Identifying mismatch repair-deficient colon cancer: near-perfect concordance between immunohistochemistry and microsatellite instability testing in a large, population-based series.

AIMS: Establishing the mismatch repair (MMR) status of colorectal cancers is important to enable the detection of underlying Lynch syndrome and inform prognosis and therapy. Current testing typically involves either polymerase chain reaction (PCR)-based microsatellite instability (MSI) testing or MMR protein immunohistochemistry (IHC). The aim of this study was to compare these two approaches in a large, population-based cohort of stage 2 and 3 colon cancer cases in Northern Ireland. METHODS AND RESULTS: The study used the Promega pentaplex assay to determine MSI status and a four-antibody MMR IHC panel. IHC was applied to tumour tissue microarrays with triplicate tumour sampling, and assessed manually. Of 593 cases with available MSI and MMR IHC results, 136 (22.9%) were MSI-high (MSI-H) and 135 (22.8%) showed abnormal MMR IHC. Concordance was extremely high, with 97.1% of MSI-H cases showing abnormal MMR IHC, and 97.8% of cases with abnormal IHC showing MSI-H status. Under-representation of tumour epithelial cells in samples from heavily inflamed tumours resulted in misclassification of several cases with abnormal MMR IHC as microsatellite-stable. MMR IHC revealed rare cases with unusual patterns of MMR protein expression, unusual combinations of expression loss, or secondary clonal loss of expression, as further illustrated by repeat immunostaining on whole tissue sections. CONCLUSIONS: MSI PCR testing and MMR IHC can be considered to be equally proficient tests for establishing MMR/MSI status, when there is awareness of the potential pitfalls of either method. The choice of methodology may depend on available services and expertise.

Fusobacterium nucleatum and oral cancer: a critical review.

There is a growing level of interest in the potential role inflammation has on the initiation and progression of malignancy. Notable examples include Helicobacter pylori-mediated inflammation in gastric cancer and more recently Fusobacterium nucleatum-mediated inflammation in colorectal cancer. Fusobacterium nucleatum is a Gram-negative anaerobic bacterium that was first isolated from the oral cavity and identified as a periodontal pathogen. Biofilms on oral squamous cell carcinomas are enriched with anaerobic periodontal pathogens, including F. nucleatum, which has prompted hypotheses that this bacterium could contribute to oral cancer development. Recent studies have demonstrated that F. nucleatum can promote cancer by several mechanisms; activation of cell proliferation, promotion of cellular invasion, induction of chronic inflammation and immune evasion. This review provides an update on the association between F. nucleatum and oral carcinogenesis, and provides insights into the possible mechanisms underlying it.

Immune status is prognostic for poor survival in colorectal cancer patients and is associated with tumour hypoxia.

BACKGROUND: Immunohistochemical quantification of the immune response is prognostic for colorectal cancer (CRC). Here, we evaluate the suitability of alternative immune classifiers on prognosis and assess whether they relate to biological features amenable to targeted therapy. METHODS: Overall survival by immune (CD3, CD4, CD8, CD20 and FOXP3) and immune-checkpoint (ICOS, IDO-1 and PD-L1) biomarkers in independent CRC cohorts was evaluated. Matched mutational and transcriptomic data were interrogated to identify associated biology. RESULTS: Determination of immune-cold tumours by combined low-density cell counts of CD3, CD4 and CD8 immunohistochemistry constituted the best prognosticator across stage II-IV CRC, particularly in patients with stage IV disease (HR 1.98 [95% CI: 1.47-2.67]). These immune-cold CRCs were associated with tumour hypoxia, confirmed using CAIX immunohistochemistry (P = 0.0009), which may mediate disease progression through common biology (KRAS mutations, CRIS-B subtype and SPP1 mRNA overexpression). CONCLUSIONS: Given the significantly poorer survival of immune-cold CRC patients, these data illustrate that assessment of CD4-expressing cells complements low CD3 and CD8 immunohistochemical quantification in the tumour bulk, potentially facilitating immunophenotyping of patient biopsies to predict prognosis. In addition, we found immune-cold CRCs to associate with a difficult-to-treat, poor prognosis hypoxia signature, indicating that these patients may benefit from hypoxia-targeting clinical trials.

