Vagal sparing gastrectomy: A systematic review and meta-analysis.

Radical gastrectomy is associated with significant functional complications. In appropriate patients may be amenable to less invasive resection aimed at preserving the vagal trunks. The aim of this systematic review and meta-analysis is to assess the functional consequences and oncological safety of vagal sparing gastrectomy (VSG) compared to conventional non-vagal sparing gastrectomy (CG). A systematic review of four databases was undertaken for studies published between 1/11990 and 15/122021, comparing patients who underwent VSG to CG. We meta-analysed the following outcomes: operative time, blood loss, nodal yield, days to flatus, body weight changes, as well as the incidence of post-operative cholelithiasis, diarrhoea, delayed gastric emptying, and dumping syndrome. Thirty studies were included in the meta-analysis with a selection of studies qualitatively analysed. VSG was associated with a lower rate of cholelithiasis (OR 0.25, 95% CI 0.15-0.41, p<0.010) and early dumping syndrome (OR 0.42, 95% CI 0.21 - 0.86; p=0.02), less blood loss (MD -51 ml, 95% CI -89.11 to -12.81 ml, p=0.009), less long term weight loss (MD 2.03%, 95% CI 0.31-3.76%, p=0.02) and a faster time to flatus (MD -0.42 days, 95% CI -0.48 - 0.36, p<0.001). There was no significant difference in nodal harvest, overall survival, and all other endpoints. VSG significantly reduces the incidence of post-operative cholelithiasis and dumping syndrome, decreases weight loss and facilitates an earlier return of gut motility. Although technically more challenging, VSG should be considered for prophylactic surgery.

Tumor-Infiltrating Neutrophils after Neoadjuvant Therapy are Associated with Poor Prognosis in Esophageal Cancer.

BACKGROUND: In esophageal cancer (EC), there is a paucity of knowledge regarding the interplay between the tumor immune microenvironment and response to neoadjuvant treatment and, therefore, which factors may influence outcomes. Thus, our goal was to investigate the changes in the immune microenvironment with neoadjuvant treatment in EC by assessing the expression of immune related genes and their association with prognosis. METHODS: We examined the transcriptome of paired pre- and post-neoadjuvant treated EC specimens. Based on these findings, we validated the presence of tumor-infiltrating neutrophils using CD15+ immunohistochemistry in a discovery cohort of patients with residual pathologic disease. We developed a nomogram as a predictor of progression-free survival (PFS) incorporating the variables CD15+ cell count, tumor regression grade, and tumor grade. RESULTS: After neoadjuvant treatment, there was an increase in genes related to myeloid cell differentiation and a poor prognosis associated with high neutrophil (CD15+) counts. Our nomogram incorporating CD15+ cell count was predictive of PFS with a C-index of 0.80 (95% confidence interval [CI] 0.68-0.9) and a concordance probability estimate (CPE) of 0.77 (95% CI 0.69-0.86), which indicates high prognostic ability. The C-index and CPE of the validation cohort were 0.81 (95% CI 0.69-0.91) and 0.78 (95% CI 0.7-0.86), respectively. CONCLUSIONS: Our nomogram incorporating CD15+ cell count can potentially be used to identify patients at high risk of recurrent disease and thus stratify patients who will benefit most from adjuvant treatment.

Potential Clinical Utility of a Targeted Circulating Tumor DNA Assay in Esophageal Adenocarcinoma.

