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Introduction Microsatellite instability (MSI) predict response to anti-PD1 immunotherapy in colorectal cancer (CRC). CRCs with MSI have higher infiltration of immune cells related to a better survival. Elevated Microsatellite Alterations at Tetranucleotides (EMAST) is a form of MSI but its association with PD-L1 expression and immune-cell infiltration is not known. Methods A consecutive, observational cohort of patients undergoing surgery for CRC. EMAST and clinicopathological characteristics were investigated against PD-L1, as well as CD3 and CD8 expression in the invasive margin or tumour centre (Immunoscore). Difference in survival between groups was assessed by log rank test. Results A total of 149 stage I–III CRCs patients, with a median follow up of 60.1 months. Patients with PD-L1+ tumours (7%) were older (median 79 vs 71 years, p = 0.045) and had EMAST+ cancers (OR 10.7, 95% CI 2.2–51.4, p = 0.001). Recurrence-free survival was longer in cancers with PD-L1+ immune cells (HR 0.35, 95% CI 0.16–0.76, p = 0.008, independent of EMAST) and high Immunoscore (HR 0.10, 95% CI 0.01–0.72, p = 0.022). Patients expressing PD-L1 in immune cells had longer disease-specific survival (HR 0.28, 95% CI 0.10–0.77, p = 0.014). Conclusions Higher Immunoscore (CD3/CD8 cells) and expression of tumour PD-L1 is found in CRCs with EMAST. Lymphocytic infiltrate and peritumoral PD-L1 expression have prognostic value in CRC.

Elevated microsatellite alterations at selected tetranucleotides (EMAST) is a poorly investigated form of microsatellite instability (MSI) in colorectal cancer (CRC). The aim of this study was to investigate the clinicopathological features of EMAST in CRC and its relation to outcome. A population-based, consecutive cohort of surgically treated stage I–III CRC patients investigated for high-frequency MSI (MSI-H) and EMAST. Clinicopathological differences were reported as odds ratios (OR) and survival was presented as hazard ratios (HR) with 95% confidence intervals (CIs). Of 161 patients included, 25% were aged > 79 years. There was a large overlap in the prevalence of EMAST (31.7%) and MSI-H (27.3%) [82.4% of EMAST were also MSI-H]. EMAST had the highest prevalence in the proximal colon (OR 15.9, 95% CI 5.6–45.1; p

Background: Novel technology has enabled researchers to better characterize pancreatic cancers at the molecular level. We wanted to explore some of the emerging discoveries, such as molecular subclassification, use of liquid biopsy and use of organoids in cancer assessment. Methods: A literature review with a search specific to the topic, with recent reviews in major journals and a focus on the last 5 years (until December 2018), was done. Results: Pancreatic ductal adenocarcinoma (PDAC) may now be classified into clinical subgroups based on the predominant genomic profiles, but consensus on one classification system is lacking. Several subtypes have been suggested, including categories such as basal-like, stroma-activated, desmoplastic, pure classical and immune classical types. Further refinement may translate into clinically meaningful groups for therapeutic or prognostic purposes. Liquid biopsies (by means of circulating cancer cells, cell-free DNA, exosomes or other constituents of cancer cells in blood) may aid in earlier diagnosis, define prognostic groups and even predict therapy response and resistance. Organoids are increasingly used for the opportunity to investigate druggable and effective targets ex vivo and should facilitate personalized and precise, targeted therapy in the near future. While immunotherapy has not yet proved to be effective, a better understanding of molecular subgroups and specific immune profiles may help identify candidates for this approach in a more selective approach. Conclusion: Novel molecular techniques have the potential to accelerate the road to improved outcomes in patients with pancreatic cancer. publishedVersion

