Immune Infiltration Subtypes Characterization and Identification of Prognosis-Related lncRNAs in Adenocarcinoma of the Esophagogastric Junction.

The incidence of adenocarcinoma of the esophagogastric junction (AEG) has markedly increased worldwide. However, the precise etiology of AEG is still unclear, and the therapeutic options thus remain limited. Growing evidence has implicated long non-coding RNAs (lncRNAs) in cancer immunomodulation. This study aimed to examine the tumor immune infiltration status and assess the prognostic value of immune-related lncRNAs in AEG. Using the ESTIMATE method and single-sample GSEA, we first evaluated the infiltration level of 28 immune cell types in AEG samples obtained from the TCGA dataset (N=201). Patients were assigned into high- and low-immune infiltration subtypes based on the immune cell infiltration's enrichment score. GSEA and mutation pattern analysis revealed that these two immune infiltration subtypes had distinct phenotypes. We identified 1470 differentially expressed lncRNAs in two immune infiltration subtypes. From these differentially expressed lncRNAs, six prognosis-related lncRNAs were selected using the Cox regression analysis. Subsequently, an immune risk signature was constructed based on combining the values of the six prognosis-associated lncRNAs expression levels and multiple regression coefficients. To determine the risk model's prognostic capability, we performed a series of survival analyses with Kaplan-Meier methods, Cox proportional hazards regression models, and the area under receiver operating characteristic (ROC) curve. The results indicated that the immune-related risk signature could be an independent prognostic factor with a significant predictive value in patients with AEG. Furthermore, the immune-related risk signature can effectively predict the response to immunotherapy and chemotherapy in AEG patients. In conclusion, the proposed immune-related lncRNA prognostic signature is reliable and has high survival predictive value for patients with AEG and is a promising potential biomarker for immunotherapy.

Changes in the therapeutic landscape of oesophago-gastric cancers.

PURPOSE OF REVIEW: This article reviews recent randomised clinical trials on systemic treatment of oesophago-gastric cancers in the perioperative and metastatic setting. RECENT FINDINGS: Adding nivolumab to first-line chemotherapy improved survival in patients with metastatic gastric/gastro-oesophageal junction/oesophageal adenocarcinoma with PD-L1 combined positive score (CPS) ≥ five in a global trial and progression-free survival in metastatic gastric/gastro-oesophageal junction cancers in an Asian trial. The addition of pembrolizumab to first-line chemotherapy improved survival in metastatic oesophageal cancer patients, with the most benefit in oesophageal squamous cancer and tumours with high PD-L1 expression (CPS ≥ 10). Adjuvant nivolumab improved disease-free survival (DFS) in resectable oesophageal cancer patients with residual pathologic disease after neoadjuvant chemoradiation. In human epidermal growth factor receptor 2 (HER2)-positive oesophago-gastric adenocarcinoma, a phase II trial showed improved DFS when pertuzumab and trastuzumab were added to perioperative FLOT (5-fluorouracil/leucovorin, oxaliplatin, docetaxel). Another phase II trial showed improved response rates and survival in pretreated metastatic HER2-positive gastric and gastrooesophageal junction cancer patients who received the antibody-drug conjugate trastuzumab deruxtecan compared to physician's choice of chemotherapy. SUMMARY: Chemo-immunotherapy combinations will become the new standard of care for some patients with metastatic oesophago-gastric cancers. Adjuvant nivolumab is a new option for oesophageal cancer patients with poor response after neoadjuvant chemoradiation.

Ipilimumab or FOLFOX with Nivolumab and Trastuzumab in previously untreated HER2-positive locally advanced or metastatic EsophagoGastric Adenocarcinoma - the randomized phase 2 INTEGA trial (AIO STO 0217).

