The reviewers were able to correctly determine IPAS endosonographically with a sensitivity of 77%, specificity of 74%, and positive and negative predictive values of 50% and 92%, respectively. Conclusion: There is a moderate-to-substantial interobserver agreement in describing the sonographic characteristics of the pancreatic lesions, such as the shape, echogenicity compared to spleen, echotexture, and border of the lesions. However, the interobserver agreement is only fair when deciding if the pancreatic lesion is an IPAS. The similar profile of IPAS and pancreatic neuroendocrine tumor could confound the diagnosis of IPAS, thus contributing to the decreased interobserver agreement. This study demonstrates that EUS criteria alone are not accurate for IPAS diagnosis. Fine-needle aspiration (FNA) may be required for a confirmatory diagnosis.

Background Liquid nitrogen endoscopic spray cryotherapy can safely and effectively eradicate high-grade dysplasia in Barrett's esophagus (BE-HGD). Long-term data on treatment success and safety are lacking. Objective To assess the long-term safety and efficacy of spray cryotherapy in patients with BE-HGD. Design Single-center, retrospective study. Setting Tertiary-care referral center. Patients A total of 32 patients with BE-HGD of any length. Intervention Patients were treated with liquid nitrogen spray cryotherapy every 8 weeks until complete eradication of HGD (CE-HGD) and intestinal metaplasia (CE-IM) was found by endoscopic biopsy. Surveillance endoscopy with biopsies was performed for at least 2 years. Main Outcome Measurements CE-HGD, CE-IM, durability of response, disease progression, and adverse events. Results CE-HGD was 100% (32/32), and CE-IM was 84% (27/32) at 2-year follow-up. At last follow-up (range 24-57 months), CE-HGD was 31/32 (97%), and CE-IM was 26/32 (81%). Recurrent HGD was found in 6 (18%), with CE-HGD in 5 after repeat treatment. One patient progressed to adenocarcinoma, downgraded to HGD after repeat cryotherapy. BE segment length ≥3 cm was associated with a higher recurrence of IM ( P = .004; odds ratio 22.6) but not HGD. No serious adverse events occurred. Stricture was seen in 3 patients (9%), all successfully dilated. Limitations Retrospective study design, small sample size. Conclusion In patients with BE-HGD, liquid nitrogen spray cryotherapy has an acceptable safety profile and success rate for eliminating HGD and IM and is associated with a low rate of recurrence or progression to cancer with long-term follow-up.

Summary To assess the incidence of esophageal intra-epithelial eosinophilic infiltration following endoscopic ablation of Barrett's esophagus (BE), a retrospective study of consecutive cases of endoscopic ablation of BE with dysplasia or cancer using radiofrequency ablation (RFA) and spray cryotherapy at two centers in the United States was performed. Post-ablation eosinophilia was defined as ≥5 eosinophils per high power field during post-treatment surveillance. Twenty of 122 patients (16%) undergoing ablation developed esophageal eosinophilia after ablation, including 8/77 (10%) treated with RFA and 12/44 (27%) treated with cryotherapy. No patient had clinical or endoscopic findings of or risk factors for eosinophilic esophagitis. Esophageal eosinophilia persisted in 30% over a median of 20.2 months. On multivariate analysis, post-ablation eosinophilia was independently associated with increasing BE segment length (adjusted odds ratio 1.46 for every 2-cm increase, 95% confidence interval 1.24–1.71) and cryotherapy as the ablation modality (adjusted odds ratio 5.23, 95% confidence interval 1.67–16.39). Esophageal eosinophilic infiltration after endoscopic ablation with RFA and cryotherapy is common and is associated with the BE segment length and treatment modality. The clinical significance of post-ablation eosinophilia is unclear.