Alcohol intake, tobacco smoking, and esophageal adenocarcinoma survival: a molecular pathology epidemiology cohort study.

PURPOSE: To investigate the association between cigarette smoking, alcohol consumption, and esophageal adenocarcinoma survival, including stratified analysis by selected prognostic biomarkers. METHODS: A population-representative sample of 130 esophageal adenocarcinoma patients (n = 130) treated at the Northern Ireland Cancer Centre between 2004 and 2012. Cox proportional hazards models were applied to evaluate associations between smoking status, alcohol intake, and survival. Secondary analyses investigated these associations across categories of p53, HER2, CD8, and GLUT-1 biomarker expression. RESULTS: In esophageal adenocarcinoma patients, there was a significantly increased risk of cancer-specific mortality in ever, compared to never, alcohol drinkers in unadjusted (HR 1.96 95% CI 1.13-3.38) but not adjusted (HR 1.70 95% CI 0.95-3.04) analysis. This increased risk of death observed for alcohol consumers was more evident in patients with normal p53 expression, GLUT-1 positive or CD-8 positive tumors. There were no significant associations between survival and smoking status in esophageal adenocarcinoma patients. CONCLUSIONS: In esophageal adenocarcinoma patients, cigarette smoking or alcohol consumption was not associated with a significant difference in survival in comparison with never smokers and never drinkers in fully adjusted analysis. However, in some biomarker-selected subgroups, ever-alcohol consumption was associated with a worsened survival in comparison with never drinkers. Larger studies are needed to investigate these findings, as these lifestyle habits may not only be linked to cancer risk but also cancer survival.

Spontaneous regression of ALK fusion protein-positive non-small cell lung carcinoma: a case report and review of the literature.

BACKGROUND: ALK-rearrangement is observed in < 5% non-small cell lung cancer (NSCLC) cases and prior to the advent of oral tyrosine kinase inhibitors, the natural history of oncogenic NSCLC was typically poor. Literature relating to regression of treatment-naïve NSCLC is limited, and regression without treatment has not been noted in the ALK-rearranged sub-population. CASE PRESENTATION: A 76 year old 'never smoker' female with an ALK-rearranged left upper lobe T2 N0 NSCLC experienced a stroke following elective DC cardioversion for new atrial fibrillation. Following a good recovery, updated imaging demonstrated complete regression of the left upper lobe lesion and a reduction of the previously documented mediastinal lymph node. Remaining atelectasis was non-avid on repeat PET-CT imaging, 8 months from the baseline PET-CT. When the patient developed new symptoms 6 months later a further PET-CT demonstrated FDG-avid local recurrence. She completed 55 Gy in 20 fractions but at 18 months post-radiotherapy there was radiological progression in the lungs with new pulmonary metastases and effusion and new bone metastases. Owing to poor performance status, she was not considered fit for targeted therapy and died 5 months later. CONCLUSION: All reported cases of spontaneous regression in lung cancer have been collated within. Documented precipitants of spontaneous regression across tumour types include biopsy and immune reconstitution; stroke has not been reported previously. The favourable response achieved with radical radiotherapy alone in this unusual case of indolent oncogenic NSCLC reinforces the applicability of radiotherapy in locally advanced ALK-rearranged tumours, in cases not behaving aggressively. As a common embolic event affecting the neurological and pulmonary vasculature is less likely, an immune-mediated mechanism may underpin the phenomenon described in this patient, implying that hitherto unharnessed principles of immuno-oncology may have relevance in oncogenic NSCLC. Alternatively, high electrical voltage applied percutaneously adjacent to the tumour during cardioversion in this patient may have induced local tumour cell lethality.