BACKGROUND: Esophagogastric adenocarcinoma (EGA) currently represents a main cause of cancer related death. Despite an intensified treatment for locally advanced or metastatic EGA with a doublet chemotherapy consisting of a platinum compound and a fluoropyrimidine in combination with trastuzumab for HER2-positive disease or in selected cases with docetaxel, survival remains poor. Recently, immune-oncology based strategies relevantly improved the treatment of different solid tumors and showed some promise in late or later stage trials in EGA. Notably, the combination of immunotherapy with trastuzumab to enhance anti-tumor immunity through activation of innate and adaptive immunity was beneficial in preclinical studies or clinical studies in breast cancer. METHODS: The INTEGA study is an open-label, randomized, multicenter, exploratory phase II trial designed to assess clinical performance, safety and tolerability of ipilimumab or 5-FU/folinic acid and oxaliplatin (FOLFOX) in combination with nivolumab and trastuzumab in patients with previously untreated HER2-positive, locally advanced or metastatic EGA. The primary objective is to determine the clinical performance of ipilimumab or FOLFOX in combination with nivolumab and trastuzumab in terms of overall survival. Secondary objectives are safety and tolerability, efficacy in terms of progression-free survival and objective response rate and blood-based signatures (e.g. immune response or suppression of anti-HER2 resistance) that may correlate with treatment response. DISCUSSION: Recent evidence from the phase II NCT02954536 study (oxaliplatin, capecitabine, trastuzumab and pembrolizumab) suggests the clinical feasibility of combining chemotherapy, trastuzumab and checkpoint inhibition in EGA. However, evidence for a chemotherapy-free regimen is also mounting in HER2-positive disease (NCT02689284) i.e. margetuximab and Pembrolizumab. Both studies excelled with high overall response rates and manageable toxicities. The INTEGA study aims to comparatively assess these results and select a promising new 1st line regimen which then needs to be confirmed in a randomized phase III trial. Further, the translational part of the study might help to better stratify patients and tailor treatment of either arm. TRIAL REGISTRATION: NCT03409848 24.01.2018.

First-line pembrolizumab and trastuzumab in HER2-positive oesophageal, gastric, or gastro-oesophageal junction cancer: an open-label, single-arm, phase 2 trial.

BACKGROUND: Addition of trastuzumab to first-line chemotherapy improves overall survival in patients with HER2-positive metastatic gastric cancer. We assessed the safety and activity of pembrolizumab in combination with trastuzumab and chemotherapy in first-line HER2-positive metastatic oesophagogastric (gastric, oesophageal, or gastroesophageal junction) cancer. METHODS: This study was an investigator-initiated, open-label, non-randomised, single-arm, single centre, phase 2 trial in patients aged 18 years or older with HER2-positive metastatic oesophagogastric cancer. Eligible patients had measurable or evaluable non-measurable disease, Eastern Cooperative Oncology Group performance status of 0, 1, or 2, and left ventricular ejection fraction of at least 53%. Patients were eligible to receive an initial induction cycle of 200 mg flat dose of intravenous pembrolizumab and 8 mg/kg loading dose of intravenous trastuzumab. For subsequent cycles, patients received 130 mg/m2 of intravenous oxaliplatin or 80 mg/m2 of cisplatin on day 1, 850 mg/m2 of oral capecitabine twice a day for 2 weeks followed by 1 week off (or intravenous 5-fluorouracil, 800 mg/m2 per day on days 1-5), and a 200 mg flat dose of intravenous pembrolizumab, and 6 mg/kg of trastuzumab, administered on day 1 of each 3-week cycle. The primary endpoint was 6-month progression-free survival, defined as the proportion of patients alive and free of progression at 6 months, assessed in patients who received at least one dose of trastuzumab and pembrolizumab. The regimen would be considered worthy of further investigation if 26 or more of 37 patients were progression-free at 6 months. This trial is registered with ClinicalTrials.gov, NCT02954536, and is ongoing, but closed to enrolment. FINDINGS: Between Nov 11, 2016, and Jan 23, 2019, 37 patients were enrolled. At the time of data cutoff on Aug 6, 2019, median follow-up among survivors was 13·0 months (IQR 11·7-23·5). The primary endpoint was achieved; 26 (70%; 95% CI 54-83) of 37 patients were progression-free at 6 months. The most common treatment-related adverse event of any grade was neuropathy, which was reported in 36 (97%) of 37 patients. The most common grade 3 or 4 adverse events were lymphocytopenia (seven [19%] patients with grade 3 and two [5%] with grade 4), grade 3 decreased electrolytes (six [16%] patients), and grade 3 anaemia (four [11%] patients). Serious adverse events occurred in two patients patients (both grade 3 nephritis leading to treatment discontinuation). Four patients discontinued pembrolizumab because of immune-related adverse events. There were no treatment-related deaths. INTERPRETATION: Pembrolizumab can be safely combined with trastuzumab and chemotherapy and has promising activity in HER2-positive metastatic oesophagogastric cancer. A randomised phase 3 clinical trial assessing the efficacy and safety of pembrolizumab versus placebo in combination with trastuzumab and chemotherapy in first-line HER2-positive metastatic oesophagogastric cancer is underway. FUNDING: Merck & Co.