OBJECTIVE: To explore the clinical utility of circulating tumor DNA (ctDNA) in esophageal adenocarcinoma (EAC) by developing a cost-effective and rapid technique utilising targeted amplicon sequencing. SUMMARY OF BACKGROUND DATA: Emerging evidence suggests that levels of ctDNA in the blood can be used to monitor treatment response and in the detection of disease recurrence in various cancer types. Current staging modalities for EAC such as computerised tomography of the chest/abdomen/pelvis (CT) and positron emission tomography (PET) do not reliably detect occult micro-metastatic disease, the presence of which signifies a poor prognosis. After curative-intent treatment, some patients are still at high risk of recurrent disease, and there is no widely accepted optimal surveillance tool for patients with EAC. METHODS: Sixty-two patients with EAC were investigated for the presence of ctDNA using a tumor-informed approach. We designed a custom targeted amplicon sequencing panel of target specific primers covering mutational foci in 9 of the most commonly mutated genes in EAC. Serial blood samples were taken before and after neoadjuvant treatment (NAT), and during surveillance. RESULTS: Somatic mutations were detected in pre-treatment biopsy samples of 55 out of 62 (89%) EAC patients. Mutations in TP53 (80%) were the most common. Out of these 55 patients, 20 (36%) had detectable ctDNA at baseline. The majority (90%) of patients with detectable ctDNA had either locally advanced tumors, nodal involvement or metastatic disease. In patients with locally advanced tumors, disease free survival (DFS) was more accurately stratified using pre-treatment ctDNA status [HR 4.34 (95% CI 0.93-20.21); P = 0.05] compared to nodal status on PET-CT. In an exploratory subgroup analysis, patients who are node negative but ctDNA positive have inferior DFS [HR 11.71 (95% CI 1.16-118.80) P = 0.04]. In blood samples taken before and following NAT, clearance of ctDNA after NAT was associated with a favourable response to treatment. Furthermore, patients who are ctDNA positive during post-treatment surveillance are at high risk of relapse. CONCLUSIONS: Our study shows that ctDNA has potential to provide additional prognostication over conventional staging investigation such as CT and PET. It may also have clinical utility in the assessment of response to NAT and as a biomarker for the surveillance of recurrent disease.

A MXI1-NUTM1 fusion protein with MYC-like activity suggests a novel oncogenic mechanism in a subset of NUTM1-rearranged tumors.

Most NUTM1-rearranged neoplasms (NRNs) have fusions between NUTM1 and BRD (bromodomain-containing) family members and are termed NUT carcinomas (NCs) because they show some squamous differentiation. However, some NRNs are associated with fusions between NUTM1 and members of the MAD (MAX dimerization) gene family of MYC antagonists. Here we describe a small round cell malignancy from the gastro-esophageal junction with a previously unreported fusion between NUTM1 and the MAD family member MXI1. In contrast to NCs, the MXI1-NUTM1 tumor did not show squamous differentiation and did not express MYC, TP63 or SOX2, genes known to be targets of BRD-NUTM1 proteins and critical for NC oncogenesis. Transcriptome analysis showed paradoxical enrichment of MYC target genes in the MXI1-NUTM1 tumor despite the lack of MYC expression. When expressed in vitro MXI1-NUTM1 partially phenocopied MYC, enhancing cell proliferation and cooperating with oncogenic HRAS to produce anchorage-independent cell growth. These data provide evidence that MAD family members, which are normally repressors of MYC activity, can be converted into MYC-like mimics by fusion to NUTM1. The pathological features and novel oncogenic mechanism of the MXI1-NUTM1 tumor show that identification of NUTM1 fusion partners can be important for accurate diagnostic classification of some NRN subtypes, and potentially may guide therapeutic options.

GRB7 is an oncogenic driver and potential therapeutic target in oesophageal adenocarcinoma.

