Background Immune checkpoint inhibitors are successfully introduced as anticancer treatment. However, they may induce severe immune-related adverse events (irAEs). One of the most frequent irAEs is diarrhoea. The main objective of this study was to analyse symptoms (ie, grade of diarrhoea), endoscopic and histological features and response to management in immune checkpoint inhibition-related colitis (IRC). Patients and methods We retrospectively analysed patients who developed diarrhoea on checkpoint inhibition and therefore underwent an endoscopy and/or were treated with corticosteroids. Patients were treated between August 2010 and March 2016 for metastatic melanoma or non-small cell lung cancer. Severity of IRC was scored using the endoscopic Mayo score and the van der Heide score. Results Out of a cohort of 781 patients, 92 patients were identified who developed diarrhoea and therefore underwent an endoscopy and/or were treated with corticosteroids. Patients were treated with monotherapy anticytotoxic T-lymphocyte antigen-4, antiprogrammed death receptor-1 or a combination of both. All patients had symptoms of diarrhoea (grade 1: 16%; grade 2: 39% and grade 3: 44%). A complete colonoscopy was performed in 62 (67%) patients, of whom 42 (68%) had a pancolitis (≥3 affected segments). Ulcers were seen in 32% of endoscopies. There was no significant correlation between the grade of diarrhoea at presentation and endoscopic severity scores, the presence of ulcers or histological features. In 54 episodes of diarrhoea (56%), patients received one or more cycles infliximab for steroid-refractory colitis. Patients with higher endoscopic severity scores, ulcers and/or a pancolitis needed infliximab more often. Conclusions The correlation between grade of diarrhoea and endoscopic or histological features for severity of colitis is poor. Patients with higher endoscopic severity scores, ulcers or a pancolitis needed the addition of infliximab more often. Therefore, endoscopy may have value in the evaluation of the severity of IRC and may help in decision making for optimal management.

e21062 Background: Immune checkpoint inhibition has improved survival of patients with several types of advanced cancer. With these new treatments previously unseen adverse events (AE) have become relevant. Considering the fact that most of these immune related AEs can be reversed when identified early and treated adequately, the need for updated algorithms is crucial. Colitis is one of the most severe AEs upon anti-CTLA-4 (e.g. ipilimumab) monotherapy or combination therapies. Severe colitis with deep ulcerations is associated with life-threatening complications such as perforation. Therefore, early recognition and treatment is warranted. Anti-TNF therapy (infliximab) can resolve colitis fast. We sought to identify endoscopic features that indicate the need for early treatment with infliximab. Methods: We retrospectively analyzed all patients who developed diarrhea upon checkpoint inhibition. Only patients undergoing endoscopy were included. All patients were treated at the NKI between August 2010 and March 2016 for either metastatic melanoma or non-small cell lung cancer. Results: A total of 89 patients underwent an endoscopy due to immune related diarrhea (irDiarrhea). Patients were treated with either monotherapy anti-CTLA-4, anti-PD-(L)1, a combination of both, or the combination of anti-CTLA-4 and radiofrequency ablation. Median age was 58 years (range 30-88) and 54% of patients were female. A colonoscopy was performed in 64 (72%) patients and a sigmoidoscopy in 25 (28%) patients. Ulcers were seen during 29 endoscopies (32%). In 18 endoscopies (20%) there were ≤ 10 ulcers and in 11 endoscopies (12%) > 10. Of all patients that had ulcers at endoscopy 23 (79%) required additional infliximab treatment for corticosteroid refractory colitis, while only 25 (42%) of patients without ulcers required additional infliximab ( p= 0.001). Conclusions: Patients with ulcerative irColitis require twice as often infliximab in addition to corticosteroids. Early treatment with infliximab may be indicated in ulcerative irColitis. Treatment based on endoscopic features makes colonoscopy an important diagnostic tool in irDiarrhea. Prospective analysis of endoscopy-driven management of irColitis is warranted.