Safety and efficacy of pembrolizumab in combination with S-1 plus oxaliplatin as a first-line treatment in patients with advanced gastric/gastroesophageal junction cancer: Cohort 1 data from the KEYNOTE-659 phase IIb study.

AIM: The KEYNOTE-659 study evaluated the efficacy and safety of pembrolizumab in combination with chemotherapy as the first-line treatment in Japanese patients with advanced gastric/gastroesophageal junction (G/GEJ) cancer. In this paper, we report results from cohort 1 (S-1 plus oxaliplatin [SOX] with pembrolizumab). METHODS: This was a non-randomised, multicentre, open-label phase IIb study in patients with advanced programmed death-ligand 1 (PD-L1)-positive, human epidermal growth factor receptor 2-negative G/GEJ tumours. The primary endpoint was the objective response rate (ORR) assessed by blinded independent central review (BICR). Secondary endpoints were duration of response (DOR), disease control rate (DCR), time to response (TTR), progression-free survival (PFS), overall survival (OS) and safety. Exploratory analyses were performed based on the PD-L1 combined positive score (CPS) status. RESULTS: Fifty-four patients were evaluated. The median follow-up was 10.1 months. ORR and DCR by BICR were 72.2% (95% confidence interval [CI] 58.4-83.5) and 96.3% (95% CI 87.3-99.5), respectively. Median DOR, TTR, PFS and OS were as follows: not reached, 1.5 months, 9.4 months and not reached. The ORR was 73.9% in patients with CPS ≥1 to <10 and 71.0% in those with CPS ≥10. Grade ≥3 treatment-related adverse events (TRAEs) were reported by 57.4% of patients. The most common grade ≥3 TRAEs were decreased platelet count (14.8%), decreased neutrophil count (13.0%), colitis (5.6%) and adrenal insufficiency (5.6%). CONCLUSIONS: SOX with pembrolizumab showed encouraging efficacy and a manageable safety profile for the first-line treatment of advanced G/GEJ cancer. TRIAL REGISTRATION: NCT03382600/JapicCTI-183829.

PD-1 and PD-L1 inhibitors in oesophago-gastric cancers.

Oesophago-gastric cancers (OGCs) are aggressive tumours. While better peri-operative strategies, increased number of cytotoxic agents and availability of targeted therapies have improved survival, there remains an unmet need for novel treatment approaches. Immune checkpoint inhibitors (ICIs) have marked a new era in cancer management with unprecedented results in several malignancies. Although OGC lagged behind other solid tumours, evidence has increasingly accumulated supporting the contention that modulation of the anti-cancer host immune response may be beneficial for at least some patients. Many trials have been completed in Eastern and Western countries, some of these leading to the approval of ICIs in the refractory setting, and favorable opinion from regulatory agencies is expected also in treatment-naïve, advanced OGC. Furthermore, studies are evaluating ICIs in the early stage setting and exploring the potential of combination treatments. In this article we discuss the biological bases underlying the successful development of ICIs in OGC and review the available data on PD-1 and PD-L1 monoclonal antibodies in this disease. Also, we present ongoing clinical trials of these ICIs that could shape the future treatment landscape of OGC.

Prognostic Value of the Systemic Inflammation Response Index in Patients with Adenocarcinoma of the Oesophagogastric Junction: A Propensity Score-Matched Analysis.

Systemic inflammation is closely related to the occurrence and development of tumours. Based on preoperative neutrophil, monocyte, and lymphocyte counts, a new systemic inflammation response index (SIRI) was established, and the predictive ability of the SIRI for the survival of patients with adenocarcinoma of the oesophagogastric junction (AEG) was evaluated by propensity score matching (PSM) analysis. A total of 302 AEG patients undergoing radical surgery were studied. Univariate and multivariate analyses were performed using Cox proportional hazards regression models. Time-dependent receiver operating characteristic (ROC) curves were used to compare the predictive capabilities of the SIRI. PSM was implemented to balance the baseline characteristics. The results showed that the SIRI, PLR, NLR, and MLR were associated with overall survival (OS) in AEG patients based on the Kaplan-Meier survival analysis. Multivariate analysis demonstrated that the SIRI was an independent prognostic factor. The AUC for the SIRI was significantly greater than that for the NLR, PLR, and MLR in predicting the 3- and 5-year OS of AEG patients. In PSM analysis, the SIRI remained an independent prognostic indicator of OS in AEG patients. The SIRI is a novel, simple, and inexpensive prognostic predictor for AEG. The prognostic value of the SIRI is superior to that of the PLR, NLR, and MLR. The SIRI can be used to distinguish the prognosis of AEG patients with different TNM stages and can be an important supplement to TNM staging.

