LC-MS/MS based detection of circulating proinsulin derived peptides in patients with altered pancreatic beta cell function.

Routine immunoassays for insulin and C-peptide have the potential to cross-react with partially processed proinsulin products, although in healthy patients these are present at such low levels that the interference is insignificant. Elevated concentrations of proinsulin and des-31,32 proinsulin arising from pathological conditions, or injected insulin analogues, however can cause significant assay interferences, complicating interpretation. Clinical diagnosis and management therefore sometimes require methods that can distinguish true insulin and C-peptide from partially processed proinsulin or injected insulin analogues. In this scenario, the high specificity of mass spectrometric analysis offers potential benefit for patient care. A high throughput targeted LC-MS/MS method was developed as a fit for purpose investigation of insulin, insulin analogues, C-peptide and proinsulin processing intermediates in plasma samples from different patient groups. Using calibration standards and bovine insulin as an internal standard, absolute concentrations of insulin and C-peptide were quantified across a nominal human plasma postprandial range and correlated strongly with immunoassay-based measurements. The ability to distinguish between insulin, insulin analogues and proinsulin intermediates in a single extraction is an improvement over existing immunological based techniques, offering the advantage of exact identification of the species being measured. The method promises to aid in the detection of circulating peptides which have previously been overlooked but may interfere with standard insulin and C-peptide immunoassays.

International Delphi consensus guidelines for follow-up after prophylactic total gastrectomy: the Life after Prophylactic Total Gastrectomy (LAP-TG) study.

BACKGROUND: Prophylactic total gastrectomy (PTG) remains the only means of preventing gastric cancer for people with genetic mutations predisposing to Hereditary Diffuse Gastric Cancer (HDGC), mainly in the CDH1 gene. The small but growing cohort of people undergoing PTG at a young age are expected to have a life-expectancy close to the general population, however, knowledge of the long-term effects of, and monitoring requirements after, PTG is limited. This study aims to define the standard of care for follow-up after PTG. METHODS: Through a combination of literature review and two-round Delphi consensus of major HDGC/PTG units and physicians, and patient advocates, we produced a set of recommendations for follow-up after PTG. RESULTS: There were 42 first round, and 62 second round, responses from clinicians, allied health professionals and patient advocates. The guidelines include recommendations for timing of assessments and specialties involved in providing follow-up, micronutrient supplementation and monitoring, bone health and the provision of written information. CONCLUSION: While the evidence supporting the guidelines is limited, expert consensus provides a framework to best manage people following PTG, and could support the collection of information on the long-term effects of PTG.

Peptidomics of enteroendocrine cells and characterisation of potential effects of a novel preprogastrin derived-peptide on glucose tolerance in lean mice.

OBJECTIVES: To analyse the peptidomics of mouse enteroendocrine cells (EECs) and human gastrointestinal (GI) tissue and identify novel gut derived peptides. METHODS: High resolution nano-flow liquid chromatography mass spectrometry (LC-MS/MS) was performed on (i) flow-cytometry purified NeuroD1 positive cells from mouse and homogenised human intestinal biopsies, (ii) supernatants from primary murine intestinal cultures, (iii) intestinal homogenates from mice fed high fat diet. Candidate bioactive peptides were selected on the basis of species conservation, high expression/biosynthesis in EECs and evidence of regulated secretionin vitro. Candidate novel gut-derived peptides were chronically administered to mice to assess effects on food intake and glucose tolerance. RESULTS: A large number of peptide fragments were identified from human and mouse, including known full-length gut hormones and enzymatic degradation products. EEC-specific peptides were largely from vesicular proteins, particularly prohormones, granins and processing enzymes, of which several exhibited regulated secretion in vitro. No regulated peptides were identified from previously unknown genes. High fat feeding particularly affected the distal colon, resulting in reduced peptide levels from GCG, PYY and INSL5. Of the two candidate novel peptides tested in vivo, a peptide from Chromogranin A (ChgA 435-462a) had no measurable effect, but a progastrin-derived peptide (Gast p59-79), modestly improved glucose tolerance in lean mice. CONCLUSION: LC-MS/MS peptidomic analysis of murine EECs and human GI tissue identified the spectrum of peptides produced by EECs, including a potential novel gut hormone, Gast p59-79, with minor effects on glucose tolerance.