Efficacious therapeutic approaches are urgently needed to improve outcomes in patients with oesophageal adenocarcinoma (OAC). However, oncogenic drivers amenable to targeted therapy are limited and their functional characterisation is essential. Among few targeted therapies available, anti-human epidermal growth factor receptor 2 (HER2) therapy showed only modest benefit for patients with OAC. Herein, we investigated the potential oncogenic role of growth factor receptor bound protein 7 (GRB7), which is reported to be co-amplified with HER2 (ERBB2) in OAC. GRB7 was highly expressed in 15% of OAC tumours, not all of which could be explained by co-amplification with HER2, and was associated with a trend for poorer overall survival. Knockdown of GRB7 decreased proliferation and clonogenic survival, and induced apoptosis. Reverse phase protein array (RPPA) analyses revealed a role for PI3K, mammalian target of rapamycin (mTOR), MAPK, and receptor tyrosine kinase signalling in the oncogenic action of GRB7. Furthermore, the GRB7 and HER2 high-expressing OAC cell line Eso26 showed reduced cell proliferation upon GRB7 knockdown but was insensitive to HER2 inhibition by trastuzumab. Consistent with this, GRB7 knockdown in vivo with an inducible shRNA significantly inhibited tumour growth in cell line xenografts. HER2 expression did not predict sensitivity to trastuzumab, with Eso26 xenografts remaining refractory to trastuzumab treatment. Taken together, our study provides strong evidence for an oncogenic role for GRB7 in OAC and suggests that targeting GRB7 may be a potential therapeutic strategy for this cancer. © 2020 The Authors. The Journal of Pathology published by John Wiley & Sons, Ltd. on behalf of The Pathological Society of Great Britain and Ireland.

Clinical pathways and outcomes of patients with Barrett's esophagus in tertiary care settings: a prospective longitudinal cohort study in Australia, 2008-2016.

BACKGROUND: Clinical services for Barrett's esophagus have been rising worldwide including Australia, but little is known of the long-term outcomes of such patients. Retrospective studies using data at baseline are prone to both selection and misclassification bias. We investigated the clinical characteristics and outcomes of Barrett's esophagus patients in a prospective cohort. METHODS: We recruited patients diagnosed with Barrett's esophagus in tertiary settings across Australia between 2008 and 2016. We compared baseline and follow-up epidemiological and clinical data between Barrett's patients with and without dysplasia. We calculated age-adjusted incidence rates and estimated minimally and fully adjusted hazard ratios (HR) to identify those clinical factors related to disease progression. RESULTS: The cohort comprised 268 patients with Barrett's esophagus (median follow-up 5 years). At recruitment, 224 (84%) had no dysplasia, 44 (16%) had low-grade or indefinite dysplasia (LGD/IND). The age-adjusted incidence of esophageal adenocarcinoma (EAC) was 0.5% per year in LGD/IND compared with 0.1% per year in those with no dysplasia. Risk of progression to high-grade dysplasia/EAC was associated with prior LGD/IND (fully adjusted HR 6.55, 95% confidence interval [CI] 1.96-21.8) but not long-segment disease (HR 1.03, 95%CI 0.29-3.58). CONCLUSIONS: These prospective data suggest presence of dysplasia is a stronger predictor of progression to cancer than segment length in patients with Barrett's esophagus.

Intramuscular Transplantation Improves Engraftment Rates for Esophageal Patient-Derived Tumor Xenografts.

BACKGROUND: Recently, there has been an increase in the availability of targeted molecular therapies for cancer treatment. The application of these approaches to esophageal cancer, however, has been hampered by the relative lack of appropriate models for preclinical testing. Patient-derived tumor xenograft (PDTX) models are gaining popularity for studying many cancers. Unfortunately, it has proven difficult to generate xenografts from esophageal cancer using these models. The purpose of this study was to improve the engraftment efficiency of esophageal PDTXs. METHODS: Fresh pieces of esophageal tumors obtained from endoscopic biopsies or resected specimens were collected from 23 patients. The tumors were then coated in Matrigel and transplanted in immunocompromised mice subcutaneously (n = 6) and/or using a novel implantation technique whereby the tumor is placed in a dorsal intramuscular pocket (n = 18). They are then monitored for engraftment. RESULTS: With the novel intramuscular technique, successful engraftment was achieved for all 18 patient tumors. Among these PDTXs, 13 recapitulated the original patient tumors with respect to degree of differentiation, molecular and genetic profiles, and chemotherapeutic response. Lymphomatous transformation was observed in the other five PDTXs. Successful engraftment was achieved for only one of six patient tumors using the classic subcutaneous approach. DISCUSSION: We achieved a much higher engraftment rate of PDTXs using our novel intramuscular transplant technique than has been reported in other published studies. It is hoped that this advancement will help expedite the development and testing of new therapies for esophageal cancer.