Density of CD8-positive tumor-infiltrating T-lymphocytes is an independent prognostic factor in adenocarcinoma of the esophagogastric junction.

Tumor-infiltrating lymphocytes (TILs) have commonly been associated with markedly improved prognosis in a variety of human cancers, including carcinomas of the upper and lower gastrointestinal tract. Especially the presence of T-cells (cytotoxic as well as helper cells) seems to define a subgroup of patients with prolonged overall and event-free survival. The density of TILs was assessed via immunohistochemistry for CD8 and CD103 in a population of 228 adenocarcinomas of the esophagogastric junction. Density of CD8+ T-lymphocytes was inversely correlated with depth of tumor infiltration (p=0.013) while no correlation with any of the analyzed clinicopathologic factors could be established for CD103-density. High density of CD8-positive T-cells additionally showed significantly longer overall survival (OS) with a p-value of 0.024 while density of CD103+ cells was associated with prolonged tumor free survival (p-value 0.011). Independence could be demonstrated applying Cox proportional hazard analysis (Hazard Ratio 0.742; 95%-Confidence Interval 0.579-0.951; p=0.019). High density of CD8-positive T-lymphocytes identifies a patient subgroup with significantly prolonged overall survival, is correlated with tumor stage and might open up new therapeutic possibilities via immunomodulating drugs.

Checkpoint blockade in esophagogastric cancer.

There are few effective treatment options for metastatic esophagogastric adenocarcinomas after progression on second-line chemotherapy. Immune checkpoint blockade therapy is a promising treatment strategy for selected advanced esophagogastric cancer, and the PD-1 inhibitor pembrolizumab has recently been approved for metastatic or recurrent gastric or gastroesophageal junction cancer that has progressed beyond second-line systemic therapy. We review the current data supporting immune checkpoint blockade therapy in advanced esophagogastric adenocarcinoma.

Patterns of PD-L1 expression and CD8 T cell infiltration in gastric adenocarcinomas and associated immune stroma.

OBJECTIVE: Recent data supports a significant role for immune checkpoint inhibitors in the treatment of solid tumours. Here, we evaluate gastric and gastro-oesophageal junction (G/GEJ) adenocarcinomas for their expression of programmed death-ligand 1 (PD-L1), infiltration by CD8+ T cells and the relationship of both factors to patient survival. DESIGN: Thirty-four resections of primary invasive G/GEJ were stained by immunohistochemistry for PD-L1 and CD8 and by DNA in situ hybridisation for Epstein-Barr virus (EBV). CD8+ T cell densities both within tumours and at the tumour-stromal interface were analysed using whole slide digital imaging. Patient survival was evaluated according to PD-L1 status and CD8 density. RESULTS: 12% of resections showed tumour cell membranous PD-L1 expression and 44% showed expression within the immune stroma. Two cases (6%) were EBV positive, with one showing membranous PD-L1 positivity. Increasing CD8+ densities both within tumours and immune stroma was associated with increasing percentage of tumour (p=0.027) and stromal (p=0.005) PD-L1 expression. Both tumour and immune stromal PD-L1 expression and high intratumoral or stromal CD8+ T cell density (>500/mm2) were associated with worse progression-free survival (PFS) and overall survival (OS). CONCLUSIONS: PD-L1 is expressed on both tumour cells and in the immune stroma across all stages and histologies of G/GEJ. Surprisingly, we demonstrate that increasing CD8 infiltration is correlated with impaired PFS and OS. Patients with higher CD8+ T cell densities also have higher PD-L1 expression, indicating an adaptive immune resistance mechanism may be occurring. Further characterisation of the G/GEJ immune microenvironment may highlight targets for immune-based therapy.

High Density of Tumor-infiltrating B-Lymphocytes and Plasma Cells Signifies Prolonged Overall Survival in Adenocarcinoma of the Esophagogastric Junction.