Single-bite versus double-bite technique for mapping biopsies during endoscopic surveillance for hereditary diffuse gastric cancer: a single-center, randomized trial.

BACKGROUND : Endoscopic surveillance is recommended in patients with hereditary diffuse gastric cancer (HDGC) who refuse or want to delay surgery. Because early signet-ring cell carcinoma (SRCC) can be inconspicuous, the current surveillance endoscopy protocol entails 30 random biopsies, which are time-consuming. This study aimed to compare single-bite and double-bite techniques in HDGC surveillance. METHODS : Between October 2017 and December 2018, consecutive patients referred for HDGC surveillance were prospectively randomized to the single- or double-bite arm. The primary outcome was the diagnostic yield for SRCC foci. Secondary outcomes were: procedural time for random biopsies; comfort score; biopsy size; and quality of specimens, the latter assessed by the presence of muscularis mucosa, crush artifact, and proportion usable for diagnostic assessment. RESULTS : 25 patients were randomized to the single-bite arm and 23 to the double-bite arm. SRCC foci were detected in three and four patients in the single- and double-bite arms, respectively (P = 0.70). The procedural time for the double-bite arm (12 minutes, interquartile range [IQR] 4) was significantly shorter than for the single-bite arm (15 minute, IQR 6; P = 0.01), but comfort scores were similar. The size of the biopsies in the double-bite arm was significantly smaller than in single-bite arm (2.5 mm vs. 3.0 mm; P < 0.001) but this did not affect the presence of muscularis mucosa (P = 0.73), artifact level (P = 0.11), and diagnostic utility (P = 0.051). CONCLUSION : For patients undergoing HDGC surveillance, the double-bite technique is significantly faster than the single-bite technique. The diagnostic yield for SRCC and the biopsy quality were similar across both groups.

Ileo-colonic delivery of conjugated bile acids improves glucose homeostasis via colonic GLP-1-producing enteroendocrine cells in human obesity and diabetes.

BACKGROUND: The bile acid (BA) pathway plays a role in regulation of food intake and glucose metabolism, based mainly on findings in animal models. Our aim was to determine whether the BA pathway is altered and correctable in human obesity and diabetes. METHODS: We conducted 3 investigations: 1) BA receptor pathways were studied in NCI-H716 enteroendocrine cell (EEC) line, whole human colonic mucosal tissue and in human colonic EEC isolated by Fluorescence-activated Cell Sorting (ex vivo) from endoscopically-obtained biopsies colon mucosa; 2) We characterized the BA pathway in 307 participants by measuring during fasting and postprandial levels of FGF19, 7αC4 and serum BA; 3) In a placebo-controlled, double-blind, randomised, 28-day trial, we studied the effect of ileo-colonic delivery of conjugated BAs (IC-CBAS) on glucose metabolism, incretins, and lipids, in participants with obesity and diabetes. FINDINGS: Human colonic GLP-1-producing EECs express TGR5, and upon treatment with bile acids in vitro, human EEC differentially expressed GLP-1 at the protein and mRNA level. In Ussing Chamber, GLP-1 release was stimulated by Taurocholic acid in either the apical or basolateral compartment. FGF19 was decreased in obesity and diabetes compared to controls. When compared to placebo, IC-CBAS significantly decreased postprandial glucose, fructosamine, fasting insulin, fasting LDL, and postprandial FGF19 and increased postprandial GLP-1 and C-peptide. Increase in faecal BA was associated with weight loss and with decreased fructosamine. INTERPRETATIONS: In humans, BA signalling machinery is expressed in colonic EECs, deficient in obesity and diabetes, and when stimulated with IC-CBAS, improved glucose homeostasis. ClinicalTrials.gov number, NCT02871882, NCT02033876. FUNDING: Research support and drug was provided by Satiogen Pharmaceuticals (San Diego, CA). AA, MC, and NFL report grants (AA- C-Sig P30DK84567, K23 DK114460; MC- NIH R01 DK67071; NFL- R01 DK057993) from the NIH. JR was supported by an Early Career Grant from Society for Endocrinology.