Incremental diagnostic utility of gastric distension FDG PET/CT.

PURPOSE: To assess the diagnostic utility of gastric distension (GD) FDG PET/CT in both patients with known gastric malignancy and those not known to have gastric malignancy but with incidental focal FDG uptake in the stomach. METHODS: This retrospective analysis included 88 patients who underwent FDG PET/CT following GD with hyoscine N-butylbromide (Buscopan®) and water ingestion as part of routine clinical evaluation between 2004 and 2014. FDG PET/CT scans before and after GD were reported blinded to the patient clinical details in 49 patients undergoing pretreatment staging of gastric malignancy and 39 patients who underwent GD following incidental suspicious gastric uptake. The PET findings were validated by a composite clinical standard. RESULTS: In the 49 patients undergoing pretreatment staging of gastric malignancy, GD improved PET detection of the primary tumour (from 80 % to 90 %). PET evaluation of tumour extent was concordant with endoscopic/surgical reports in 31 % (interpreter 1) and 45 % (interpreter 2) using pre-GD images and 73 % and 76 % using GD images. Interobserver agreement also improved with GD (κ = 0.29 to 0.69). Metabolic and morphological quantitative analysis demonstrated a major impact of GD in normal gastric wall but no significant effect in tumour, except a minor increase in SUV related to a delayed acquisition time. The tumour to normal stomach SUVmax ratio increased from 3.8 ± 2.9 to 9.2 ± 8.6 (mean ± SD) with GD (p < 0.0001), facilitating detection and improved assessment of the primary tumour. In 25 (64 %) of the 39 patients with incidental suspicious gastric uptake, acquisition after GD correctly excluded a malignant process. In 10 (71 %) of the remaining 14 patients with persistent suspicious FDG uptake despite GD, malignancy was confirmed and in 3 (21 %) an active but benign pathology was diagnosed. CONCLUSION: GD is a simple way to improve local staging with FDG PET in patients with gastric malignancy. In the setting of incidental suspicious gastric uptake, GD is also an effective tool for ruling out malignancy and leads to the avoidance of unnecessary endoscopy.

The Genetics of Barrett's Esophagus: A Familial and Population-Based Perspective.

Barrett's esophagus (BE) is intestinal metaplasia of the lower esophagus and a precursor lesion for esophageal adenocarcinoma (EAC). Both are important health issues as they have rising incidences in the Western world. Improving the management of BE relies on understanding the underlying biology of this disease, but the exact biological mechanisms have been difficult to determine. BE is generally thought to be an acquired condition that develops secondarily to chronic gastroesophageal reflux. However, multiple reports of familial clustering of patients with BE and/or EAC suggest a possible inherited predisposition to BE may be driving this condition, at least in a subset of patients. Identifying the genetic variants that predispose to BE in these families would open up the possibility for blood-based screening tests that could inform decision-making in regard to surveillance strategies, particularly for relatives of patients with BE and/or EAC. Perhaps more importantly, understanding the genetic mechanisms that predispose to BE may provide valuable insights into the biology of this condition and potentially identify novel targets for therapeutic intervention. Here we review the current evidence for a genetic predisposition to BE and discuss the potential implications of these findings.

APR-246 potently inhibits tumour growth and overcomes chemoresistance in preclinical models of oesophageal adenocarcinoma.