BACKGROUND: Tumor-infiltrating lymphocytes (TILs) have been shown to be of prognostic significance in a variety of tumors. Not only T-cell, but also B-cell infiltration is commonly associated with improved survival. MATERIALS AND METHODS: We assessed the density of tumor-infiltrating B-cells, as well as that of plasma cells, in 210 adenocarcinomas of the esophagogastric junction through immunohistochemical analysis using antibodies against CD20 and CD138. RESULTS: No correlation between density of B-cells or plasma cells and various clinicopathologic features could be established. High density of tumor-infiltrating B-cells, as well as plasma cells, showed significantly better overall survival (OS) compared to patients with no infiltrates (p=0.047 and p=0.022, respectively). Cox proportional hazard analysis could verify B-cell infiltration as an independent prognostic factor (hazard ratio (HR)=0.683; 95% confidence interval (CI)=0.517-0.901; p=0.007). CONCLUSION: Plasma cell and B-cell infiltration correlates with prolonged OS and might identify a patient subset with favorable disease course.

Targeted Treatment of Esophagogastric Cancer.

Adenocarcinoma of the esophagogastric junction (EGJ) and stomach remains one of the most common causes of cancer-related death worldwide. Although there is increasing data on the mutational landscape of esophagogastric cancer, phase III trials often yield negative results, and there is a paucity of approved targeted agents. For the time being, the subset of patients carrying HER2-positive metastatic tumors can receive trastuzumab in addition to chemotherapy. Furthermore, ramucirumab has been found to be active both as a single agent and in combination with paclitaxel. Herein, we give an overview of currently approved targeted treatments for locally advanced/resectable as well as unresectable/metastatic EGJ/gastric adenocarcinoma, summarizing the underlying clinical studies. Moreover, further potential targets still under investigation are presented.

Evaluation of Barrett esophagus by multiphoton microscopy.

CONTEXT: Multiphoton microscopy (MPM) based on 2-photon excitation fluorescence and second-harmonic generation allows simultaneous visualization of cellular details and extracellular matrix components of fresh, unfixed, and unstained tissue. Portable multiphoton microscopes, which could be placed in endoscopy suites, and multiphoton endomicroscopes are in development, but their clinical utility is unknown. OBJECTIVE: To examine fresh, unfixed endoscopic biopsies obtained from the distal esophagus and gastroesophageal junction to (1) define the MPM characteristics of normal esophageal squamous mucosa and gastric columnar mucosa, and (2) evaluate whether diagnosis of intestinal metaplasia/Barrett esophagus (BE) could be made reliably with MPM. DESIGN: The study examined 35 untreated, fresh biopsy specimens from 25 patients who underwent routine upper endoscopy. A Zeiss LSM 710 Duo microscope (Carl Zeiss, Thornwood, New York) coupled to a Spectra-Physics (Mountain View, California) Tsunami Ti:sapphire laser was used to obtain a MPM image within 4 hours of fresh specimen collection. After obtaining MPM images, the biopsy specimens were placed in 10% buffered formalin and submitted for routine histopathologic examination. Then, the MPM images were compared with the findings in the hematoxylin-eosin-stained, formalin-fixed, paraffin-embedded sections. The MPM characteristics of the squamous, gastric-type columnar and intestinal-type columnar epithelium were analyzed. In biopsies with discrepancy between MPM imaging and hematoxylin-eosin-stained sections, the entire tissue block was serially sectioned and reevaluated. A diagnosis of BE was made when endoscopic and histologic criteria were satisfied. RESULTS: Based on effective 2-photon excitation fluorescence of cellular reduced pyridine nucleotides and flavin adenine dinucleotide and lack of 2-photon excitation fluorescence of mucin and cellular nuclei, MPM could readily identify and distinguish among squamous epithelial cells, goblet cells, gastric foveolar-type mucous cells, and parietal cells in the area of gastroesophageal junction. Based on the cell types identified, the mucosa was defined as squamous, columnar gastric type (cardia/fundic-type), and metaplastic columnar intestinal-type/BE. Various types of mucosa seen in the study of 35 biopsies included normal squamous mucosa only (n = 14; 40%), gastric cardia-type mucosa only (n = 2; 6%), gastric fundic mucosa (n = 6; 17%), and both squamous and gastric mucosa (n = 13; 37%). Intestinal metaplasia was identified by the presence of goblet cells in 10 of 25 cases (40%) leading to a diagnosis of BE on MPM imaging and only in 7 cases (28%) by histopathology. In 3 of 35 biopsies (9%), clear-cut goblet cells were seen by MPM imaging but not by histopathology, even after the entire tissue block was sectioned. Based on effective 2-photon excitation fluorescence of elastin and second-harmonic generation of collagen, connective tissue in the lamina propria and the basement membrane was also visualized with MPM. CONCLUSIONS: Multiphoton microscopy has the ability to accurately distinguish squamous epithelium and different cellular elements of the columnar mucosa obtained from biopsies around the gastroesophageal junction, including goblet cells that are important for the diagnosis of BE. Thus, use of MPM in the endoscopy suite might provide immediate microscopic images during endoscopy, improving screening and surveillance of patients with BE.

