Objective Pancreatic ductal adenocarcinoma (PDAC) is an aggressive malignancy with a 5-year survival of less than 5%. Transcriptomic analysis has identified two clinically relevant molecular subtypes of PDAC: classical and basal-like. The classical subtype is characterised by a more favourable prognosis and better response to chemotherapy than the basal-like subtype. The classical subtype also expresses higher levels of lineage specifiers that regulate endodermal differentiation, including the nuclear receptor hepatocyte nuclear factor 4 α (HNF4α). The objective of this study is to evaluate the role of HNF4α, SIX4 and SIX1 in regulating the growth and molecular subtype of PDAC.
Design We manipulate the expression of HNF4α, SIX4 and SIX1 in multiple in vitro and in vivo PDAC models. We determine the consequences of manipulating these genes on PDAC growth, differentiation and molecular subtype using functional assays, gene expression analysis and cross-species comparisons with human datasets.
Results We show that HNF4α restrains tumour growth and drives tumour cells toward an epithelial identity. Gene expression analysis of murine models and human tumours shows that HNF4α activates expression of genes associated with the classical subtype. HNF4α also directly represses SIX4 and SIX1, two mesodermal/neuronal lineage specifiers expressed in the basal-like subtype. Finally, SIX4 and SIX1 drive proliferation and regulate differentiation in HNF4α-negative PDAC.
Conclusion Our data show that HNF4α regulates the growth and molecular subtype of PDAC by multiple mechanisms, including activation of the classical gene expression programme and repression of SIX4 and SIX1, which may represent novel dependencies of the basal-like subtype.
- pancreatic cancer
- gene expression
- molecular mechanisms
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Contributors SAC and ELS designed experiments. SAC, VKB, KCB and HC performed experiments. SAC, LT-H, JMV, CS, RM, JG, and ELS analysed data. JS generated PDXs. ELS performed histopathological review. SAC and ELS wrote the manuscript. All authors discussed results, reviewed and revised the manuscript.
Funding ELS was supported grants from National Institutes of Health (NIH) (R21CA194764, R01CA237404, R01CA240317 and R01CA212415), by a Career Award for Medical Scientists from the Burroughs Wellcome Fund and a V Scholar Award. The research reported in this publication was supported by institutional funds (Huntsman Cancer Foundation and Department of Pathology, University of Utah) and NIH grant P30CA042014 awarded to Huntsman Cancer Institute and to the NC Programme at Huntsman Cancer Institute. Research reported in this publication used shared resources (including flow cytometry, high-throughput genomics, bioinformatics, and biorepository and molecular pathology) at the University of Utah was supported by the P30CA042014. Research reported in this publication used shared resources (including flow cytometry, high-throughput genomics, bioinformatics and biorepository and molecular pathology) at the University of Utah and was supported by the National Cancer Institute of the NIH under Award no: P30CA042014. Work in the flow cytometry core was also supported by the National Centre for Research Resources of the NIH under Award no: 1S20RR026802.
Competing interests RM has filed a pending US Patent Application 15/518,900 that involves using gene expression to direct therapy.
Patient and public involvement Patients and/or the public were not involved in the design, conduct, reporting or dissemination plans of this research. Patients were not directly involved in this study. Patient Derived Xenograft (PDX) PDAC models were derived at the University of Utah.
Patient consent for publication Not required.
Provenance and peer review Not commissioned; externally peer reviewed.
Data availability statement Data are available in a public, open access repository. All data relevant to the study are included in the article or uploaded as supplementary information. Gene expression, ChIP-Seq and ATAC-seq data are accessible in the NCBI Gene Expression Omnibus database under accession numbers GSE138145, GSE144750 and GSE138463.
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