TET2 drives 5hmc marking of GATA6 and epigenetically defines pancreatic ductal adenocarcinoma transcriptional subtypes

Abstract

Background and Aims: Pancreatic ductal adenocarcinoma (PDAC) is characterised by advanced disease stage at presentation, aggressive disease biology and resistance to therapy resulting in extremely poor five-year survival <10%. PDAC is classified into transcriptional subtypes with distinct survival characteristics, although how these arise is not known. Epigenetic deregulation, rather than genetics, has been proposed to underpin progression but exactly why is unclear and hindered by analysis of clinical samples. Methods: Genome-wide epigenetic mapping of DNA modifications 5-hydroxymethylcytosine (5mc) and 5-hydroxymethylcytosine (5hmc) using oxidative bisulphite sequencing (oxBS). Bioinformatics using iCluster and mutational profiling to identify overlap with transcriptional signatures in FFPE from resected patients and confirmation in vivo. Results: We find that more aggressive squamous-like PDAC subtypes result from epigenetic inactivation of loci including GATA6 that promote differentiated classical-pancreatic subtypes. We show that squamous-like PDAC transcriptional subtypes are associated with greater loss of 5hmc due to reduced expression of the 5mc-hydroxylase TET2. Furthermore, we find that SMAD4 directly supports TET2 levels in the pancreas and classical-pancreatic tumors and loss of SMAD4 expression is associated reduced 5hmc, GATA6 and squamous-like tumors. Importantly, enhancing TET2 stability using Metformin and VitaminC/ascorbic acid (AA) restores 5hmc and GATA6 levels, reverting squamous-like tumor phenotypes and WNT-dependence in vitro and in vivo. Conclusions: We identify epigenetic deregulation of pancreatic differentiation as an underpinning event behind the emergence of transcriptomic subtypes in PDAC. Our data shows that restoring epigenetic control increases biomarkers of classical-pancreatic tumors and raises the possibility that combination of Vitamin C and Metformin may prolong survival in patients with squamous-like pancreatic cancer.

Molecular Landscape and Prognostic Biomarker Analysis of Advanced Pancreatic Cancer and Predictors of Treatment Efficacy of AG Chemotherapy

“…Studies have classified PDAC into classical, quasimesenchymal, and exocrine-like subtypes. Classical PDAC showed that higher GATA6 mRNA expression is associated with a better prognosis ( 35 , 36 ). Additionally, low GATA6 expression is associated with shorter OS ( 37 ).…”

Chemical-induced Gene Expression Ranking and its Application to Pancreatic Cancer Drug Repurposing

“…Furthermore, we aimed to discover drugs that can induce drug sensitivity of pancreatic cancer sub-types that are resistant to existing anti-cancer therapies rather than screen compounds that can kill cancer cells directly. A recent study has shown that the combination of metformin and vitamin C can restore TET2 and GATA6 activities in aggressive squamous-like pancreatic ductal adenocarcinoma sub-type, which are the biomarkers of classical-pancreatic tumor, thereby improving therapeutic responses and survival of aggressive pancreatic sub-type 43 . The main step of this screening pipeline is to compare the chemical-induced gene expression profiles generated by CIGER with gene expression profile computed from pancreatic cancer cell lines treated by metformin and vitamin C. For drug gene expression profiles, we send queries to the DrugBank database to retrieve the list of all existing drugs (i.e., 11179 drugs) with their corresponding SMILES representations and then use CIGER trained on the LINCS L1000 dataset to generate profiles for these drugs from their SMILES representations.…”

“…The main step of this screening pipeline is to compare the chemical-induced gene expression profiles generated by CIGER with gene expression profile computed from pancreatic cancer cell lines treated by metformin and vitamin C. For drug gene expression profiles, we send queries to the DrugBank database to retrieve the list of all existing drugs (i.e., 11179 drugs) with their corresponding SMILES representations and then use CIGER trained on the LINCS L1000 dataset to generate profiles for these drugs from their SMILES representations. For gene expression profile of pancreatic cancer treated by metformin and vitamin C, we perform differential expression analysis with DESeq2 44 between metformin and vitamin C treated samples and mock-treated samples 43 . Then, we compute the similarity with respect to ranking information between the gene expression profiles of treatment and drugs across 10 cell lines by GSEA and Precision@200 scores to find potential treatment for this disease.…”

“…As PI3K/AKT/mTOR signaling is one of the most important intracellular pathways that regulate the cell cycle, it can be targeted by drugs to regulate the metabolism in cancer cells, resulting in phenotype shift, increased cell death, and decreased cell proliferation 46,47 . Biguanide and its medication drug (i.e., Metformin) have been shown to be effective for pancreatic cancer tumor growth inhibition 43,48,49 . Note that the drugs selected by CIGER are not available in the training set of LINCS L1000 dataset, thereby showing its real potential application in drug discovery that enables high-throughput phenotypic drug screening by utilizing the molecular structure's information only.…”

“…Treatment Gene Expression Profile For pancreatic cancer treatment gene expression profile, we perform differential expression analysis between metformin and vitamin C treated samples and mock-treated samples with DESeq2 44 . A recent study showed that metformin and vitamin C treatment can restore TET2 activity in aggressive squamous-like pancreatic ductal adenocarcinoma subtype, and increase biomarker of classical-pancreatic tumor, which are related to improved therapeutic responses and survival 43 . To search for drugs that can restore epigenetic control in pancreatic tumors like metformin and vitamin C, we used pancreatic cancer gene expression data from this study, where the human pancreatic tumor cell line PSN1, orthotopically implanted into mice, was treated with Metformin combined with vitamin C or mock treatment.…”