Complete colonoscopy for cancer screening requires cecal intubation. Failure to reach and examine the cecum may result in missed right colon pathology. We developed and validated a novel classification scheme for the endoscopic appearance of the normal appendiceal orifice (AO). We analyzed 1,456 AO images and grouped them into four categories based on distinguishing features: ‘diverticuloid’, ‘umbilicoid’, ‘crescent’ and ‘linear’. An expert panel classified the images and modified these categories, combining crescent and linear categories into ‘curvilinear’. A 100-image subset was classified twice by a validation cohort consisting of gastroenterology faculty and fellows. Inter-observer agreement among the expert panel, and intra- and inter-observer agreement among the validation cohort were analyzed using Fleiss’ kappa statistic. The distribution of AO images was 67% curvilinear, 19% umbilicoid and 10% diverticuloid; 85 images (4%) were not classifiable. There was substantial inter-observer agreement among the expert panel (kappa, 0.72). Inter-observer agreement among the validation cohort was moderate (kappa, 0.53 and 0.55 for the first and second viewing, respectively). Intra-observer kappa values among the validation cohort were 0.69 for the overall classification, 0.65 for diverticuloid, 0.70 for umbilicoid and 0.70 for curvilinear, indicating substantial agreement. This simple, validated classification scheme for the endoscopic appearance of the normal AO can be used both as a research and clinical tool to measure endoscopic quality, improve cecal examination and document successful cecal intubation.

The following on endoscopic treatments of Barrett's esophagus includes commentaries on animal experiments on cryotherapy; indications for cryotherapy, choice of dosimetry, number of sessions, and role in Barrett's esophagus and adenocarcinoma; recent technical developments of RFA technology and long-term effects; the comparative effects of diverse ablation procedures and the rate of recurrence following treatment; and the indications for treatment of dysplasia and the role of radiofrequency ablation.

Accumulating evidence highlights the promising results seen with endoscopic spray cryotherapy in the treatment of dysplasia associated with Barrett esophagus and esophageal carcinoma. Published studies show that the success of spray cryotherapy to eradicate Barrett high-grade dysplasia is comparable to that for other therapies, with a favourable safety profile and high levels of patient comfort. For patients with untreatable esophageal cancer, spray cryotherapy offers a therapeutic option with the potential for complete eradication in early-stage disease and palliation in advanced cases. The mechanism of tissue injury in cryotherapy is unique, with direct cytotoxic effects and ischemic effects from vascular injury. Increased tumor cell death through induction of apoptosis and immunologic effects require further study.

Background and Aims Because of the extremely low neoplastic progression rate in Barrett's esophagus, it is difficult to diagnose patients with concomitant adenocarcinoma early in their disease course. If biomarkers existed in normal squamous esophageal epithelium to identify patients with concomitant esophageal adenocarcinoma, potential applications would be far-reaching. The aim of the current study was to identify global gene expression patterns in normal esophageal epithelium capable of revealing simultaneous esophageal adenocarcinoma, even located remotely in the esophagus. Methods Tissues comprised normal esophageal epithelia from 9 patients with esophageal adenocarcinoma, 8 patients lacking esophageal adenocarcinoma or Barrett's, and 6 patients with Barrett's esophagus alone. cDNA microarrays were performed, and pattern recognition in each of these subgroups was achieved using shrunken nearest centroid predictors. Results Our method accurately discriminated normal esophageal epithelia of 8/8 patients without esophageal adenocarcinoma or Barrett's esophagus and of 6/6 patients with Barrett's esophagus alone from normal esophageal epithelia of 9/9 patients with Barrett's esophagus and concomitant esophageal adenocarcinoma. Moreover, we identified genes differentially expressed between the above subgroups. Thus, based on their corresponding normal esophageal epithelia alone, our method accurately diagnosed patients who had concomitant esophageal adenocarcinoma. Conclusions These global gene expression patterns, along with individual genes culled from them, represent potential biomarkers for the early diagnosis of esophageal adenocarcinoma from normal esophageal epithelia. Genes discovered in normal esophagus that are differentially expressed in patients with vs. without esophageal adenocarcinoma merit further pursuit in molecular genetic, functional, and therapeutic interventional studies.

