Milea Timbergen

98 particularly in view of the reported interaction between CTNNB1 and DNMT1 15 . Therefore, the genome-wide DNA methylation profiles of DTF tumours were explored, comparing CTNNB1 S45F and T41A mutated tumours. To this end, 29 FFPE samples of primary untreated DTF tumours were analysed using MeD-seq. Fifteen samples harboured a CTNNB1 T41A mutation and 14 samples a CTNNB1 S45F mutation, both mutations that are commonly observed. The patients had a median age of 36 years (interquartile range 26-47 years) and females were in the majority (n = 19, 66%). Most tumours were located extra-abdominally (69%) and had a median size of 55 mm (interquartile range 34-87 mm) (Table 1). Table 1. Clinical characteristics of patients and tumour samples (n = 29) included in this study Total group n = 29 T41A n (%) S45F n (%) Sex Female 19 (66%) 11 (73%) 8 (57%) Male 10 (34%) 4 (27%) 6 (43%) Tumour location Extra-abdominal 20 (69%) 7 (47%) 13 (93%) Abdominal wall 6 (21%) 5 (33%) 1 (7%) Intra-abdominal 3 (10%) 3 (20%) 0 Median age at diagnosis in years (IQR) 36 (26-47) 38 (33-48) 31 (20-45) Median size in mm (IQR) a,b 55 (34-87) 53 (29-59) 68 (50-103) a based on initial imaging; b n = 3 missing values; IQR, interquartile range Differentially methylated regions between S45F and T41A DTF samples identi- fied by MeD-seq A genome-wide MeD-seq analysis was carried out using DNA isolated from 29 FFPE DTF samples. A total of 365 differentially methylated regions (DMRs) were found to be significantly different between S45F, and T41A DTF tumours. After excluding DMRs located on the X- and Y-chromosomes, a group of 354 DMRs remained. Of these 354 DMRs, 97 (27%) DMRs were hypomethylated in S45F (versus T41A), and 257 (83%) were hypermethylated in S45F (vs. T41A). Supplemental Materials 1 provides an overview of all 354 DMRs. A supervised hierarchical clustering of the DTF samples based on all detected DMRs, did not reveal clearly separated groups based on CTNNB1 mutational status (Figure 1). Likewise, no distinct cluster patterns were observed on basis of sex (Figure 1), tumour site (Supplemental Figure 1) and tumour size (Supplemental Figure 2). As a meta-analysis by our group revealed tumour size to be an independent predictor for recurrence 7 a supervised cluster analysis was performed based on DMRs between small DTF (n = 6, ≤ 34 mm) and large tumours (n = 6, > 87 mm). Interestingly, the small and large DTF tumours were almost completely distinguished only one small tumour grouped together with the large tumours (Supplemental Figure 3). 4