Promotion of gemcitabine resistance in pancreatic cancer cells by three-dimensional collagen I through HMGA2-dependent histone acetyltransferase expression.

Abstract

172 Background: Pancreatic ductal adenocarcinoma (PDAC) is associated with a pronounced stromal reaction that has been shown to contribute to chemo-resistance. We have previously shown that PDAC cells are resistant to gemcitabine chemotherapy in the collagen microenvironment due to increased expression of the chromatin remodeling protein high mobility group A2 (HMGA2). Methods: Pancreatic TMAs were stained with trichrome and for histone H3K9, H3K27 acetylation (Ac), and histone acetyltransferase (HAT) expression. PDAC cells were plated onto tissue culture plastic or in three-dimensional (3D) collagen gels and protein expression assessed by Western blotting. DNA damage response was assessed by comet and clonogenic assays. Results: PDAC tumors display higher levels of H3K9Ac and H3K27Ac in fibrotic regions. Moreover, PDAC cells upregulate H3K9Ac and H3K27Ac along with GCN5, PCAF and p300 HATs in 3D collagen compared to tissue culture plastic. Knocking down HMGA2 attenuates the effect of collagen on H3K9Ac, H3K27Ac and p300, PCAF and GCN5 expression. We also show that human PDAC tumors with HMGA2 expression demonstrate increased H3K9Ac and H3K27Ac. Additionally, we show that cells in 3D collagen gels demonstrate reduced tailing with the comet assay, increased clonogenic potential and increased γH2AX following gemcitabine treatment, suggesting an increased response and repair to damaged DNA in the 3D collagen microenvironment. Significantly, down-regulation of HMGA2 or p300, PCAF and GCN5 HATs decreases gemcitabine-induced γH2AX detected and attenuates clonogenic potential. Conclusions: Collagen microenvironment limits the effectiveness of gemcitabine through HMGA2-dependent HAT expression. HMGA2 expression is associated with histone acetylation and HAT expression in human PDAC tumors, particularly in area of fibrosis, suggesting that fibrosis may contribute to chemo-resistance through increased HMGA2-HAT signaling. Overall, our results increase our understanding of how the collagen microenvironment contributes to chemo-resistance and identify HATs as potential therapeutic targets against this deadly cancer.