Targeting of the Histone 3 Lysine 9 Methyltransferase Pathway in Kras‐Induced Cell Growth and Pancreatic Cancer

Pancreatic ductular adenocarcinoma (PDAC) ranks fourth as a cause of cancer death in the USA and is almost universally fatal, with the annual number of deaths equivalent to the number of newly diagnosed cases. Valuable research in the field has revealed genetic aberrations that contribute to PDAC development and progression, with KRAS being one of the most frequent mutations in more than 90% of patient samples. However, to date, any efforts to directly target KRAS have failed in the clinic. Thus, there is indisputably an urgent need to further improve our understanding of molecular mechanisms underlying PDAC development as to identify novel therapeutic targets, including druggable important downstream targets and nodes orchestrated by oncogenic KRAS. In the current study, using a cell model that allows inducible expression of mutant KRAS G12D , we find that protein levels of the dimethyl‐K9H3 histone methyl transferase (HMT), G9a, and its complex partners are increased in response to activation of the oncogenic Kras pathway. Furthermore, the activation of this oncogenic pathway results in the formation of the G9a‐GLP‐Wiz trimer complex, as determined by affinity protein purification, combined with mass spectrometry. In vivo experiments involving the cross of the Pdx1‐CRE/LSL‐KRAS G12D mice with G9a fl/fl animals demonstrate that a loss of the H3K9Me2 mark in the nucleus of exocrine cells is accompanied by a significantly reduced number of PanIN lesions. RNA‐Seq experiments from these animals reveal that these mice have reduced levels of typical molecular markers of PanINs and have an upregulation of p21, a cyclin‐dependent kinase inhibitor that is a major effector of p53 activity and facilitates cell cycle arrest in response to DNA damage. Increased levels of p21 protein are also observed in lysates from Pdx1‐CRE/LSL‐KRAS G12D /G9a −/− animals compared to Pdx1‐CRE/LSL‐KRAS G12D control mice. Pharmacological inhibition and siRNA‐mediated knockdown of G9a in cells recapitulates this effect with p21 upregulation. Furthermore, we find G9a and its H3K9Me2 mark occupy the p21 promoter, suggesting G9a is involved in the direct regulation of this gene. Congruently, pharmacological inhibition of G9a using BRD4770 displays an inhibitory effect on KRAS G12D ‐induced cell proliferation, as well as pancreatic tumor growth in a subcutaneous xenograft model. Combined, these data provide evidence for a key role of the meK9H3‐G9a pathway as a mediator of the oncogenic Kras response and defines a novel point of potential therapeutic intervention for PDAC. Support or Funding Information NIH R01CA178627; NIH R01DK58185; Advancing a Healthier Wisconsin Endowment This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal .