We generate rapid, complex models of serrated CRC to determine the contribution of specific genetic alterations to carcinogenesis. Analysis of our models alongside patient data has led to the identification of a potential susceptibility for this tumour type.

Background & Aims Colorectal cancer is an epigenetically heterogeneous disease, however, the extent and spectrum of the CpG island methylator phenotype (CIMP) is not clear. Methods Genome-scale methylation and transcript expression were measured by DNA Methylation and RNA expression microarray in 216 unselected colorectal cancers, and findings were validated using The Cancer Genome Atlas 450K and RNA sequencing data. Mutations in epigenetic regulators were assessed using CIMP-subtyped Cancer Genome Atlas exomes. Results CIMP-high cancers dichotomized into CIMP-H1 and CIMP-H2 based on methylation profile. KRAS mutation was associated significantly with CIMP-H2 cancers, but not CIMP-H1 cancers. Congruent with increasing methylation, there was a stepwise increase in patient age from 62 years in the CIMP-negative subgroup to 75 years in the CIMP-H1 subgroup ( P < .0001). CIMP-H1 predominantly comprised consensus molecular subtype 1 cancers (70%) whereas consensus molecular subtype 3 was over-represented in the CIMP-H2 subgroup (55%). Polycomb Repressive Complex-2 (PRC2)-marked loci were subjected to significant gene body methylation in CIMP cancers ( P < 1.6 × 10 -78 ). We identified oncogenes susceptible to gene body methylation and Wnt pathway antagonists resistant to gene body methylation. CIMP cluster–specific mutations were observed in chromatin remodeling genes, such as in the SWItch/Sucrose Non-Fermentable and Chromodomain Helicase DNA-Binding gene families. Conclusions There are 5 clinically and molecularly distinct subgroups of colorectal cancer. We show a striking association between CIMP and age, sex, and tumor location, and identify a role for gene body methylation in the progression of serrated neoplasia. These data support our recent findings that CIMP is uncommon in young patients and that BRAF mutant polyps in young patients may have limited potential for malignant progression.

Conventional adenomas are initiated by APC gene mutation that activates the WNT signal. Serrated neoplasia is commonly initiated by BRAF or KRAS mutation. WNT pathway activation may also occur, however, to what extent this is owing to APC mutation is unknown. We examined aberrant nuclear β-catenin immunolocalization as a surrogate for WNT pathway activation and analyzed the entire APC gene coding sequence in serrated and conventional pathway polyps and cancers. WNT pathway activation was a common event in conventional pathway lesions with aberrant nuclear immunolocalization of β-catenin and truncating APC mutations in 90% and 89% of conventional adenomas and 82% and 70% of BRAF wild-type cancers, respectively. WNT pathway activation was seen to a lesser extent in serrated pathway lesions. It occurred at the transition to dysplasia in serrated polyps with a significant increase in nuclear β-catenin labeling from sessile serrated adenomas (10%) to sessile serrated adenomas with dysplasia (55%) and traditional serrated adenomas (9%) to traditional serrated adenomas with dysplasia (39%) (P=0.0001). However, unlike the conventional pathway, truncating APC mutations were rare in the serrated pathway lesions especially sessile serrated adenomas even when dysplastic (15%) and in the BRAF mutant cancers with microsatellite instability that arise from them (8%). In contrast, APC missense mutations that were rare in conventional pathway adenomas and cancers (3% in BRAF wild-type cancers) were more frequent in BRAF mutant cancers with microsatellite instability (32%). We conclude that increased WNT signaling is important in the transition to malignancy in the serrated pathway but that APC mutation is less common and the spectrum of mutations is different than in conventional colorectal carcinogenesis. Moderate impact APC mutations and non-APC-related causes of increased WNT signaling may have a more important role in serrated neoplasia than the truncating APC mutations common in conventional adenomas.