DNA methylation alterations may provide important insights into gene-environment interaction in cancer, aging and complex diseases such as inflammatory bowel disease (IBD). We aim firstly to determine whether the circulating DNA methylome in patients requiring surgery may predict Crohn’s disease (CD) recurrence following intestinal resection; and secondly to compare the circulating methylome seen in patients with established CD with that we had reported in a series of inception cohorts.
TOPPIC was a placebo-controlled, randomised controlled trial of 6-mecaptopurine at 29 UK centres in CD patients undergoing ileocolic resection between 2008 and 2012. Genomic DNA was extracted from whole blood samples from 229 of the 240 patients taken prior to intestinal surgery and analysed using 450KHuman Methylation and Infinium Omni Express Exome arrays (Illumina, San Diego). Co-primary objectives were to determine whether methylation alterations may predict clinical disease recurrence; and to assess whether the epigenetic alterations previously reported in newly diagnosed IBD were present in the CD patients recruited into the TOPPIC study. Differential methylation (DMP) and variance (DVP) analysis was performed comparing patients with and without clinical evidence of recurrence. Secondary analyses included investigation of methylation associations with smoking, genotype (MeQTLs), and chronological age. Validation of our previously published case-control observation of the methylome was performed using historical control data (CD n = 123, Control n=198).
Crohn’s disease recurrence in patients following surgery is associated with five differentially methylated positions (DMPs, Holm p <0.05), including probes mapping to WHSC1 (p=4.1 × 10, Holm p = 0.002) and EFNA3 (p=4.9 × 10, Holm p = 0.02). Five differentially variable positions (DVPs) are demonstrated in the group of patients with evidence of disease recurrence including a probe mapping to MAD1L1 (p = 6.4 × 10). DNA methylation clock analyses demonstrated significant age acceleration in Crohn's disease compared with controls (GrimAge + 2 years (95% Confidence intervals 1.2 to 2.7 years)); with some evidence for accelerated aging in Crohn's patients with disease recurrence following surgery (GrimAge +1.04 years (95% CI -0.04 to 2.22)). Significant methylation differences between CD cases and controls were seen by comparing this cohort in conjunction with previously published control data, including validation of our previously described differentially methylated positions (RPS6KA2 p=1.2×10, SBNO2=1.2×10) and regions (TXK [FDR p = 3.6 ×10], WRAP73 [FDR p = 1.9 ×10], VMP1 [FDR p = 1.7 ×10] and ITGB2 [FDR p=1.4×10]).
We demonstrate differential methylation and differentially variable methylation in patients developing clinical recurrence within 3 years of surgery. Moreover, we report replication of the CD-associated methylome, previously characterised only in adult and paediatric inception cohorts, in patients with medically refractory disease needing surgery.
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