OBJECTIVES: p53 is a critical tumour suppressor and is mutated in 70% of oesophageal adenocarcinomas (OACs), resulting in chemoresistance and poor survival. APR-246 is a first-in-class reactivator of mutant p53 and is currently in clinical trials. In this study, we characterised the activity of APR-246 and its effect on p53 signalling in a large panel of cell line xenograft (CLX) and patient-derived xenograft (PDX) models of OAC. DESIGN: In vitro response to APR-246 was assessed using clonogenic survival, cell cycle and apoptosis assays. Ectopic expression, gene knockdown and CRISPR/Cas9-mediated knockout studies of mutant p53 were performed to investigate p53-dependent drug effects. p53 signalling was examined using quantitative RT-PCR and western blot. Synergistic interactions between APR-246 and conventional chemotherapies were evaluated in vitro and in vivo using CLX and PDX models. RESULTS: APR-246 upregulated p53 target genes, inhibited clonogenic survival and induced cell cycle arrest as well as apoptosis in OAC cells harbouring p53 mutations. Sensitivity to APR-246 correlated with cellular levels of mutant p53 protein. Ectopic expression of mutant p53 sensitised p53-null cells to APR-246, while p53 gene knockdown and knockout diminished drug activity. Importantly, APR-246 synergistically enhanced the inhibitory effects of cisplatin and 5-fluorouracil through p53 accumulation. Finally, APR-246 demonstrated potent antitumour activity in CLX and PDX models, and restored chemosensitivity to a cisplatin/5-fluorouracil-resistant xenograft model. CONCLUSIONS: APR-246 has significant antitumour activity in OAC. Given that APR-246 is safe at therapeutic levels our study strongly suggests that APR-246 can be translated into improving the clinical outcomes for OAC patients.

Molecular changes in the phosphatidylinositide 3-kinase (PI3K) pathway are common in gastric cancer.

BACKGROUND AND OBJECTIVES: The phosphatidylinositide 3-kinase (PI3K) pathway is an important signalling pathway that is frequently activated in cancer cells. This has led to the emergence of PI3K inhibitors as potential new treatment modalities for many cancers. We have investigated the frequency of molecular changes in the PI3K pathway in gastric cancer. METHODS: A series of sixty one human gastric cancer specimens and nine human gastric cancer cell lines were screened for PIK3CA mutations and copy number gain by direct sequencing and multiplex ligation-dependent probe amplification (MLPA), respectively. PTEN protein levels were assessed by immunohistochemistry. RESULTS: Alterations in the PI3K pathway were found in 33 of 61 (54%) gastric tumours. PIK3CA mutation and copy number gain were detected in 3 (4.9%) and 8 (13.1%), respectively, of 61 gastric cancer samples while PTEN loss was detected in 24 (39%) of the tumours. Two tumours had both PTEN loss and PIK3CA copy number gain. There were no significant associations between these PI3K pathway changes and the clinical features of the tumours. CONCLUSIONS: Alterations in the PI3K pathway are frequent in gastric tumours implicating this pathway as a legitimate therapeutic target in gastric cancer.

Using gene expression profiling to predict response and prognosis in gastrointestinal cancers-the promise and the perils.

Cancer treatment is now moving toward a personalized approach, promising improved rates of response and survival. A number of studies have employed the use of microarrays to investigate the predictive potential of expression profiling in gastrointestinal (GI) cancer patients. However while many robust predictive classifiers relating to response and prognosis have been generated for GI cancer patients, these have yet to make the transition to the clinic. The main obstacle is the limited cross validation between predictive gene lists identified for the same tumor type and outcome. Differences in the experimental design, analysis, and interpretation of results all contribute to this variation, with numerous factors influencing which genes are highlighted as predictive. While predictive genomics shows immense potential, it is still a relatively new field and the validation of predictive gene lists derived from microarray data remains a challenge. Future studies must carefully consider all aspects of experimental design to ensure a clinically applicable predictive test can be developed. With this in mind, more extensive and collaborative research must be undertaken before microarray-based platforms can be used routinely in tailoring GI cancer treatment and change clinical practice. Larger cohorts and consistency in methodology will enable the findings from this research to make the transition to the clinic.