Endoscopic measurement of gastric pH associates with persistent acid reflux in patients treated with proton-pump inhibitors for gastroesophageal reflux disease.

Background: Proton-pump inhibitors (PPIs) are the mainstay of gastroesophageal reflux disease (GERD) treatment, however, up to 30% of patients have a poor symptomatic response. PH-impedance is the gold standard to assess whether this is due to persistent acid reflux. We aimed to characterize clinical predictors of persistent esophageal acid reflux on PPIs including gastric pH measured during endoscopy. Methods: We prospectively recruited patients with GERD and/or Barrett's esophagus (BE) on PPIs. All patients completed a symptom questionnaire (RDQ) and underwent gastroscopy with gastric pH analysis, immediately followed by ambulatory 24-hour pH-impedance. We used a modified cut-off of 1.3% for pathological esophageal acid exposure time (AET). Multiple linear regression model was used to analyze the correlation between AET and predictive variables. Results: We recruited 122 patients, of which 92 (75.4%) were included in the final analysis [44 male (47.8%), median age 53 years (IQR: 43-66)]. Forty-four patients (47.8%) had persistent acid reflux with a median total AET of 2.2 (IQR1.2-5.0), as compared to 0.1 (IQR 0.0-0.2) in patients without persistent reflux (n=48; P<.001). There was no difference in age, gender, BMI, PPI-regimen, diagnosis of hiatus hernia or BE, and severity of symptoms between patients with normal and abnormal AET. Median gastric pH was significantly lower in patients with abnormal AET (5.8 vs 6.6, P=0.032) and it correlated with the total AET (P=.045; R2=12.0%). With a pH cut-off of 5.05, single point endoscopic gastric pH analysis had an area under the ROC curve (AUC) of 63.0% (95%CI 51.3-74.7) for prediction of pathological esophageal AET. Conclusions: Symptoms and clinical characteristics are not useful to predict persistent acid reflux in patients on PPIs. One-point gastric pH correlates with 24-hour esophageal AET and could guide clinicians to assess response to PPIs, however, its utility needs validation in larger studies.

Characterisation of proguanylin expressing cells in the intestine - evidence for constitutive luminal secretion.

Guanylin, a peptide implicated in regulation of intestinal fluid secretion, is expressed in the mucosa, but the exact cellular origin remains controversial. In a new transgenic mouse model fluorescent reporter protein expression driven by the proguanylin promoter was observed throughout the small intestine and colon in goblet and Paneth(-like) cells and, except in duodenum, in mature enterocytes. In Ussing chamber experiments employing both human and mouse intestinal tissue, proguanylin was released predominantly in the luminal direction. Measurements of proguanylin expression and secretion in cell lines and organoids indicated that secretion is largely constitutive and requires ER to Golgi transport but was not acutely regulated by salt or other stimuli. Using a newly-developed proguanylin assay, we found plasma levels to be raised in humans after total gastrectomy or intestinal transplantation, but largely unresponsive to nutrient ingestion. By LC-MS/MS we identified processed forms in tissue and luminal extracts, but in plasma we only detected full-length proguanylin. Our transgenic approach provides information about the cellular origins of proguanylin, complementing previous immunohistochemical and in-situ hybridisation results. The identification of processed forms of proguanylin in the intestinal lumen but not in plasma supports the notion that the primary site of action is the gut itself.

Important Role of the GLP-1 Axis for Glucose Homeostasis after Bariatric Surgery.