Background Colorectal cancer develops in a multi-step manner from normal epithelium, through a pre-malignant lesion (so-called adenoma), into a malignant lesion (carcinoma), which invades surrounding tissues and eventually can spread systemically (metastasis). It is estimated that only about 5% of adenomas do progress to a carcinoma. Aim The present study aimed to unravel the biology of adenoma to carcinoma progression by mRNA expression profiling, and to identify candidate biomarkers for adenomas that are truly at high risk of progression. Methods Genome-wide mRNA expression profiles were obtained from a series of 37 colorectal adenomas and 31 colorectal carcinomas using oligonucleotide microarrays. Differentially expressed genes were validated in an independent colorectal gene expression data set. Gene Set Enrichment Analysis (GSEA) was used to identify altered expression of sets of genes associated with specific biological processes, in order to better understand the biology of colorectal adenoma to carcinoma progression. Results mRNA expression of 248 genes was significantly different, of which 96 were upregulated and 152 downregulated in carcinomas compared to adenomas. Classification of adenomas and carcinomas using the expression of these genes showed to be very accurate, also when tested in an independent expression data set. Gene-sets associated with ageing (which is related to senescence) and chromosomal instability were upregulated, and a gene-set associated with fatty acid metabolism was downregulated in carcinomas compared to adenomas. Moreover, gene-sets associated with chromosomal location revealed chromosome 4q22 loss and chromosome 20q gain of gene-set expression as being relevant in this progression. Concluding remark These data are consistent with the notion that adenomas and carcinomas are distinct biological entities. Disruption of specific biological processes like senescence (ageing), maintenance of chromosomal instability and altered metabolism, are key factors in the progression from adenoma to carcinoma.

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Colorectal adenomas form a biologically and clinically distinct intermediate stage in development of colorectal cancer (CRC) from normal colon epithelium. Only 5% of adenomas progress into adenocarcinomas, indicating that malignant transformation requires other biological alterations than those involved in adenoma formation. The present study aimed to explore which cancer-related biological processes are affected during colorectal adenoma-to-carcinoma progression and to identify key genes within these pathways that can serve as tumor markers for malignant transformation. The activity of 12 cancer-related biological processes was compared between 37 colorectal adenomas and 31 adenocarcinomas, using the pathway analysis tool Gene Set Enrichment Analysis. Expression of six gene sets was significantly increased in CRCs compared to adenomas, representing chromosomal instability, proliferation, differentiation, invasion, stroma activation, and angiogenesis. In addition, 18 key genes were identified for these processes based on their significantly increased expression levels. For AURKA and PDGFRB, increased mRNA expression levels were verified at the protein level by immunohistochemical analysis of a series of adenomas and CRCs. This study revealed cancer-related biological processes whose activities are increased during malignant transformation and identified key genes which may be used as tumor markers to improve molecular characterization of colorectal tumors.

