Mutation of SMAD4 in Pancreatic Cancer Results in an Epigenetic Switch in Molecular Subtypes Via Regulation of TET2 and Hydroxymethylation

The epigenome is increasingly appreciated as crucial modifier of cellular plasticity in cancer biology, leading to tumour heterogeneity and emergence of therapeutic resistance. Epigenetic mechanisms can explain tumours that arise without obvious driver mutations, as well as heterogeneous functional phenotypes within tumours that lack genetic explanation. While some tumours display heterogeneity within the primary tumour, pancreatic tumours are not genetically diverse, even when comparing to metastases (Reiter et al, 2018). Conversely, considerable epigenetic reprogramming is associated with progression to more aggressive phenotpyes (McDonald et al, 2018). Using a cohort of pancreatic adenocarcinoma (PDAC) patients, we have characterized the mutational landscape in combination with the transcriptome, DNA methylome and hydroxymethylome of PDAC tumours. Using an integrated, multi-genomic clustering approach, we report how pancreatic cancer is marked by a loss of cytosine-hydroxymethylation at specific loci. Specific peaks of hydroxymethylation at the boarders of regulatory regions have been described to generate protective “rims”, which prevent the spread of cytosine-methylation and maintain an activated transcriptional state (Li et al, 2018). We find that pancreatic tumours lose hydroxymethylation-rims from the promoters of differentiation genes that define pancreatic identity. This correlates with increased methyl-cytosine associated gene silencing of differentiation genes and this allows the emergence of the aggressive subtype of pancreatic cancer known as basal, quasi-mesenchymal or squamous. Moreover, we also report how expression of the main hydroxymethylation enzyme in the adult pancreas, TET2, is a SMAD4 target gene, and that alterations in epigenetics and cell phenotype explains the poorer prognosis associated with SMAD4 mutation in patients. Thus, we report how a single genomic mutation results in widespread epigenetic rewriting of the PDAC genome and alteration of cell phenotype to squamous. Pharmacological inhibition of the epigenome may lead to a reversion of tumours to molecular subtypes that are more amenable to therapy.