Abstract 3190: Using a genome-wide shRNA screen to investigate the mechanism of stroma-conferred gemcitabine resistance in pancreatic ductal adenocarcinoma
- Abstract:
- Abstract The purpose of this study is to identify mechanisms of resistance to gemcitabine in pancreatic ductal adenocarcinoma (PDAC) using an in vitro coculture system. The inherent resistance of PDAC to first line treatments such as gemcitabine has been attributed to the dense fibrotic stroma. Targeting stromal components, in particular cancer-associated fibroblasts (CAFs), the activated form of pancreatic stellate cells (PSCs), is a promising avenue for therapeutic intervention. We have demonstrated previously the induction of gemcitabine resistance when co-culturing KRAS G12D; p53 R172H; Pdx-cre (KPC) mouse PDAC-derived tumour cells with αSMA+ fibroblast-like cells from PDAC tumour (FLCs), in comparison to PDAC cells co-cultured with PSCs or in monoculture. This resistance-inducing effect is specific to cells of mesenchymal phenotype, is transient in nature and a product of cell-cell contact. To investigate the exact mechanism by which PDAC cells become resistant to gemcitabine in coculture, we undertook a genome-wide shRNA depletion viability screen, infecting KPC mouse PDAC cells with a pooled shERWOOD UltramiR shRNA library both in mono- and co-culture with FLCs, in the presence and absence of gemcitabine. Next-generation sequencing and differential expression analysis, using DESeq2, identified shRNAs depleted through 6 cycles of gemcitabine treatment in coculture, indicating they target genes driving the resistance effect. Pathway analysis of differential expression using MetaCore implicated DNA damage repair and protein modification pathways in the gemcitabine resistance effect. Depletion of p97, ATR, or Chk1 all sensitised PDAC cells to gemcitabine in coculture, corroborating findings with translational value in previous studies. Further evaluation is ongoing using validation shRNA dropout assays in combination with targeted literature reviews to collate a candidate list of targetable genes with translational value in preventing and combating chemotherapy resistance in patients with PDAC. Citation Format: Graham D. Mills, Nicolas Erard, Frances M. Richards, Gregory J. Hannon, Duncan I. Jodrell. Using a genome-wide shRNA screen to investigate the mechanism of stroma-conferred gemcitabine resistance in pancreatic ductal adenocarcinoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 3190. doi:10.1158/1538-7445.AM2017-3190
- Authors:
- GD Mills, N Erard, FM Richards, GJ Hannon, DI Jodrell
- Journal:
- Cancer Research
- Citation info:
- 77(13_Supplement):3190-3190
- Publication date:
- 1st Jul 2017
- Full text
- DOI