Immune activation by DNA damage predicts response to chemotherapy and survival in oesophageal adenocarcinoma.

OBJECTIVE: Current strategies to guide selection of neoadjuvant therapy in oesophageal adenocarcinoma (OAC) are inadequate. We assessed the ability of a DNA damage immune response (DDIR) assay to predict response following neoadjuvant chemotherapy in OAC. DESIGN: Transcriptional profiling of 273 formalin-fixed paraffin-embedded prechemotherapy endoscopic OAC biopsies was performed. All patients were treated with platinum-based neoadjuvant chemotherapy and resection between 2003 and 2014 at four centres in the Oesophageal Cancer Clinical and Molecular Stratification consortium. CD8 and programmed death ligand 1 (PD-L1) immunohistochemical staining was assessed in matched resection specimens from 126 cases. Kaplan-Meier and Cox proportional hazards regression analysis were applied according to DDIR status for recurrence-free survival (RFS) and overall survival (OS). RESULTS: A total of 66 OAC samples (24%) were DDIR positive with the remaining 207 samples (76%) being DDIR negative. DDIR assay positivity was associated with improved RFS (HR: 0.61; 95% CI 0.38 to 0.98; p=0.042) and OS (HR: 0.52; 95% CI 0.31 to 0.88; p=0.015) following multivariate analysis. DDIR-positive patients had a higher pathological response rate (p=0.033), lower nodal burden (p=0.026) and reduced circumferential margin involvement (p=0.007). No difference in OS was observed according to DDIR status in an independent surgery-alone dataset.DDIR-positive OAC tumours were also associated with the presence of CD8+ lymphocytes (intratumoural: p<0.001; stromal: p=0.026) as well as PD-L1 expression (intratumoural: p=0.047; stromal: p=0.025). CONCLUSION: The DDIR assay is strongly predictive of benefit from DNA-damaging neoadjuvant chemotherapy followed by surgical resection and is associated with a proinflammatory microenvironment in OAC.

Recommendations for determining HPV status in patients with oropharyngeal cancers under TNM8 guidelines: a two-tier approach.

BACKGROUND: TNM8 staging for oropharyngeal squamous cell carcinomas (OPSCC) surrogates p16 immunohistochemistry for HPV testing. Patients with p16+ OPSCC may lack HPV aetiology. Here, we evaluate the suitability of TNM8 staging for guiding prognosis in such patients. METHODS: HPV status was ascertained using p16 immunohistochemistry and high-risk HPV RNA and DNA in situ hybridisation. Survival by stage in a cohort of OPSCC patients was evaluated using TNM7/TNM8 staging. Survival of p16+/HPV- patients was compared to p16 status. RESULTS: TNM8 staging was found to improve on TNM7 (log rank p = 0·0190 for TNM8 compared with p = 0·0530 for TNM7) in p16+ patients. Patients who tested p16+ but were HPV- (n = 20) had significantly reduced five-year survival (33%) compared to p16+ patients (77%) but not p16- patients (35%). Cancer stage was reduced in 95% of p16+/HPV- patients despite having a mortality rate twice (HR 2.66 [95% CI: 1.37-5.15]) that of p16+/HPV+ patients under new TNM8 staging criteria. CONCLUSION: Given the significantly poorer survival of p16+/HPV- OPSCCs, these data provide compelling evidence for use of an HPV-specific test for staging classification. This has particular relevance in light of potential treatment de-escalation that could expose these patients to inappropriately reduced treatment intensity as treatment algorithms evolve.

Embracing an integromic approach to tissue biomarker research in cancer: Perspectives and lessons learned.