Bariatric surgery is widely used to treat obesity and improves type 2 diabetes beyond expectations from the degree of weight loss. Elevated post-prandial concentrations of glucagon-like peptide 1 (GLP-1), peptide YY (PYY), and insulin are widely reported, but the importance of GLP-1 in post-bariatric physiology remains debated. Here, we show that GLP-1 is a major driver of insulin secretion after bariatric surgery, as demonstrated by blocking GLP-1 receptors (GLP1Rs) post-gastrectomy in lean humans using Exendin-9 or in mice using an anti-GLP1R antibody. Transcriptomics and peptidomics analyses revealed that human and mouse enteroendocrine cells were unaltered post-surgery; instead, we found that elevated plasma GLP-1 and PYY correlated with increased nutrient delivery to the distal gut in mice. We conclude that increased GLP-1 secretion after bariatric surgery arises from rapid nutrient delivery to the distal gut and is a key driver of enhanced insulin secretion.

Comparison of Human and Murine Enteroendocrine Cells by Transcriptomic and Peptidomic Profiling.

Enteroendocrine cells (EECs) produce hormones such as glucagon-like peptide 1 and peptide YY that regulate food absorption, insulin secretion, and appetite. Based on the success of glucagon-like peptide 1-based therapies for type 2 diabetes and obesity, EECs are themselves the focus of drug discovery programs to enhance gut hormone secretion. The aim of this study was to identify the transcriptome and peptidome of human EECs and to provide a cross-species comparison between humans and mice. By RNA sequencing of human EECs purified by flow cytometry after cell fixation and staining, we present a first transcriptomic analysis of human EEC populations and demonstrate a strong correlation with murine counterparts. RNA sequencing was deep enough to enable identification of low-abundance transcripts such as G-protein-coupled receptors and ion channels, revealing expression in human EECs of G-protein-coupled receptors previously found to play roles in postprandial nutrient detection. With liquid chromatography-tandem mass spectrometry, we profiled the gradients of peptide hormones along the human and mouse gut, including their sequences and posttranslational modifications. The transcriptomic and peptidomic profiles of human and mouse EECs and cross-species comparison will be valuable tools for drug discovery programs and for understanding human metabolism and the endocrine impacts of bariatric surgery.

GDF15 Provides an Endocrine Signal of Nutritional Stress in Mice and Humans.

GDF15 is an established biomarker of cellular stress. The fact that it signals via a specific hindbrain receptor, GFRAL, and that mice lacking GDF15 manifest diet-induced obesity suggest that GDF15 may play a physiological role in energy balance. We performed experiments in humans, mice, and cells to determine if and how nutritional perturbations modify GDF15 expression. Circulating GDF15 levels manifest very modest changes in response to moderate caloric surpluses or deficits in mice or humans, differentiating it from classical intestinally derived satiety hormones and leptin. However, GDF15 levels do increase following sustained high-fat feeding or dietary amino acid imbalance in mice. We demonstrate that GDF15 expression is regulated by the integrated stress response and is induced in selected tissues in mice in these settings. Finally, we show that pharmacological GDF15 administration to mice can trigger conditioned taste aversion, suggesting that GDF15 may induce an aversive response to nutritional stress.

Peptidomic analysis of endogenous plasma peptides from patients with pancreatic neuroendocrine tumours.