Comparison of the prognostic value of tumour- and patient-related factors in patients undergoing potentially curative resection of oesophageal cancer.

BACKGROUND: Evidence is increasing that elevated systemic inflammation is associated with poor survival in patients with oesophageal carcinoma. However, it is not yet established if any specific component of systemic inflammatory response is a better predictor of cancer survival. The aim of the present study was to compare the predictive value of selected markers of systemic inflammation in patients who undergo surgical resection of oesophageal cancer. METHODS: One hundred twelve patients who underwent potentially curative resection for oesophageal carcinoma, including type I and type II tumours of the gastro-oesophageal junction (Siewert and Stein in Dis Esophagus 9:173-182, 1996), between 1996 and 2008 were included in the study. Patients had laboratory measurement of white cells, neutrophils, lymphocytes, platelet counts, albumin, and C-reactive protein. Glasgow Prognostic Score (mGPS), neutrophil lymphocyte ratio (NLR), platelet lymphocyte ratio (PLR), and metastatic lymph node ratio (LNR) were calculated. RESULTS: On multivariate analysis, only the LNR (HR 2.87, 95% CI 1.99-4.15, p < 0.001) and the mGPS (HR 4.31, 95% CI 2.20-8.45, p < 0.001) were independently associated with cancer-specific survival in oesophageal cancer. An elevated mGPS was associated with high white cell count (p < 0.05) and poorer survival (p = 0.001). CONCLUSION: The present study indicates that the mGPS, an acute-phase protein-based prognostic score, better predicts cancer survival compared with the cellular components of systemic inflammation in patients with oesophageal carcinoma.

Reduction of interstitial cells of Cajal (ICC) associated with neuronal nitric oxide synthase (n-NOS) in patients with achalasia.

BACKGROUND: The etiology of achalasia is still unknown. The current theories of chronic inflammation leading to autoimmune response with destruction and loss of the inhibitory myenteric ganglion cells enlighten its pathogenesis in a limited way only. Interstitial cells of Cajal (ICC) have been shown to be involved in nitrergic neurotransmission of the lower esophageal sphincter (LES). AIM: To investigate the significance of ICC and neuronal nitric oxide synthase (n-NOS) in esophageal wall tissue of patients undergoing surgery for achalasia. METHODS: In 53 patients with a median age of 45 (6-78) yr undergoing surgery for achalasia, the immunoreactivity of ICC (CD117/c-kit) and n-NOS was assessed. In 42 patients, biopsies were taken from the LES high-pressure zone during Heller myotomy, whereas in 11 patients with end-stage achalasia and a decompensated megaesophagus, the complete esophagus was resected. A semiquantitative analysis was carried out and ICC and n-NOS impairments were classified into four grades. Staining intensity was correlated with preoperative clinical, radiologic, and manometric findings and with long-term postoperative Eckardt score. RESULTS: Grade III/IV ICC reduction (severe reduction to complete loss) was seen in 59.5% of all biopsy specimens of the LES high-pressure zone. Patients with grade III/IV ICC reduction had a significantly longer duration of achalasia symptoms (3 [0-43] yr) than patients with minor to marked (grade I/II) impairment (1 [0-16] yr, P= 0.028). A majority (72.5%) of tissue samples revealed severe reduction to complete loss of n-NOS immunoreactivity. The preoperative Eckardt score was statistically significantly different between patients with grade I/II and those with grade III/IV n-NOS reductions (P= 0.031). CD117 (c-kit) positivity was statistically significantly correlated with n-NOS staining intensity (correlation coefficient r= 0.781, P < 0.0001). CONCLUSION: The present results suggest that in the pathogenesis of achalasia, especially in the development of the LES high-pressure zone, depletion of ICC networks and potential changes in the electrical activity of smooth muscle cells may play a crucial role. The reduction in CD117-positive ICC in a few patients also seemed to be of relevance, even if the cells of Auerbach's plexus were unscathed. The associated reduced NOS release might underlie the profound ICC impairment and could possibly be responsible for the lack of LES relaxation, because of missing inhibitory neurotransmission. It is unclear, however, whether the ICC loss is primarily caused by the accelerated attrition of mature cells or their impaired regeneration.