Modern approaches to biomedical research and diagnostics targeted towards precision medicine are generating 'big data' across a range of high-throughput experimental and analytical platforms. Integrative analysis of this rich clinical, pathological, molecular and imaging data represents one of the greatest bottlenecks in biomarker discovery research in cancer and other diseases. Following on from the publication of our successful framework for multimodal data amalgamation and integrative analysis, Pathology Integromics in Cancer (PICan), this article will explore the essential elements of assembling an integromics framework from a more detailed perspective. PICan, built around a relational database storing curated multimodal data, is the research tool sitting at the heart of our interdisciplinary efforts to streamline biomarker discovery and validation. While recognizing that every institution has a unique set of priorities and challenges, we will use our experiences with PICan as a case study and starting point, rationalizing the design choices we made within the context of our local infrastructure and specific needs, but also highlighting alternative approaches that may better suit other programmes of research and discovery. Along the way, we stress that integromics is not just a set of tools, but rather a cohesive paradigm for how modern bioinformatics can be enhanced. Successful implementation of an integromics framework is a collaborative team effort that is built with an eye to the future and greatly accelerates the processes of biomarker discovery, validation and translation into clinical practice.

Integrated tumor identification and automated scoring minimizes pathologist involvement and provides new insights to key biomarkers in breast cancer.

Digital image analysis (DIA) is becoming central to the quantitative evaluation of tissue biomarkers for discovery, diagnosis and therapeutic selection for the delivery of precision medicine. In this study, automated DIA using a new purpose-built software platform (QuPath) is applied to a cohort of 293 breast cancer patients to score five biomarkers in tissue microarrays (TMAs): ER, PR, HER2, Ki67 and p53. This software is able to measure IHC expression following fully automated tumor recognition in the same immunohistochemical (IHC)-stained tissue section, as part of a rapid workflow to ensure objectivity and accelerate biomarker analysis. The digital scores produced by QuPath were compared with manual scores by a pathologist and shown to have a good level of concordance in all cases (Cohen's κ>0.6), and almost perfect agreement for the clinically relevant biomarkers ER, PR and HER2 (κ>0.86). To assess prognostic value, cutoff thresholds could be applied to both manual and automated scores using the QuPath software, and survival analysis performed for 5-year overall survival. DIA was shown to be capable of replicating the statistically significant stratification of patients achieved using manual scoring across all biomarkers (P<0.01, log-rank test). Furthermore, the image analysis scores were shown to consistently lead to statistical significance across a wide range of potential cutoff thresholds, indicating the robustness of the method, and identify sub-populations of cases exhibiting different expression patterns within the p53 and Ki67 data sets that warrant further investigation. These findings have demonstrated QuPath's suitability for fast, reproducible, high-throughput TMA analysis across a range of important biomarkers. This was achieved using our tumor recognition algorithms for IHC-stained sections, trained interactively without the need for any additional tumor recognition markers, for example, cytokeratin, to obtain greater insight into the relationship between biomarker expression and clinical outcome applicable to a range of cancer types.

Validation of the systematic scoring of immunohistochemically stained tumour tissue microarrays using QuPath digital image analysis.

AIMS: Output from biomarker studies involving immunohistochemistry applied to tissue microarrays (TMA) is limited by the lack of an efficient and reproducible scoring methodology. In this study, we examine the functionality and reproducibility of biomarker scoring using the new, open-source, digital image analysis software, QuPath. METHODS AND RESULTS: Three different reviewers, with varying experience of digital pathology and image analysis, applied an agreed QuPath scoring methodology to CD3 and p53 immunohistochemically stained TMAs from a colon cancer cohort (n = 661). Manual assessment was conducted by one reviewer for CD3. Survival analyses were conducted and intra- and interobserver reproducibility assessed. Median raw scores differed significantly between reviewers, but this had little impact on subsequent analyses. Lower CD3 scores were detected in cases who died from colorectal cancer compared to control cases, and this finding was significant for all three reviewers (P-value range = 0.002-0.02). Higher median p53 scores were generated among cases who died from colorectal cancer compared with controls (P-value range = 0.04-0.12). The ability to dichomotise cases into high versus low expression of CD3 and p53 showed excellent agreement between all three reviewers (kappa score range = 0.82-0.93). All three reviewers produced dichotomised expression scores that resulted in very similar hazard ratios for colorectal cancer-specific survival for each biomarker. Results from manual and QuPath methods of CD3 scoring were comparable, but QuPath scoring revealed stronger prognostic stratification. CONCLUSIONS: Scoring of immunohistochemically stained tumour TMAs using QuPath is functional and reproducible, even among users of limited experience of digital pathology images, and more accurate than manual scoring.