RATIONALE: Diagnosis of pancreatic neuroendocrine tumours requires the study of patient plasma with multiple immunoassays, using multiple aliquots of plasma. The application of mass spectrometry based techniques could reduce the cost and amount of plasma required for diagnosis. METHODS: Plasma samples from two patients with pancreatic neuroendocrine tumours were extracted using an established acetonitrile-based plasma peptide enrichment strategy. The circulating peptidome was characterised using nano and high flow rate liquid chromatography/mass spectrometry (LC/MS) analyses. To assess the diagnostic potential of the analytical approach, a large sample batch (68 plasmas) from control subjects, and aliquots from subjects harbouring two different types of pancreatic neuroendocrine tumour (insulinoma and glucagonoma), were analysed using a 10-min LC/MS peptide screen. RESULTS: The untargeted plasma peptidomics approach identified peptides derived from the glucagon prohormone, chromogranin A, chromogranin B and other peptide hormones and proteins related to control of peptide secretion. The glucagon prohormone derived peptides that were detected were compared against putative peptides that were identified using multiple antibody pairs against glucagon peptides. Comparison of the plasma samples for relative levels of selected peptides showed clear separation between the glucagonoma and the insulinoma and control samples. CONCLUSIONS: The combination of the organic solvent extraction methodology with high flow rate analysis could potentially be used to aid diagnosis and monitor treatment of patients with functioning pancreatic neuroendocrine tumours. However, significant validation will be required before this approach can be clinically applied.

Gastrectomy with Roux-en-Y reconstruction as a lean model of bariatric surgery.

BACKGROUND: Altered enteroendocrine hormone responses are widely believed to underlie the beneficial effects of bariatric surgery in type 2 diabetes. While elevated postprandial glucagon-like peptide-1 (GLP-1) is considered one of the mediators, increased postprandial glucagon levels have recently been implicated. OBJECTIVES: We investigated hormonal responses in lean patients after prophylactic total gastrectomy (PTG), as a model of Roux-en-Y gastric bypass without the confounding effects of obesity or massive weight loss. SETTING: University hospital, United Kingdom. METHODS: Ten participants after PTG and 9 healthy volunteers were recruited for oral glucose tolerance tests. Plasma glucose, insulin, GLP-1, peptide YY, glucose-dependent insulinotropic-polypeptide, glucagon, oxyntomodulin, glucagon(1-61), and glicentin levels were assessed using immunoassays and/or mass spectrometry. RESULTS: PTG participants exhibited accelerated plasma glucose appearance, followed, in 3 of 10 cases, by hypoglycemia (<3 mM glucose). Plasma GLP-1, peptide YY, glucose-dependent insulinotropic-polypeptide, glicentin, and oxyntomodulin responses were elevated, and glucagon appeared to rise in PTG participants when measured with a glucagon-specific enzyme-linked immunosorbent assay. We revisited the specificity of this assay, and demonstrated significant cross-reactivity with glicentin and oxyntomodulin at concentrations observed in PTG plasma. Reassessment of glucagon with the same assay using a modified protocol, and by liquid chromatography-mass spectrometry, demonstrated suppression of glucagon secretion after oral glucose tolerance tests in both PTG and control cohorts. CONCLUSIONS: Care should be taken when assessing glucagon levels in the presence of elevated plasma levels of other proglucagon products. Substantial elevation of GLP-1 and insulin responses after PTG likely contribute to the observed hypoglycemia, and mirror similar hormone levels and complications observed in bariatric weight loss patients.

Roux-en-Y Gastric Bypass Surgery in the Management of Familial Partial Lipodystrophy Type 1.

Context: Familial partial lipodystrophy type 1 (FPLD1) is an extreme form of central adiposity, with peripheral lipodystrophy associated with severe manifestations of the metabolic syndrome, often poorly responsive to standard therapeutic approaches. Body mass index in FPLD1 varies but, in many cases, is below the level at which metabolic surgery is usually considered as a therapeutic option. Design: We detailed the metabolic response to gastric bypass surgery of three patients with FPLD1, refractory to medical therapy. Results: Roux-en-Y gastric bypass (RYGB) was associated with weight loss and substantial improvements in glycemic control and insulin sensitivity. All three patients were able to stop using insulin. Glucose tolerance testing in one patient demonstrated an increase in L-cell-derived gut hormone responses postoperatively. Conclusion: RYGB surgery substantially improved glycemic control in three patients with FPLD1, two of whom had body mass indices below 30 kg/m2. RYGB should be considered in patients with partial lipodystrophy and refractory metabolic disease.