Introduction: Colorectal adenomas are common precursors of colorectal cancer (CRC). About only 5% of adenomas progress to cancer and this progression is mostly associated with overall onset/increase of chromosomal instability. Gain of 13q is often implicated in this progression of adenoma to carcinoma (1). Until recently however no genes were identified to be the drivers of this amplicon. Recently, Firestein and collaborators (2) showed that cyclin 8 (CDK8), at 13q12.13, functions as an oncogene in CRC. The aim of this study was to evaluate CDK8 gene dosage effects in colorectal adenoma to carcinoma progression. Material and methods: Sixty seven colorectal tumours (34 adenomas and 28 carcinomas) were analysed by array CGH (5k BAC platform, including contig coverage of 13q) and by expression microarray (30k Compugen library). Integration of DNA copy number dosage and gene expression was performed using the Ace-it tool (3). Results: In the tumours analysed we observed 13q copy number gain in 9% and 46% of adenomas and carcinomas, respectively. Integrating copy number and mRNA expression with the differential upregulation of genes between carcinomas and adenomas, provided us with a list of 43 genes. Within this list, CDK8 ranked 7th in significancy (p=0.003). Conclusions: Copy number gain of 13q has a gene dosage effect on CDK8 mRNA expression, indicating a role of this gene in colorectal adenoma to carcinoma progression. (1) Hermsen M, Postma C, Baak J, Weiss M, Rapallo A, Sciutto A, Roemen G, Arends JW, Williams R, Giaretti W, De Goeij A, Meijer G. Colorectal adenoma to carcinoma progression follows multiple pathways of chromosomal instability. Gastroenterology 2002 Oct;123(4):1109-19. (2) Firestein R, Bass AJ, Kim SY, Dunn IF, Silver SJ, Guney I, Freed E, Ligon AH, Vena N, Ogino S, Chheda MG, Tamayo P, Finn S, Shrestha Y, Boehm JS, Jain S, Bojarski E, Mermel C, Barretina J, Chan JA, Baselga J, Tabernero J, Root DE, Fuchs CS, Loda M, Shivdasani RA, Meyerson M, Hahn WC. CDK8 is a colorectal cancer oncogene that regulates beta-catenin activity. Nature 2008 Sep 25;455(7212):547-51. (3) van Wieringen WN, Belien JA, Vosse SJ, Achame EM, Ylstra B. ACE-it: a tool for genome-wide integration of gene dosage and RNA expression data. Bioinformatics. 2006 Aug 1;22(15):1919-20. Note: This abstract was not presented at the AACR 101st Annual Meeting 2010 because the presenter was unable to attend. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3086.

MicroRNAs are small non-coding RNA molecules, which regulate central mechanisms of tumorigenesis. In colorectal tumours, the combination of gain of 8q and 13q is one of the major factors associated with colorectal adenoma to adenocarcinoma progression. Functional studies on the miR-17-92 cluster localised on 13q31 have shown that its transcription is activated by c-myc, located on 8q, and that it has oncogenic activities. We investigated the contribution of the miR-17-92 cluster during colorectal adenoma to adenocarcinoma progression. Expression levels of the miR-17-92 cluster were determined in 55 colorectal tumours and in 10 controls by real-time RT–PCR. Messenger RNA c-myc expression was also determined by real-time RT–PCR in 48 tumours with array comparative genomic hybridisation (aCGH) data available. From the six members of the miR-17-92 cluster, all except miR-18a, showed significant increased expression in colorectal tumours with miR-17-92 locus gain compared with tumours without miR-17-92 locus gain. Unsupervised cluster analysis clustered the tumours based on the presence of miR-17-92 locus gain. Significant correlation between the expression of c-myc and the six miRNAs was also found. Increased expression of miR-17-92 cluster during colorectal adenoma to adenocarcinoma progression is associated to DNA copy number gain of miR17-92 locus on 13q31 and c-myc expression.

BACKGROUND Colorectal cancer (CRC) is biologically a heterogeneous disease, which may affect response to drug therapy. We investigated the correlation of genome-wide DNA copy number profiles of primary tumors with response to systemic chemotherapy in advanced CRC. PATIENTS AND METHODS DNA was isolated from formaldehyde-fixed paraffin-embedded primary tumors of 32 patients with advanced CRC, which were selected based on either a good response (n = 16) or a poor response (n = 16) to first-line combination therapy with capecitabine and irinotecan. High-resolution DNA copy number profiles were obtained by means of 30 K oligonucleotide-based array comparative genomic hybridization (aCGH). RESULTS Unsupervised hierarchical cluster analysis of the aCGH data revealed two clusters of 19 and 13 tumors, respectively, and cluster membership showed a significant correlation with response status (P < 0.03). The nonresponders had fewer chromosomal alterations compared with the responders, in particular less losses were found (P < 0.03). Most prominent differences between the two groups were losses of regions 18p11.32-q11.2 (P < 0.02) and 18q12.1-q23 (P < 0.03), which were more frequently observed in responders. CONCLUSIONS Differences in DNA copy number profiles of primary CRCs are associated with response to systemic combination chemotherapy with capecitabine and irinotecan. Responders overall had more chromosomal alterations, especially loss of chromosome 18.