Tissue-based next generation sequencing: application in a universal healthcare system.

In the context of solid tumours, the evolution of cancer therapies to more targeted and nuanced approaches has led to the impetus for personalised medicine. The targets for these therapies are largely based on the driving genetic mutations of the tumours. To track these multiple driving mutations the use of next generation sequencing (NGS) coupled with a morphomolecular approach to tumours, has the potential to deliver on the promises of personalised medicine. A review of NGS and its application in a universal healthcare (UHC) setting is undertaken as the technology has a wide appeal and utility in diagnostic, clinical trial and research paradigms. Furthermore, we suggest that these can be accommodated with a unified integromic approach. Challenges remain in bringing NGS to routine clinical use and these include validation, handling of the large amounts of information flow and production of a clinically useful report. These challenges are particularly acute in the setting of UHC where tests are not reimbursed and there are finite resources available. It is our opinion that the challenges faced in applying NGS in a UHC setting are surmountable and we outline our approach for its routine application in diagnostic, clinical trial and research paradigms.

Molecular profiling of signet ring cell colorectal cancer provides a strong rationale for genomic targeted and immune checkpoint inhibitor therapies.

BACKGROUND: Signet ring cell colorectal cancer (SRCCa) has a bleak prognosis. Employing molecular pathology techniques we investigated the potential of precision medicine in this disease. METHODS: Using test (n=26) and validation (n=18) cohorts, analysis of mutations, DNA methylation and transcriptome was carried out. Microsatellite instability (MSI) status was established and immunohistochemistry (IHC) was used to test for adaptive immunity (CD3) and the immune checkpoint PDL1. RESULTS: DNA methylation data split the cohorts into hypermethylated (n=18, 41%) and hypomethylated groups (n=26, 59%). The hypermethylated group predominant in the proximal colon was enriched for CpG island methylator phenotype (CIMP), BRAF V600E mutation and MSI (P<0.001). These cases also had a high CD3+ immune infiltrate (P<0.001) and expressed PDL1 (P=0.03 in intra-tumoural lymphoid cells). The hypomethylated group predominant in the distal colon did not show any characteristic molecular features. We also detected a common targetable KIT mutation (c.1621A>C) across both groups. No statistically significant difference in outcome was observed between the two groups. CONCLUSIONS: Our data show that SRCCa phenotype comprises two distinct genotypes. The MSI+/CIMP+/BRAF V600E+/CD3+/PDL1+ hypermethylated genotype is an ideal candidate for immune checkpoint inhibitor therapy. In addition, one fourth of SRCCa cases can potentially be targeted by KIT inhibitors.

Statin use, candidate mevalonate pathway biomarkers, and colon cancer survival in a population-based cohort study.