Background In colon cancer, the location and density of tumor-infiltrating lymphocytes (TILs) can classify patients into low and high-risk groups for prognostication. While a commercially available ‘Immunoscore®’ exists, the incurred expenses and copyrights may prevent universal use. The aim of this study was to develop a robust and objective quantification method of TILs in colon cancer. Methods A consecutive, unselected series of specimens from patients with colon cancer were available for immunohistochemistry and assessment of TILs by automated digital pathology. CD3 + and CD8 + cells at the invasive margin and in tumor center were assessed on consecutive sections using automated digital pathology and image analysis software (Visiopharm®). An algorithm template for whole slide assessment, generated cell counts per square millimeters (cells/mm2), from which the immune score was calculated using distribution volumes. Furthermore, immune score was compared with clinical and histopathological characteristics to confirm its relevance. Results Based on the quantified TILs numbers by digital image analyses, patients were classified into low (n = 83, 69.7%), intermediate (n = 14, 11.8%) and high (n = 22, 18.5%) immune score groups. High immune score was associated with stage I–II tumors (p = 0.017) and a higher prevalence of microsatellite instable (MSI) tumors (p = 0.030). MSI tumors had a significantly higher numbers of CD3 + TILs in the invasive margin and CD8 + TILs in both tumor center and invasive margin, compared to microsatellite stable (MSS) tumors. Conclusion A digital template to quantify an easy-to-use immune score corresponds with clinicopathological features and MSI in colon cancer.

INTRODUCTION: EMAST is a poorly understood form of microsatellite instability (MSI) in colorectal cancer (CRC) for which loss of MSH3 has been proposed as the underlying mechanism, based on experimental studies. We aimed to evaluate whether MSH3 loss is associated with EMAST in CRC. METHODS: A consecutive cohort of patients with stage I-III CRC. Digital image analysis using heatmap-derived hot spots investigated MSH3 expression by immunohistochemistry. Fragment analysis of multiplex PCR was used to assess MSI and EMAST, and results cross-examined with MSH3 protein expression. RESULTS: Of 152 patients, EMAST was found in 50 (33%) and exclusively in the colon. Most EMAST-positive cancers had instability at all 5 markers, and EMAST overlapped with MSI-H in 42/50 cases (84%). The most frequently altered tetranucleotide markers were D8S321 (38.2% of tumors) and D20S82 (34.4%). Subjective evaluation of MSH3 expression by IHC in tumor found ≤10% negative tumor cells in all samples, most being ≤5% negative. Digital analysis improved the detection but showed a similar spread of MSH3 loss (range 0.1–15.7%, mean 2.2%). Hotspot MSH3 negativity ranged between 0.1 to 95.0%, (mean 8.6%) with significant correlation with the whole slide analysis (Spearman's rho = 0.677 P < .001). Loss of MSH3 expression did not correlate with EMAST. CONCLUSIONS: In a well-defined cohort of patients with CRC, loss of MSH3 was not associated with EMAST. Further investigation into the mechanisms leading to EMAST in CRC is needed. publishedVersion

The tumor-node-metastasis (TNM) system fails to accurately predict disease recurrence in a considerable number of patients. Although node-negative (stage II) colon cancer is considered to have an overall good prognosis, the 5-year cancer-specific survival is reported at 81–83% in patients who did not have adjuvant chemotherapy. Thus, reliance on node status alone has led to undertreatment in a subgroup of stage II patients with an unfavorable prognosis. The search for new and better prognosticators in stage II colon cancer has suggested several proposed biomarkers of better prognostication and prediction. However, few such biomarkers have reached widespread clinical utility. For the clinician swimming in the sea of emerging biomarkers, it may be hard to recognize the true floating aid from the surrounding debris in the search for more precise decision-making. Proposed markers include microsatellite instability (MSI) and KRAS and BRAF mutations, but a number of gene panels and consensus molecular subtypes are proposed for clinical prediction and prognostication as well. Although several studies suggest such biomarkers or panels to have a prognostic role in subgroups of patients, a number of studies are reported in heterogeneous groups with in part discordant findings, which again distorts the predictive and prognostic ability of each marker. Lack of homogeneous cohorts, underpowered studies in strict subgroups and challenges in analytical and clinical validity may hamper the progress toward widespread clinical utility. The harvest of prognostic biomarkers in colon cancer has yielded a huge number of candidates for which it is now time to separate the wheat from the chaff.

