Targeting thioredoxin reductase in pancreatic cancer


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Khaled Alhadyan1,Stewart G. Martin2,Sarah J. Storr2,Martyn Inman3,Abed M. Zaitoun4,Dileep Lobo5,Chris Moody3
1,2Academic Unit of Clinical Oncology, University of Nottingham, School of Medicine, Nottingham University Hospitals NHS Trust, City Hospital Campus, Nottingham,3School of Chemistry, University of Nottingham,4Department of Pathology, Nottingham University Hospitals, Queen’s Medical Centre, Nottingham,5Division of Gastrointestinal Surgery, University of Nottingham, Nottingham Digestive Diseases Centre National Institute of Health Research Biomedical Research Unit, Nottingham

Abstract

Background

Current treatment options for pancreatic cancer have failed to improve 5-year survival rates which remain very low, at 3.7%; there is an urgent need to identify new treatment modalities. Redox proteins are key members of cellular antioxidant systems, required to maintain redox homeostasis, with the thioredoxin system being an important component. We have developed novel indolequinones (IQs) as thioredoxin reductase inhibitors and are assessing them in pancreatic cancer models as single agents and in combination with ionising radiation.

Method

Western blotting assessed protein levels +/- drug. In vitro cytotoxicity (proliferation and clonogenic survival) of two agents (IQ9 and IQ10) was assessed in PANC-1, MIAPaCa-2 and BxPc-3 human pancreatic cancer cell lines. Cells were treated with the clonogenic IQ IC50 dose for 48 hours, then irradiated with a single dose of 1, 2, 4 or 6 Gy. Following irradiation cells were plated for clonogenic survival. ROS levels were measured using DCHFDA flow cytometry, following treatment with the drugs for 1, 4, 24 and 48 hours either alone or with additional H2O2 exposure.

Results

IQ treatment caused reduced expression of Thioredoxin. Drugs exhibit potent in vitro cytotoxicity at 24, 48 and 72 hours with IC50 values in the low nM range (~ 90nM at 72h). IC50 doses combined with subsequent hydrogen peroxide (H2O2) exposure increased oxidative stress ~2 fold. Preliminary data suggest radiosensitisation of PANC-1 and BxPc-3 cells, with a 50% and 25% decrease in surviving fraction at 6 Gray respectively, with no effect on MiaPaca2’s.

Conclusion

TrxR represents a potential novel target in pancreatic cancer. Indolequinone’s are highly effective in pancreatic cancer cell lines as single agents and enhance radiotherapeutic efficacy in certain lines. Future work will include 3D culture, use of non-malignant pancreatic cell lines and in vivo radiosensitising experiments.