Abstract
BackgroundPancreatic ductal adenocarcinoma (PDAC) patients suffer poor outcomes, including a five-year survival of below 10%. Poor outcomes result in part from therapeutic resistance that limits the impact of cytotoxic first-line therapy. Novel therapeutic approaches are needed, but currently no targeted therapies exist to treat PDAC.MethodsTo assess cellular resistance mechanisms common to four cytotoxic chemotherapies (gemcitabine, 5-fluorouracil, irinotecan, and oxaliplatin) used to treat PDAC patients, we performed four genome-wide CRISPR activation (CRISPRact) and CRISPR knock-out (CRISPRko) screens in two common PDAC cell lines (Panc-1 and BxPC3). We used pathway analysis to identify gene sets enriched among our hits and conducted RNA-sequencing and chromatin immunoprecipitation-sequencing (ChIP-seq) to characterize top hits from our screen. We used scratch assays to assess changes in cellular migration with HDAC1 overexpression.ResultsOur data revealed activation of ABCG2, a well-described efflux pump, as the most consistent mediator of resistance in each of our screens. CRISPR-mediated activation of genes involved in transcriptional co-repressor complexes also conferred resistance to multiple drugs. Expression of many of these genes, including HDAC1, is associated with reduced survival in PDAC patients. Up-regulation of HDAC1 in vitro increased promoter occupancy and expression of several genes involved in the epithelial-to-mesenchymal transition (EMT). These cells also displayed phenotypic changes in cellular migration consistent with activation of the EMT pathway. The expression changes resulting from HDAC1 activation were also observed with activation of several other co-repressor complex members. Finally, we developed a publicly available analysis tool, PancDS, which integrates gene expression profiles with our screen results to predict drug sensitivity in resected PDAC tumors and cell lines.ConclusionOur results provide a comprehensive resource for identifying cellular mechanisms of drug resistance in PDAC, mechanistically implicate HDAC1, and co-repressor complex members broadly, in multi-drug resistance, and provide an analytical tool for predicting treatment response in PDAC tumors and cell lines.
Highlights
Pancreatic ductal adenocarcinoma (PDAC) patients suffer poor outcomes, including a five-year survival of below 10%
Our results provide a comprehensive resource for identifying cellular mechanisms of drug resistance in PDAC, mechanistically implicate HDAC1, and co-repressor complex members broadly, in multi-drug resistance, and provide an analytical tool for predicting treatment response in PDAC tumors and cell lines
With largely non-specific symptoms and invasive procedures required for diagnosis, only 20% of PDAC patients are eligible for surgical resection, leaving a majority of patients with chemotherapy and radiation as their sole course of treatment [1]
Summary
Pancreatic ductal adenocarcinoma (PDAC) patients suffer poor outcomes, including a five-year survival of below 10%. Poor outcomes result in part from therapeutic resistance that limits the impact of cytotoxic first-line therapy. Despite decades of clinical trials evaluating dozens of potential therapeutics, pancreatic ductal adenocarcinoma (PDAC) has remained largely refractory to improvement of five-year survival rates, which are still less than 10% [1, 2]. Even patients eligible for surgical resection tend to have recurrent disease within 2 years at which time the cancer typically is refractory to adjuvant chemotherapy [3]. The limited effectiveness of cytotoxic chemotherapy has led to a long effort to identify alternative treatment strategies. An alternative strategy is to develop therapies that reverse resistance or increase sensitivity to existing treatments
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