An enormous body of literature exists on colorectal adenoma to carcinoma progression, both related to clinical and biological aspects. Inherently, hardly any longitudinal data on humans exist; reason why most knowledge is extrapolated from cross sectional studies, leaving a substantial number of important unresolved issues. What has become clear is that colorectal cancer is a heterogeneous and complex disease in terms of underlying molecular biology, etiology and clinical phenotype. Yet, the interactions between these areas we are only starting to unravel. Examples of this dilemma are, the time it takes for adenomas to progress to cancer, the proportion of colorectal cancer actually arising form e.g. flat or serrated adenomas, the best surrogate endpoint for screening studies, and the clinical and biological meaning of CIMP. Optimal understanding of these aspects is mandatory to arrive to the best strategies for colorectal, especially against the background of new molecular tests for early diagnosis of colorectal cancer coming up.

OBJECTIVE:This study aimed to identify the oncogenes at 20q involved in colorectal adenoma to carcinoma progression by measuring the effect of 20q gain on mRNA expression of genes in this amplicon.METHODS:Segmentation of DNA copy number changes on 20q was performed by array CGH (comparative genomic hybridisation) in 34 non-progressed colorectal adenomas, 41 progressed adenomas (ie, adenomas that present a focus of cancer) and 33 adenocarcinomas. Moreover, a robust analysis of altered expression of genes in these segments was performed by microarray analysis in 37 adenomas and 31 adenocarcinomas. Protein expression was evaluated by immunohistochemistry on tissue microarrays.RESULTS:The genes C20orf24, AURKA, RNPC1, TH1L, ADRM1, C20orf20 and TCFL5, mapping at 20q, were significantly overexpressed in carcinomas compared with adenomas as a consequence of copy number gain of 20q.CONCLUSION:This approach revealed C20orf24, AURKA, RNPC1, TH1L, ADRM1, C20orf20 and TCFL5 genes to be important in chromosomal instability-related adenoma to carcinoma progression. These genes therefore may serve as highly specific biomarkers for colorectal cancer with potential clinical applications.

Colorectal cancer (CRC) is a complex and heterogeneous disease in which genomic instability and DNA promoter methylation play important roles. The aim of this study was to investigate the relationship between chromosomal instability (CIN), microsatellite instability (MSI) and promoter methylation of CRC-associated genes. Therefore, 71 CRCs were analysed for CIN and MSI by comparative genomic hybridization and the mononucleotide marker BAT-26, respectively. Promoter methylation of the tumour suppressor and DNA repair genes hMLH1, O(6)-MGMT, APC, p14(ARF), p16(INK4A), RASSF1A, GATA-4, GATA-5 and CHFR was analysed using methylation-specific polymerase chain reaction. These integrative analyses showed that in CIN+ CRCs, promoter methylation of GATA-4 and p16(INK4A) was inversely related to chromosomal loss at 15q11-q21 and gain at 20q13, respectively (P values: 3.8 x 10(-2) and 4.5 x 10(-2), respectively). Interestingly, promoter methylation of RASSF1A, GATA-4, GATA-5 and CHFR, as well as a high methylation index (MI), was positively related to chromosomal gain at 8q23-qter (P values: 1.5 x 10(-2), 3.8 x 10(-2), 3.9 x 10(-2), 4.9 x 10(-2) and 8.2 x 10(-3), respectively). MSI was associated with BRAF mutation, promoter methylation of hMLH1, APC and p16(INK4A) and a high MI (total number of methylated genes) (P values: 2.4 x 10(-2), 2.5 x 10(-3), 1.8 x 10(-2), 4.6 x 10(-2) and 1.0 x 10(-2), respectively). Therefore, we conclude that promoter methylation of pivotal tumour suppressor and DNA repair genes is associated with specific patterns of chromosomal changes in CRC, which are different from methylation patterns in MSI tumours.