Loss of SMAD4 Is Sufficient to Promote Tumorigenesis in a Model of Dysplastic Barrett's Esophagus.

BACKGROUND & AIMS: Esophageal adenocarcinoma (EAC) develops from its precursor Barrett's esophagus through intermediate stages of low- and high-grade dysplasia. However, knowledge of genetic drivers and molecular mechanisms implicated in disease progression is limited. Herein, we investigated the effect of Mothers against decapentaplegic homolog 4 (SMAD4) loss on transforming growth factor ß (TGF-ß) signaling functionality and in vivo tumorigenicity in high-grade dysplastic Barrett's cells. METHODS: An in vivo xenograft model was used to test tumorigenicity of SMAD4 knockdown or knockout in CP-B high-grade dysplastic Barrett's cells. RT2 polymerase chain reaction arrays were used to analyze TGF-ß signaling functionality, and low-coverage whole-genome sequencing was performed to detect copy number alterations upon SMAD4 loss. RESULTS: We found that SMAD4 knockout significantly alters the TGF-ß pathway target gene expression profile. SMAD4 knockout positively regulates potential oncogenes such as CRYAB, ACTA2, and CDC6, whereas the CDKN2A/B tumor-suppressor locus was regulated negatively. We verified that SMAD4 in combination with CDC6-CDKN2A/B or CRYAB genetic alterations in patient tumors have significant predictive value for poor prognosis. Importantly, we investigated the effect of SMAD4 inactivation in Barrett's tumorigenesis. We found that genetic knockdown or knockout of SMAD4 was sufficient to promote tumorigenesis in dysplastic Barrett's esophagus cells in vivo. Progression to invasive EAC was accompanied by distinctive and consistent copy number alterations in SMAD4 knockdown or knockout xenografts. CONCLUSIONS: Altogether, up-regulation of oncogenes, down-regulation of tumor-suppressor genes, and chromosomal instability within the tumors after SMAD4 loss implicates SMAD4 as a protector of genome integrity in EAC development and progression. Foremost, SMAD4 loss promotes tumorigenesis from dysplastic Barrett's toward EAC.

The Upper Gastrointestinal Cancer Registry (UGICR): a clinical quality registry to monitor and improve care in upper gastrointestinal cancers.

PURPOSE: The Upper Gastrointestinal Cancer Registry (UGICR) was developed to monitor and improve the quality of care provided to patients with upper gastrointestinal cancers in Australia. PARTICIPANTS: It supports four cancer modules: pancreatic, oesophagogastric, biliary and primary liver cancer. The pancreatic cancer (PC) module was the first module to be implemented, with others being established in a staged approach. Individuals are recruited to the registry if they are aged 18 years or older, have received care for their cancer at a participating public/private hospital or private clinic in Australia and do not opt out of participation. FINDINGS TO DATE: The UGICR is governed by a multidisciplinary steering committee that provides clinical governance and oversees clinical working parties. The role of the working parties is to develop quality indicators based on best practice for each registry module, develop the minimum datasets and provide guidance in analysing and reporting of results. Data are captured from existing data sources (population-based cancer incidence registries, pathology databases and hospital-coded data) and manually from clinical records. Data collectors directly enter information into a secure web-based Research Electronic Data Capture (REDCap) data collection platform. The PC module began with a pilot phase, and subsequently, we used a formal modified Delphi consensus process to establish a core set of quality indicators for PC. The second module developed was the oesophagogastric cancer (OGC) module. Results of the 1 year pilot phases for PC and OGC modules are included in this cohort profile. FUTURE PLANS: The UGICR will provide regular reports of risk-adjusted, benchmarked performance on a range of quality indicators that will highlight variations in care and clinical outcomes at a health service level. The registry has also been developed with the view to collect patient-reported outcomes (PROs), which will further add to our understanding of the care of patients with these cancers.

Inhibiting system xC- and glutathione biosynthesis - a potential Achilles' heel in mutant-p53 cancers.