Alterations in the density of interstitial cells of Cajal in achalasia.

The aim of this study was to assess the quantity of interstitial cells of Cajal (ICC) in the lower esophageal sphincter (LES) in achalasia. LES muscle was obtained from 11 achalasia and nine esophageal cancer (control) patients during surgery. Immunohistochemistry was performed and average cell counts per high-power field (HPF) were obtained. Overall, more ICC were observed in achalasia (median = 14.0 cells/HPF; range = 0-22.6 cells/HPF) as compared with controls (median = 6.2 cells/HPF; range = 1.6-10.8 cells/HPF) (P = 0.047). There were two subsets of findings within the achalasia group: 8/11(73%) had an increased quantity of ICC (median = 17.1 cells/HPF; range = 11.6-22.6; P = 0.015) as compared with controls, whereas the remaining 3/11(27%) had a paucity of ICC (median = 0 cells/HPF; range = 0-2; P = 0.02). ICC levels were positively correlated with age of the patient (P = 0.043). Our study demonstrates that subsets of abnormal ICC levels are observed in idiopathic achalasia of the esophagus.

Serum parathyroid hormone-related peptide is associated with systemic inflammation and adverse prognosis in gastroesophageal carcinoma.

BACKGROUND: Parathyroid hormone-related peptide (PTHrP) is a tumor-derived circulating factor that has been associated with hypercalcemia of malignancy. The role of PTHrP as a prognostic indicator remains unclear. Studies suggest that it may function as a growth factor; and, recently, the ability of PTHrP to induce cytokine expression has been described. PTHrP also has been proposed as a procachectic factor. In this study, the authors investigated the prognostic value of PTHrP in patients who had gastroesophageal carcinoma without hypercalcemia and determined whether PTHrP was associated with systemic inflammation and adverse nutritional status. METHODS: Patients were recruited at the time of diagnosis. Serum was collected for determination of c-terminal fragment PTHrP (cPTHrP) levels (by radioimmunoassay) and calcium levels as well as levels of serum cytokines and acute-phase proteins (with an enzyme-linked immunosorbent assay). Nutritional assessment of patients was undertaken at the same time as serum collection. Patients underwent routine staging, and survival duration was recorded. RESULTS: One hundred fifty-one patients with esophagogastric carcinoma were recruited. Six of 151 patients (4.0%) patients were hypercalcemic, and 26 patients (17.2%) had elevated serum cPTHrP levels. There was no association between the cPTHrP level and either serum calcium concentrations (P = 0.72) or adverse nutritional status. Elevated cPTHrP, however, was associated with significantly higher serum levels of soluble tumor necrosis factor receptor (P = 0.008) and with significantly lower levels of transferrin (P = 0.009) and albumin (P = 0.02). There was also a weak association with C-related protein levels (P = 0.06). Elevated cPTHrP levels also were associated with an adverse prognosis, as determined by reduced survival duration, on univariate analysis (P = 0.038), but not on multivariate analysis (P = 0.15). CONCLUSIONS: Elevated serum cPTHrP levels were present in approximately 17% of patients with gastroesophageal carcinoma in the absence of hypercalcemia and was associated with markers of systemic inflammation and with an adverse prognosis.

Lymphocyte subsets and T(h)1/T(h)2 immune responses in patients with adenocarcinoma of the oesophagus or oesophagogastric junction: relation to pTNM stage and clinical outcome.

INTRODUCTION: Recent studies have indicated that the cytokines produced by CD4(+) T helper type 1 (T(h)1) and type 2 (T(h)2) cells are critically important in antitumour immunity and perhaps clinical outcome. From this perspective, we investigated the immunocompetence of patients with previously untreated cancer of the oesophagus or oesophagogastric junction (OGJ) in relation to stage of disease and postoperative survival. METHODS: Blood samples were taken prior to surgery from 32 patients with adenocarcinoma of the oesophagus or OGJ. Ten healthy volunteers served as normal controls. T-cell and monocyte subpopulations were determined using flow cytometry. Monocyte as well as T(h)1- and T(h)2-lymphocyte cytokine levels were assessed in stimulated whole blood cultures. RESULTS: Absolute T-cell and monocyte (subset) counts as well as monocyte cytokine levels were similar among patients and controls. Production of T(h)1-type cytokines was higher in patients than in controls (IFN-gamma, p=0.01; IL-2, p=0.05), whereas T(h)2-type cytokine levels were comparable (IL-4, p=0.5; IL-13, p=0.3). T-cell CD4(+)/CD8(+) ratios decreased as pTNM stage worsened (stage I/II vs stage III/IV, p=0.009). Of all measured immunological parameters, only IL-2 production significantly affected both overall survival ( p=0.015) and disease-free survival ( p=0.0062). High IL-2 levels corresponded with a favourable prognosis. CONCLUSIONS: Patients awaiting surgery for adenocarcinoma of the oesophagus or oesophagogastric junction demonstrated a shift in the T(h)1/T(h)2 balance-in favour of T(h)1-compared with healthy volunteers. The ability of T cells to produce IL-2 was related to survival indicating a crucial role of T(h)1-type cells in antitumour immunosurveillance.