Background: More accurate predictive and prognostic biomarkers for patients with colorectal cancer (CRC) primaries or colorectal liver metastasis (CLM) are needed. Outside clinical trials, the translational integration of emerging pathways and novel techniques should facilitate exploration of biomarkers for improved staging and prognosis. Methods: An observational study exploring predictive and prognostic biomarkers in a population-based, consecutive cohort of surgically treated colorectal cancers and resected colorectal liver metastases. Long-term outcomes will be cancer-specific survival, recurrence-free survival and overall survival at 5 years from diagnosis. Beyond routine clinicopathological and anthropometric characteristics and laboratory and biochemistry results, the project allows for additional blood samples and fresh-frozen tumour and normal tissue for investigation of circulating tumour cells (CTCs) and novel biomarkers (e.g. immune cells, microRNAs etc.). Tumour specimens will be investigated by immunohistochemistry in full slides. Extracted DNA/RNA will be analysed for genomic markers using specific PCR techniques and next-generation sequencing (NGS) panels. Flow cytometry will be used to characterise biomarkers in blood. Collaboration is open and welcomed, with particular interest in mutual opportunities for validation studies. Status and perspectives: The project is ongoing and recruiting at an expected rate of 120–150 patients per year, since January 2013. A project on circulating tumour cells (CTCs) has commenced, with analysis being prepared. Investigating molecular classes beyond the TNM staging is under way, including characteristics of microsatellite instability (MSI) and elevated microsatellite alterations in selected tetranucleotides (EMAST). Hot spot panels for known mutations in CRC are being investigated using NGS. Immune-cell characteristics are being performed by IHC and flow cytometry in tumour and peripheral blood samples. The project has ethical approval (REK Helse Vest, #2012/742), is financially supported with a Ph.D.-Grant (EMAST project; Folke Hermansen Cancer Fund) and a CTC-project (Norwegian Research Council; O. Nordgård). The ACROBATICC clinical and molecular biobank repository will serve as a long-term source for novel exploratory analysis and invite collaborators for mutual validation of promising biomarker results. The project aims to generate results that can help better discern prognostic groups in stage II/III cancers; explore prognostic and predictive biomarkers, and help detail the biology of colorectal liver metastasis for better patient selection and tailored treatment. The project is registered at http://www.ClinicalTrials.gov NCT01762813. publishedVersion

Background Ageing is the inevitable time-dependent decline in physiological organ function that eventually leads to death. Age is a major risk factor for many of the most common medical conditions, such as cardiovascular disease, cancer, diabetes and Alzheimer's disease. This study reviews currently known hallmarks of ageing and their clinical implications. Methods A literature search of PubMed/MEDLINE was conducted covering the last decade. Results Average life expectancy has increased dramatically over the past century and is estimated to increase even further. Maximum longevity, however, appears unchanged, suggesting a universal limitation to the human organism. Understanding the underlying molecular processes of ageing and health decline may suggest interventions that, if used at an early age, can prevent, delay, alleviate or even reverse age-related diseases. Hallmarks of ageing can be grouped into three main categories. The primary hallmarks cause damage to cellular functions: genomic instability, telomere attrition, epigenetic alterations and loss of proteostasis. These are followed by antagonistic responses to such damage: deregulated nutrient sensing, altered mitochondrial function and cellular senescence. Finally, integrative hallmarks are possible culprits of the clinical phenotype (stem cell exhaustion and altered intercellular communication), which ultimately contribute to the clinical effects of ageing as seen in physiological loss of reserve, organ decline and reduced function. Conclusion The sum of these molecular hallmarks produces the clinical picture of the elderly surgical patient: frailty, sarcopenia, anaemia, poor nutrition and a blunted immune response system. Improved understanding of the ageing processes may give rise to new biomarkers of risk or prognosis, novel treatment targets and translational approaches across disciplines that may improve outcomes.