BACKGROUND: Statin use after colorectal cancer diagnosis may improve survival but evidence from observational studies is conflicting. The anti-cancer effect of statins may be restricted to certain molecular subgroups. In this population-based cohort study, the interaction between p53 and 3-hydroxy-3-methylglutaryl coenzyme-A reductase (HMGCR) expression, KRAS mutations, and the association between statin use and colon cancer survival was assessed. METHODS: The cohort consisted of 740 stage II and III colon cancer patients diagnosed between 2004 and 2008. Statin use was determined through clinical note review. Tissue blocks were retrieved to determine immunohistochemical expression of p53 and HMGCR in tissue microarrays and the presence of KRAS mutations in extracted DNA. Cox proportional hazards models were used to calculate hazard ratios (HRs) and 95% confidence intervals (CIs) for colorectal cancer-specific and overall survival. RESULTS: Statin use was not associated with improved cancer-specific survival in this cohort (HR=0.91, 95% CI 0.64-1.28). Statin use was also not associated with improved survival when the analyses were stratified by tumour p53 (wild-type HR=1.31, 95% CI 0.67-2.56 vs aberrant HR=0.80, 95% CI 0.52-1.24), HMGCR (HMGCR-high HR=0.69, 95% CI 0.40-1.18 vs HMGCR-low HR=1.10, 95% CI 0.66-1.84), and KRAS (wild-type HR=0.73, 95% CI 0.44-1.19 vs mutant HR=1.21, 95% CI 0.70-2.21) status. CONCLUSIONS: Statin use was not associated with improved survival either independently or when stratified by potential mevalonate pathway biomarkers in this population-based cohort of colon cancer patients.

Back to the future: routine morphological assessment of the tumour microenvironment is prognostic in stage II/III colon cancer in a large population-based study.

AIMS: Both morphological and molecular approaches have highlighted the biological and prognostic importance of the tumour microenvironment in colorectal cancer (CRC). Despite this, microscopic assessment of the tumour microenvironment has not been adopted into routine practice. The study aim was to identify those tumour microenvironmental features that are most likely to provide prognostic information and be feasible to use in routine pathology reporting practice. METHODS AND RESULTS: On the basis of existing evidence, we selected specific morphological features relating to peritumoral inflammatory and stromal responses, agreed criteria for scoring, and assessed these in representative haematoxylin and eosin (H&E)-stained whole tumour sections from a population-based cohort of 445 stage II/III colon cancer cases. Moderate/severe peritumoral diffuse lymphoid inflammation and Crohn's disease-like reaction were associated with significantly reduced risks of CRC-specific death [adjusted hazard ratio (HR) 0.48, 95% confidence interval (CI) 0.31-0.76, and HR 0.60, 95% CI 0.42-0.84, respectively]. The presence of >50% tumour stromal percentage, as assessed by global evaluation of tumour area, was associated with a significantly increased risk of CRC-specific death (HR 1.60 95% CI 1.06-2.41). A composite 'fibroinflammatory score' (0-3), combining dichotomized scores of these three features, showed a highly significant association with survival outcomes. Those with a score of ≥2 had an almost 2.5-fold increased risk of CRC-specific death (HR 2.44, 95% CI 1.56-3.81) as compared with those scoring zero. These associations were stronger in microsatellite instability (MSI)-high tumours, potentially identifying a subset of MSI-high colon cancers that lack characteristic morphological features and have an associated worse prognosis. CONCLUSIONS: In summary, reporting on H&E staining of selected microscopic features of the tumour microenvironment, independently or in combination, offers valuable prognostic information in stage II/III colon cancer, and may allow morphological correlation with developing molecular classifications of prognostic and predictive relevance.

Globodontia in the Otodental Syndrome: A Rare Defect of Tooth Morphology Occurring with Hearing Loss in an Eight-Year-Old.

Otodental syndrome is a hereditary disorder comprising globodontia and sensorineural hearing loss. Globodontia is characterized by distinctively bulbous, enlarged crowns of molar and primary canine teeth. Anomalies including taurodontism and hypodontia also occur. We report on the dental treatment and multidisciplinary management of an eight-year-old girl with this rare condition. Referral to Clinical Genetics and Oral Pathology was instrumental in establishing a diagnosis of otodental syndrome for this young patient and her mother, who had similar dental defects. CPD/Clinical Relevance: To increase awareness among practitioners of this rare dental disorder and highlight the need for multidisciplinary management of such cases.