Human papilloma virus (HPV) has been found to be a precursor and risk factor for both cervical and anal dysplasia. Cervical dysplasia, which is the precursor to carcinoma, is associated with immunosuppression from a variety of causes; reports of anal dysplasia associated with immune suppression exist as well. A recent study published in abstract form only demonstrated that women with inflammatory bowel disease (IBD) had high rates of cervical dysplasia and that those on immune suppressants had even higher rates of dysplasia. We report a case of a 50-year-old woman with refractory ulcerative colitis chronically treated with 6-mercaptopurine that developed severe squamous dysplasia of the rectum. The dysplastic mucosa was found to be positive for p16 (associated with high-risk HPV) after immunostaining. A total colectomy was performed. This case highlights the importance of immune suppression in the development of dysplasia of the anus/cervix secondary to HPV infection.

Retrospective series have shown the efficacy of endoscopic spray cryotherapy in eradicating high-grade dysplasia (HGD) in Barrett's esophagus (BE); however, prospective data are lacking, and efficacy for low-grade dysplasia (LGD) is unclear. The aim of this study was to assess the efficacy and safety of spray cryotherapy in patients with LGD or HGD. A multicenter, prospective open-label registry enrolled patients with dysplastic BE. Spray cryotherapy was performed every 2-3 months until there was no endoscopic evidence of BE and no histological evidence of dysplasia, followed by surveillance endoscopies up to 2 years. Primary outcome measures were complete eradication of dysplasia (CE-D) and complete eradication of all intestinal metaplasia (CE-IM). Ninety-six subjects with Barrett's dysplasia (67% HGD; 65% long-segment BE; mean length 4.5 cm) underwent 321 treatments (mean 3.3 per subject). Mean age was 67 years, 83% were male. Eighty patients (83%) completed treatment with follow-up endoscopy (mean duration 21 months). In patients with LGD, rate of CE-D was 91% (21/23) and rate of CE-IM was 61% (14/23). In HGD, CE-D rate was 81% (46/57) and CE-IM was 65% (37/57). In patients with short-segment BE (SSBE) with any dysplasia, CE-D was achieved in 97% (30/31) and CE-IM in 77% (24/31). There were no esophageal perforations or related deaths. One subject developed a stricture, which did not require dilation. One patient was hospitalized for bleeding in the setting of non-steroidal anti-inflammatory drug use. In the largest prospective cohort to date, data suggest endoscopic spray cryotherapy is a safe and effective modality for eradication of BE with LGD or HGD, particularly with SSBE.

