Cdc7 is a potent anti-cancer target in pancreatic cancer due to abrogation of the DNA origin activation checkpoint
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Matthew T. Huggett1,2, Slavica Tudzarova2, Ian Proctor2, Marco Loddo2,3, Margaret G. Keane1, Kai Stoeber2, Gareth H. Williams2,3, Stephen P. Pereira1
1UCL Institute for Liver and Digestive Health and UCL Cancer Institute, University College London, London, UK
2The Research Department of Pathology, UCL Cancer Institute, University College London, London, UK
3Oncologica Ltd, The Science Village, Chesterford Research Park, Cambridge, UK
Gareth Williams, e-mail: email@example.com
Keywords: pancreatic cancer, cell cycle, DNA replication, Cdc7
Received: July 22, 2015 Accepted: January 23, 2016 Published: February 23, 2016
Purpose: Cdc7 is a serine/threonine kinase which is responsible for the ‘firing’ of replication origins leading to initiation of DNA replication. Inhibition or depletion of Cdc7 in normal cells triggers a DNA origin activation checkpoint causing a reversible G1 arrest. Here we investigate Cdc7 as a novel therapeutic target in pancreatic cancer.
Experimental design: Cdc7 target validation was performed by immunoexpression profiling in a cohort of 73 patients with pancreatic adenocarcinoma including 24 controls. Secondly Cdc7 kinase was targeted in Capan-1 and PANC-1 pancreatic cancer cell line models using either an siRNA against Cdc7 or alternatively a small molecule inhibitor (SMI) of Cdc7 (PHA-767491).
Results: Cdc7 was significantly overexpressed in pancreatic adenocarcinoma compared to benign pancreatic tissue (median LI 34.3% vs. 1.3%; P<0.0001). Cdc7 knockdown using siRNA in Capan-1 and PANC-1 cells resulted in marked apoptotic cell death when compared with control cells. A prominent sub-G1 peak was seen on flow cytometry (sub-G1 51% vs. 3% and 45% vs. 0.7% in Capan-1 and PANC-1 cells, respectively). Annexin V labelling confirmed apoptosis in 64% vs. 11% and 75% vs. 8%, respectively. Western blotting showed cleavage of PARP-1 and caspase-3 and presence of γH2A.X. TUNEL assay showed strong staining in treated cells. These results were mirrored following Cdc7 kinase inhibition with PHA-767491.
Conclusions: Our findings show that Cdc7 is a potent anti-cancer target in pancreatic adenocarcinoma and that Cdc7 immunoexpression levels might be used as a companion diagnostic to predict response to therapeutic siRNAs or SMIs directed against this kinase.
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