The prognostic significance of the aberrant extremes of p53 immunophenotypes in breast cancer.

AIMS: The utility of p53 as a prognostic assay has been elusive. The aims of this study were to describe a novel, reproducible scoring system and assess the relationship between differential p53 immunohistochemistry (IHC) expression patterns, TP53 mutation status and patient outcomes in breast cancer. METHODS AND RESULTS: Tissue microarrays were used to study p53 IHC expression patterns: expression was defined as extreme positive (EP), extreme negative (EN), and non-extreme (NE; intermediate patterns). Overall survival (OS) was used to define patient outcome. A representative subgroup (n = 30) showing the various p53 immunophenotypes was analysed for TP53 hotspot mutation status (exons 4-9). Extreme expression of any type occurred in 176 of 288 (61%) cases. As compared with NE expression, EP expression was significantly associated (P = 0.039) with poorer OS. In addition, as compared with NE expression, EN expression was associated (P = 0.059) with poorer OS. Combining cases showing either EP or EN expression better predicted OS than either pattern alone (P = 0.028). This combination immunophenotype was significant in univariate but not multivariate analysis. In subgroup analysis, six substitution exon mutations were detected, all corresponding to extreme IHC phenotypes. Five missense mutations corresponded to EP staining, and the nonsense mutation corresponded to EN staining. No mutations were detected in the NE group. CONCLUSIONS: Patients with extreme p53 IHC expression have a worse OS than those with NE expression. Accounting for EN as well as EP expression improves the prognostic impact. Extreme expression positively correlates with nodal stage and histological grade, and negatively with hormone receptor status. Extreme expression may relate to specific mutational status.

Assessment of Ki-67 and mitoses in pituitary neuroendocrine tumours-Consistency counts.

Pituitary neuroendocrine tumour Ki-67 proliferation index varies according to the number of tumour cells assessed. Consistent Ki-67 scoring approaches and re-evaluation of the recommended Ki-67 3% cut-off are required to clarify controversies in pituitary neuroendocrine tumour Ki-67 proliferation index assessment.

Insights on epidemiology, morbidity and mortality of Cushing's disease in Northern Ireland.

Cushing's disease is a rare condition that occurs due to an adrenocorticotrophin-producing corticotrophinoma arising from the pituitary gland. The consequent hypercortisolaemia results in multisystem morbidity and mortality. This study aims to report incidence, clinicopathological characteristics, remission outcomes and mortality in a regional pituitary neurosurgical cohort of patients diagnosed with Cushing's disease in Northern Ireland (NI) from 2000 to 2019. Clinical, biochemical and radiological data from a cohort of patients operated for Cushing's disease were retrospectively collected and analysed. Fifty-three patients were identified, resulting in an estimated annual incidence of Cushing's disease of 1.39-1.57 per million population per year. Females accounted for 72% (38/53) of the cohort. The majority (74%, 39/53) of corticotrophinomas were microadenomas and in 44% (17/39) of these no tumour was identified on preoperative magnetic resonance imaging. Histopathological characterisation was similarly difficult, with no tumour being identified in the histopathological specimen in 40% (21/53) of cases. Immediate postoperative remission rates were 53% and 66% when considering serum morning cortisol cut-offs of ≤ 50 nmol/L (1.8 µg/dL) and ≤ 138 nmol/L (5 µg/dL), respectively, in the week following pituitary surgery. Approximately 70% (37/53) of patients achieved longer-term remission with a single pituitary surgery. Three patients had recurrent disease. Patients with Cushing's disease had a significantly higher mortality rate compared to the NI general population (standardised mortality ratio 8.10, 95% CI 3.3-16.7, P < 0.001). Annual incidence of Cushing's disease in NI is consistent with other Northern European cohorts. Functioning corticotrophinomas are a clinically, radiologically and histopathologically elusive disease with increased mortality compared to the general population.