Patients with Barrett's esophagus (BE) are at increased risk of developing esophageal adenocarcinoma (EAC). Clinical neoplastic progression risk factors, such as age and the length of the esophageal BE segment, have been identified. However, improved molecular biomarkers predicting increased progression risk are needed for improved risk assessment and stratification. Using real-time quantitative methylation-specific PCR, we screened 10 genes (HPP1, RUNX3, RIZ1, CRBP1, 3-OST-2, APC, TIMP3, p16, MGMT, p14) for promoter hypermethylation in 77 EAC, 93 BE, and 64 normal esophagus (NE) specimens. A subset of genes manifesting significant differences in methylation frequencies between BE and EAC was then analysed in 20 dysplastic specimens. All 10 genes except p14 were frequently methylated in EACs, with RUNX3, HPP1, CRBP1, RIZ1, and OST-2 representing novel methylation targets in EAC and/or BE. p16, RUNX3, and HPP1 displayed increasing methylation frequencies in BE vs EAC. Furthermore, these increases in methylation occurred early, at the interface between BE and low-grade dysplasia (LGD). To demonstrate the silencing effect of hypermethylation, we selected the EAC cells BIC1, in which the HPP1 promoter is natively methylated, and subjected them to 5-aza-2'-deoxycytidine (Aza-C) treatment. Real-time RT-PCR indicated increased HPP1 mRNA levels after 3 days of Aza-C treatment, as well as decreased levels of methylated HPP1 DNA. Hypermethylation of a subset of six genes (APC, TIMP3, CRBP1, p16, RUNX3, and HPP1) was then tested in a retrospective longitudinal study of 99 BE and nine LGD specimens obtained from 53 BE patients undergoing surveillance endoscopy. Only high-grade dysplasia (HGD) or EAC were defined as progression end points. Two patient groups were compared: eight progressors (P) and 45 nonprogressors (NP), using Cox proportional hazards models to determine the relative progression risks of age, BE segment length, and methylation events. Multivariate analyses revealed that only hypermethylation of p16 (odds ratio (OR) 1.74, 95% confidence interval (CI) 1.33-2.20), RUNX3 (OR 1.80, 95% CI 1.08-2.81), and HPP1 (OR 1.77, 95% CI 1.06-2.81) were independently associated with an increased risk of progression, whereas age, BE segment length, and hypermethylation of TIMP3, APC, or CRBP1 were not independent risk factors. In combined analyses, risk was detectable up to, but not earlier than, 2 years preceding neoplastic progression. Hypermethylation of p16, RUNX3, and HPP1 in BE or LGD may represent independent risk factors for the progression of BE to HGD or EAC. These findings have implications regarding risk stratification, early EAC detection, and the appropriate endoscopic surveillance interval for patients with BE.

Endoscopy plays a critical role in the management of patients with malignancies involving the gastrointestinal tract. Endoscopic ultrasound has provided essential staging information, made more complete by the ability to perform fine needle aspiration of suspicious lymph nodes. Novel endoscopic resection and ablative techniques are expanding therapeutic choices in premalignant and malignant conditions. Obstruction, virtually anywhere along the length of the gastrointestinal tract, can be relieved with new stents. All of these advances have made the therapeutic gastroenterologist a key member of the team managing patients with tumors of the gastrointestinal tract.

Background Breast cancer can metastasize to the esophagus and the mediastinum. EUS-guided FNA (EUS-FNA) is being used increasingly as a less invasive alternative to mediastinoscopy for procuring a tissue diagnosis of mediastinal disease and may be useful for the diagnosis of breast cancer metastatic to the esophagus and the mediastinum. Methods Twelve women (age range 54-82 years) with a history of breast cancer presented with dysphagia or other symptoms between 1 and 15 years after initial diagnosis and treatment. CT and endoscopy with biopsies suggested a mediastinal mass or lymphadenopathy with extrinsic esophageal compression but failed to provide a tissue diagnosis. EUS-FNA was performed for diagnosis. Results Cytologic evaluation of specimens obtained by EUS-FNA confirmed breast cancer metastases in 11 of 12 patients (91%). Recurrent disease was found in intramural masses and periesophageal lymph nodes. No complication resulted from any EUS-FNA procedure. Conclusions EUS-FNA is safe and effective for the diagnosis of breast cancer metastases to the esophagus and the mediastinum. EUS-FNA may be useful as a first-line method of evaluation when breast cancer metastasis to the esophagus and the mediastinum is suspected.