Phenotype of Barrett's esophagus and intestinal metaplasia of the distal esophagus and gastroesophageal junction: an immunohistochemical study of cytokeratins 7 and 20, Das-1 and 45 MI.

The pathogenesis of short segment Barrett's esophagus (SSBE) and intestinal metaplasia (IM) of the gastroesophageal junction (IMGEJ) are poorly understood. Also, these conditions are difficult to distinguish from one another based solely on endoscopic and pathologic criteria. Therefore, the aim of this study was to evaluate the immunophenotypic features of SSBE and IMGEJ and to compare the results with lesions of known etiologies: long segment BE (LSBE) caused by reflux disease and Helicobacter pylori-induced IM of the gastric antrum (IMGA). Routinely processed mucosal biopsy specimens from 11 patients with LSBE, 17 with SSBE, 10 with IMGEJ, 16 with IMGA, 17 with a normal nonmetaplastic GEJ, and 7 patients with a normal gastric antrum were immunohistochemically stained with monoclonal antibodies to: Das1, an antibody shown to react specifically with colonic goblet cells; 45M1, an antibody that recognizes the M1 gastric mucin antigen; and cytokeratin (CK) 7 and 20, antibodies that have previously been reported to show specific staining patterns in BE versus IMGA. Also evaluated was nonintestinalized mucinous epithelium from LSBE, SSBE, and also the normal GEJ and gastric antrum. LSBE, SSBE, and IMGEJ showed similar prevalences of Das1 (91% versus 88% versus 100%) and 45M1 reactivity (100% versus 100% versus 100%), and a similar pattern of CK7/20 reactivity (diffuse strong CK7 staining of the surface and crypt epithelium, and strong surface and superficial crypt CK20 staining) (91% versus 94% versus 90%). In contrast, although 45M1 reactivity in IMGA (93%) was similar to that of the other three groups, IMGA showed a significantly lower prevalence of Das positivity (13%, p < 0.001), and only a 14% prevalence of the CK7/20 staining pattern that was predominant in the other three groups (p < 0.001). Das1, 45M1, and CK7/20 staining were similar in nonintestinalized "cardia-type" mucinous epithelium from LSBE, SSBE, and the GEJ, but all were distinct from the normal gastric antrum. In summary, the immunophenotypic features of SSBE and IMGEJ are similar and closely resemble those seen in classic LSBE, but are distinct from IMGA. This may indicate that IM in LSBE, SSBE and at the GEJ have similar biologic properties. Based on our data, SSBE and IMGEJ cannot be distinguished on the basis of their immunophenotype.

Localization of galanin immunoreactivity in the opossum esophagus.

Galanin-like immunoreactivity was studied at 7 levels of the opossum esophagus, lower esophageal sphincter (LES) and adjacent portion of the stomach by indirect immunofluorescence; it was restricted to nervous structures. The majority of myenteric and submucous neurons were galanin-positive and received positive axo-somatic terminations. They also sent out axons staining positively. Galanin-positive fibers and a few atypically located neurons formed a mucous plexus at the bases of mucous glands. Varicose galanin fibers innervated the muscularis mucosae, circular and longitudinal muscle layers, while thick fascicles traversed the muscularis mucosae and circular muscle, possibly interconnecting the myenteric, submucous and mucous plexuses. Galanin-positive fibers did not supply blood vessels. There was no obvious gradient of innervation density along the esophagus, but the sphincter appeared to be more densely innervated than the esophageal body. There was no galanin-positive input to striated muscle. In view of its widespread distribution, this neuropeptide may serve multiple functions in the esophagus.