The Neutrophil-to-Lymphocyte Ratio as a Marker of Lymph Node Status in Patients with Rectal Cancer.

AIM: The aim of the study was to investigate the role of the neutrophil-to-lymphocyte ratio (NLR) as a prognostic marker of rectal cancers. METHODS: We undertook a retrospective review of patients with rectal cancer. Pre-treatment NLR was assessed for association and predictive values against clinicopathological staging and post-treatment outcomes. RESULTS: A total of 140/180 cases were included in the final analysis [male:female 2:1; mean age 68 years (interquartile range 58-75)]. The pre-operative mean NLR was 5.4 ± 6.8. There was a strong positive correlation between NLR and C-reactive protein (Spearman's rho 64.3%, p < 0.001). A high NLR was associated with a positive nodal status on MRI (5.2 vs. 3.8, p = 0.03) and histopathological (4.8 vs. 3.8, p = 0.02) assessment. The NLR showed an average value for predicting MRI and pathological nodal status on receiver operating characteristic analysis [area under the curve = 0.72 (95% CI = 0.54-0.91), p = 0.031 and area under the curve = 0.64 (95% CI = 0.52-0.077), p = 0.021, respectively]. On multivariate analysis, the total lymph node retrieved at operation was the best predictor of pathological nodal involvement; NLR did not show any predictive value. Patients with an NLR >4 showed reduced recurrence-free (60 vs. 86 months, p = 0.52) and overall survival (57 vs. 84 months, p = 0.40) without statistical significance. CONCLUSION: Raised pre-treatment NLR may indicate nodal involvement in patients with rectal cancer.

Surgery and Obstetric Care are Highly Cost-Effective Interventions in a Sub-Saharan African District Hospital: A Three-Month Single-Institution Study of Surgical Costs and Outcomes.

BACKGROUND: The Lancet recently sponsored a commission examining the role of surgery in global health. There is a paucity of published information on the cost-effectiveness of surgery in low- and middle-income countries, a key metric in the prioritisation of limited resources. METHODS: All patients undergoing emergency laparotomy, elective and emergency inguinal hernia repair, elective and emergency caesarean section, amputation, fracture manipulation, or fracture fixation over a 3 months period in a single district African hospital were assessed. World Health Organisation global burden of disease (GBD) methodology was used to calculate the disability-adjusted life years (DALYs) saved for each patient (using global and local life expectancy). Fully loaded costs were calculated for each patient's care and providing the overall surgical service. Cost-effectiveness was calculated in year 2012 US$ per DALY saved for each procedure and overall. RESULTS: A total of 428 patients were included, with an overall cost-effectiveness of $10.70 per DALY averted. The cost-effectiveness of individual procedures (global life expectancy) was: Amputation­$17.66; Emergency caesarean section­$7.42; Elective caesarean section­$20.50; Emergency laparotomy­$8.62; Elective hernia repair­$15.26; Emergency hernia repair­$4.36; Fracture/dislocation reduction­$69.03; Fracture/dislocation fixation­$225.89. CONCLUSIONS: Surgery is a highly cost-effective healthcare measure in the setting of an African district hospital. The presented outcomes demonstrate that surgery is on a par with better-recognised and funded interventions such as HIV anti-retrovirals, malaria prevention and diarrhoea treatment. There are recognised limitations with the GBD methodology used here; however, this remains the best way to investigate the cost-effectiveness of health interventions. This study provides useful information on an, at present, under-studied field.

The Baux score is dead. Long live the Baux score: a 27-year retrospective cohort study of mortality at a regional burns service.