N-butyl-2-cyanoacrylate has been reported to be effective for bleeding varices but is not available in the United States. We report the initial US experience with cyanoacrylate in this prospective trial and evaluate its safety, efficacy, and relative costs.Patients with active or recent gastric variceal bleeding were eligible. Cyanoacrylate therapy was performed until variceal occlusion was achieved. Rebleeding was assessed at 72 h (acute phase), 6 wk (subacute phase), and 1 yr (chronic phase). Survival was assessed at 3 months and 1 yr. Cost analysis was performed comparing the first 17 patients to historical control patients not treated with cyanoacrylate.A total of 44 patients were enrolled, 37 with cirrhosis and seven with noncirrhotic portal hypertension (NCPH). In cirrhotic patients, rebleeding was seen in two of 37 (5%) at 72 h, one of 30 (3%) at 6 wk, and five of 28 (18%) at 1 yr. Survival without shunt at 3 months was 30 of 34 (88%) and at 1 yr was 24 of 31 (77%). In NCPH patients, rebleeding was seen in two of seven (29%) at 72 h. These patients received definitive therapy for NCPH after diagnosis. Mortality and costs were substantially higher in the non-cyanoacrylate group. The odds of death were greater by 7-fold in the non-cyanoacrylate group than within the cyanoacrylate group (95% CI = 1.18-41.36, p = 0.0318). At 3 months, there was a 3.18-fold difference (95% CI = 1.05-9.64, p = 0.0411) in accrued costs; at 1 yr, the difference was 2.55-fold (95% CI = 0.96-6.94, p = 0.0585). The cost-effective ratio was estimated as 108,237 US dollars/death averted, reflecting marked cost reduction with improved survival in the cyanoacrylate-treated group. This is believed to result largely from avoidance of shunt interventions.Cyanoacrylate treatment of gastric varices is safe, clinically effective, and cost effective.

Purpose : Metallic esophageal stents frequently present during the treatment of esophageal cancer while using either external beam radiotherapy or brachytherapy. The dosimetric effects due to these metallic stents have not been reported. This work investigates these dose effects for various stent models presented during a radiotherapy procedure. Methods and Materials : Two types of representative stent models, shell and ring stents, with various designs (e.g., composition and shell thickness or ring spacing), were studied. Three Monte Carlo code systems (EGS4/BEAM, EGSnrc/DOSRZnrc, and MCNP) were used to calculate the dose distributions for 6- and 15-MV external photon beams and for a 192Ir brachytherapy source with and without a metallic esophageal stent in place. Results : For a single external beam, a dose enhancement is generally observed in front of the stent (upstream) in the region within 4-mm distance of the stent surface. The enhancement at 0.5-mm distance from the stent surface can be as high as 20%. The dose behind the stent (downstream) is generally reduced. For a parallel-opposed pair (POP), a dose enhancement is always observed in the region within 3-mm distance of the stent surface. The enhancement at 0.5-mm distance from the stent surface can be as high as 10% for the 15-MV POP and 8% for the 6-MV POP. The dose effects depend on stent design (e.g., composition, thickness of shell stent, or ring spacing in ring stents). This dependence is reduced for a POP. In the case of the 192Ir brachytherapy source, a dose enhancement is observed in the region within 1-mm distance from the stent surface. The dose enhancement is approximately 5% at 0.5-mm distance from the stent surface. Conclusion : The dose perturbations due to the presence of a metallic esophageal stent during the treatment of esophageal cancer while using either external beam radiotherapy or brachytherapy should be recognized. These perturbations result in an overdose in esophageal mucosa. The overdose is within 5%–10% at a depth of 0.5 mm in the esophageal wall.