Effective therapeutic strategies to target mutant tumor protein p53 (TP53, best known as p53) cancers remain an unmet medical need. We found that mutant p53 impairs the function of nuclear factor (erythroid-derived 2)-like 2 (NFE2L2, commonly known as NRF2), suppresses solute carrier family 7 member 11 (SLC7A11) expression, and diminishes cellular glutamate/cystine exchange. This decreases glutathione biosynthesis, resulting in redox imbalance. Mutant p53 tumors are thus inherently susceptible to further perturbations of the SLC7A11/glutathione axis.

Inhibiting the system xC-/glutathione axis selectively targets cancers with mutant-p53 accumulation.

TP53, a critical tumour suppressor gene, is mutated in over half of all cancers resulting in mutant-p53 protein accumulation and poor patient survival. Therapeutic strategies to target mutant-p53 cancers are urgently needed. We show that accumulated mutant-p53 protein suppresses the expression of SLC7A11, a component of the cystine/glutamate antiporter, system xC-, through binding to the master antioxidant transcription factor NRF2. This diminishes glutathione synthesis, rendering mutant-p53 tumours susceptible to oxidative damage. System xC- inhibitors specifically exploit this vulnerability to preferentially kill cancer cells with stabilized mutant-p53 protein. Moreover, we demonstrate that SLC7A11 expression is a novel and robust predictive biomarker for APR-246, a first-in-class mutant-p53 reactivator that also binds and depletes glutathione in tumours, triggering lipid peroxidative cell death. Importantly, system xC- antagonism strongly synergizes with APR-246 to induce apoptosis in mutant-p53 tumours. We propose a new paradigm for targeting cancers that accumulate mutant-p53 protein by inhibiting the SLC7A11-glutathione axis.

Preclinical models of esophageal adenocarcinoma for drug development.

The advent of multi-omic profiling of tumors, together with the increasing affordability of high-throughput drug screening programs, has helped usher in a new era of molecular targeted therapies for many solid malignancies. However, there has been limited success in esophageal adenocarcinoma. Until recently, preclinical drug development for this cancer has been largely limited to a small number of cell line models. Now, the increasing availability of patient-derived xenografts and novel metastatic models are helping to bridge the gap between scientific discovery and patient care. These platforms are valuable adjuncts for drug testing. Nevertheless, further work is required to develop systems that model the tumor microenvironment, including cancer cell interactions with the immune system, which will be particularly relevant for the translation of novel immunotherapies for esophageal adenocarcinoma.

Novel metastatic models of esophageal adenocarcinoma derived from FLO-1 cells highlight the importance of E-cadherin in cancer metastasis.

There is currently a paucity of preclinical models available to study the metastatic process in esophageal cancer. Here we report FLO-1, and its isogenic derivative FLO-1LM, as two spontaneously metastatic cell line models of human esophageal adenocarcinoma. We show that FLO-1 has undergone epithelial-mesenchymal transition and metastasizes following subcutaneous injection in mice. FLO-1LM, derived from a FLO-1 liver metastasis, has markedly enhanced proliferative, clonogenic, anti-apoptotic, invasive, immune-tolerant and metastatic potential. Genome-wide RNAseq profiling revealed a significant enrichment of metastasis-related pathways in FLO-1LM cells. Moreover, CDH1, which encodes the adhesion molecule E-cadherin, was the most significantly downregulated gene in FLO-1LM compared to FLO-1. Consistent with this, repression of E-cadherin expression in FLO-1 cells resulted in increased metastatic activity. Importantly, reduced E-cadherin expression is commonly reported in esophageal adenocarcinoma and independently predicts poor patient survival. Collectively, these findings highlight the biological importance of E-cadherin activity in the pathogenesis of metastatic esophageal adenocarcinoma and validate the utility of FLO-1 parental and FLO-1LM cells as preclinical models of metastasis in this disease.