BACKGROUND: To assess trends in mortality after burn injuries treated in a regional specialist burns service between 1982 and 2008. METHODS: Patient and burn-specific information and mortality were collated from written admission ledgers and the hospital coding department for 11,109 patients. The data set was divided into age cohorts (0-14, 15-44, 45-64, and >65 years) and time cohorts (1982-1991, 1992-2000, and 2000-2008). Lethal area 50 (LA50) was calculated by logistic regression and probit analysis. Mortality was related to the Baux score (age + total % burned surface area) by logistic regression. RESULTS: In the time period 2000 to 2008, the LA50 values with approximate 95% confidence intervals (CIs) were 100% (CI, 85.5-100%) in the 0 to 14 cohort (LA10, 78.3%; CI, 64.1-92.5%), 76.4% (CI, 69.1-83.8%) in the 15 to 44 cohort, 58.6% (CI, 50.8-66.5%) in the 45 to 64 cohort, and 30.8% (CI, 24.7-36.9%) in the >65 cohort. The point of futility (the Baux Score at which predicted mortality is 100%) was 160 and the Baux50 (the Baux score at which predicted mortality is 50%) was 109.6 (CI, 105.9-113.4) in the 2000 to 2008 cohort. CONCLUSIONS: Mortality is markedly improved over earlier data from this study and other historical series and compares favorably with outcomes published from the US National Burn Repository. The Baux Score continues to provide an indication of the risk of mortality. Survival after major burn injury is increasingly common, and decisions by nonspecialist about initial triage, management, and futility of care should be made after consultation with a specialist burn service.

Real-time outcome monitoring following oesophagectomy using cumulative sum techniques.

AIM: To examine the feasibility of prospective, real-time outcome monitoring in a United Kingdom oesophago-gastric cancer surgery unit. METHODS: The first 100 hybrid (laparoscopic abdominal phase, open thoracic phase) Ivor-Lewis oesophagectomies performed by a United Kingdom oesophago-gastric cancer surgery unit were assessed retrospectively using cumulative sum (CUSUM) techniques. The monitored outcome was 30-d post-operative mortality, with the accepted mortality risk defined as 5%. A variable life adjusted display (VLAD) was constructed by plotting a graph of cumulative mortality minus cumulative mortality risk on the y axis vs sequential case number on the x axis. This was modified to a zeroed VLAD by preventing the plot from crossing the y = 0 axis - essentially creating two plots, one examining trends where cumulative mortality was higher than mortality risk (i.e., worse than expected outcomes) where y > 0, and vice versa. Alert lines were set at y = ± 2. At any point where a plot breaches an alert line, it is felt that the 30-d post-operative mortality rate has deviated significantly from that expected and an internal review should be performed. RESULTS: One hundred cases were assessed, with a mean age of 66.4 years, mean T stage of 2.1, and mean N stage of 0.48. Three cases were commenced using a laparoscopic technique and converted to open surgery due to technical factors. Median length of inpatient stay was 15 d. The crude 30 d mortality was 5% and the incidence of clinically significant anastomotic leak was 6%. The VLAD demonstrated a plot of cumulative mortality minus cumulative mortality risk (i.e., 5% per case) which remained in the range -1.4 to +0.5 excess mortalities. With the alert set at two greater or fewer than predicted mortalities, this method does not approach the point of triggering internal review. It is however arguable that a run of performance that is better than expected, causing the plot to be well below y = 0, would mask a subsequent run of poor performance by requiring a rise of greater than two excess mortalities to trigger the alert line. The zeroed VLAD removes this problem by preventing the plot that is examining above expected mortality from passing below y = 0, and vice versa. In this study period, no audit triggers were reached. It is therefore possible to independently assess runs of good, or poor performance and so target internal audit to the appropriate series of cases. It is important to note this technique allows targeted internal review, in response to both above and below average outcomes. This study has demonstrated the feasibility of prospective outcome monitoring using the above techniques, actual real-time implementation has the potential to pick up and reinforce good practices when performance is better than predicted, and provide an early warning system for when performance falls below that predicted. Further development is possible, including more patient specific risk adjustment using the oesophago-gastric surgery physiological and operative severity score for the enumeration of mortality and morbidity score. CONCLUSION: CUSUM techniques provide a potential method of prospective, real-time outcome monitoring in oesophageal cancer surgery.