Precise clinical staging of esophageal cancer before treatment is important. Thoracoscopic/laparoscopic (Ts/Ls) staging has been proposed as a promising staging method. This study was conducted to evaluate the potential benefits of Ts/Ls staging over conventional noninvasive clinical staging in patients with esophageal cancer. From 1991 to 1999, 111 patients with esophageal cancer underwent Ts/Ls staging by the University of Maryland Medical System. Pretreatment staging workup included computed tomography, magnetic resonance imaging, and esophageal ultrasonography, followed by Ts/Ls surgical staging. Thoracoscopy was successfully performed in 102 patients and was aborted in 4 patients because of pleural adhesions. Laparoscopy was successfully done in 76 patients and was aborted in 1 patient because of peritoneal adhesion. Sixty-seven patients had both Ts and Ls staging, whereas 35 patients and 9 patients, respectively, had only Ts or Ls staging. Thirteen of 19 patients with clinical T4 disease were downstaged to T3 disease, and 8 patients with clinical T3 disease were upstaged to T4 by Ts/Ls staging. No clinical T1-2 disease was found to be associated with local invasion (T4) by Ts/Ls. Forty-eight and 19 patients had mediastinal and celiac lymph node metastases clinically diagnosed, respectively. Nine (18.8%) and 12 (63.2%) of them were proved by Ts and Ls, respectively. An additional 5 and 16 patients were found to have unexpected mediastinal and celiac lymph node metastases, respectively, by Ts/Ls. Biopsy specimens of pleura, lung, or liver were obtained by Ts/Ls procedures in 17 patients because of suspicious findings of routine imaging studies or unexpected findings during the staging operation. Five patients were found to have distant metastasis, and the presence of metastases in others was excluded. The correlation between Ts/Ls staging and conventional noninvasive clinical staging in the diagnosis of T4 disease, mediastinal lymph node metastasis, celiac lymph node metastasis, and M1 disease was 18.8%, 14.5%, 25.5%, and 20.0%, respectively. Ts/Ls provides more accurate information for evaluating local invasion, lymph node metastasis, and distant metastasis. The poor correlation of staging diagnosis between Ts/Ls and conventional noninvasive clinical examinations suggests that the accuracy of current noninvasive clinical staging is questionable and needs to be improved.

Background: The liver is a common site of metastases for various malignancies. EUS-guided fine needle aspiration (EUS-FNA) of liver masses has only been reported in small series from single centers. Methods: A retrospective questionnaire was sent by e-mail to 130 EUS-FNA centers around the world regarding indications, complications, and findings of EUS-FNA of the liver. Results: Twenty-one centers reported 167 cases of EUS-FNA of the liver. A complication was reported in 6 (4%) of 167 cases including the following: death in 1 patient with an occluding biliary stent and biliary sepsis, bleeding (1), fever (2), and pain (2). EUS-FNA diagnosed malignancy in 23 of 26 (89%) cases after nondiagnostic fine needle aspiration under transabdominal US guidance. EUS localized an unrecognized primary tumor in 17 of 33 (52%) cases in which CT had demonstrated only liver metastases. EUS image characteristics were not predictive of malignant versus benign lesions. Conclusion: EUS-guided FNA of the liver appears to be a safe procedure with a major complication rate of approximately 1%. EUS-FNA should be considered when a liver lesion is poorly accessible to US-, or CT-guided FNA should be considered when US- or CT-guided FNA fail to make a diagnosis, when a liver lesion(s) is detected (de novo) by EUS, and for investigation of possible upper GI primary tumors in the setting of liver metastases. (Gastrointest Endosc 2002;55:859-62.)

In order to identify and contrast global gene expression profiles defining the premalignant syndrome, Barrett's esophagus, as well as frank esophageal cancer, we utilized cDNA microarray technology in conjunction with bioinformatics tools. We hybridized microarrays, each containing 8000 cDNA clones, to RNAs extracted from 13 esophageal surgical or endoscopic biopsy specimens (seven Barrett's metaplasias and six esophageal carcinomas). Hierarchical cluster analysis was performed on these results and displayed using a color-coded graphic representation (Treeview). The esophageal samples clustered naturally into two principal groups, each possessing unique global gene expression profiles. After retrieving histologic reports for these tissues, we found that one main cluster contained all seven Barrett's samples, while the remaining principal cluster comprised the six esophageal cancers. The cancers also clustered according to histopathological subtype. Thus, squamous cell carcinomas (SCCAs) constituted one group, adenocarcinomas (ADCAs) clustered separately, and one signet-ring carcinoma was in its own cluster, distinct from the ADCA cluster. We conclude that cDNA microarrays and bioinformatics show promise in the classification of esophageal malignant and premalignant diseases, and that these methods can be applied to small biopsy samples.