Doxycycline down-regulates DNA-PK and radiosensitizes tumor initiating cells: Implications for more effective radiation therapy.

Authors

Rebecca Lamb, The Breakthrough Breast Cancer Research Unit, Institute of Cancer Sciences, University of Manchester; The Manchester Centre for Cellular Metabolism (MCCM), Institute of Cancer Sciences, University of ManchesterFollow
Marco Fiorillo, The Breakthrough Breast Cancer Research Unit, Institute of Cancer Sciences, University of Manchester; The Manchester Centre for Cellular Metabolism (MCCM), Institute of Cancer Sciences, University of Manchester; The Department of Pharmacy, Health and Nutritional Sciences, The University of CalabriaFollow
Amy Chadwick, The Breakthrough Breast Cancer Research Unit, Institute of Cancer Sciences, University of Manchester; The Manchester Centre for Cellular Metabolism (MCCM), Institute of Cancer Sciences, University of Manchester
Bela Ozsvari, The Breakthrough Breast Cancer Research Unit, Institute of Cancer Sciences, University of Manchester; The Manchester Centre for Cellular Metabolism (MCCM), Institute of Cancer Sciences, University of ManchesterFollow
Kimberly J Reeves, The Breakthrough Breast Cancer Research Unit, Institute of Cancer Sciences, University of Manchester; The Manchester Centre for Cellular Metabolism (MCCM), Institute of Cancer Sciences, University of Manchester
Duncan L Smith, The Cancer Research UK Manchester Institute, University of ManchestFollow
Robert B Clarke, The Breakthrough Breast Cancer Research Unit, Institute of Cancer Sciences, University of ManchesterFollow
Sacha J Howell, The Breakthrough Breast Cancer Research Unit, Institute of Cancer Sciences, University of Manchester
Anna Rita Cappello, The Department of Pharmacy, Health and Nutritional Sciences, The University of CalabriaFollow
Ubaldo E. Martinez-Outshoorn, MD, The Sidney Kimmel Cancer Center, Thomas Jefferson UniversityFollow
Maria Peiris-Pagès, The Breakthrough Breast Cancer Research Unit, Institute of Cancer Sciences, University of Manchester; The Manchester Centre for Cellular Metabolism (MCCM), Institute of Cancer Sciences, University of ManchesterFollow
Federica Sotgia, The Breakthrough Breast Cancer Research Unit, Institute of Cancer Sciences, University of Manchester; The Manchester Centre for Cellular Metabolism (MCCM), Institute of Cancer Sciences, University of ManchesterFollow
Michael P Lisanti, The Breakthrough Breast Cancer Research Unit, Institute of Cancer Sciences, University of Manchester; The Manchester Centre for Cellular Metabolism (MCCM), Institute of Cancer Sciences, University of ManchesterFollow

Document Type

Article

Publication Date

6-10-2015

Comments

This article has been peer reviewed. It was published in: Oncotarget.

Volume 6, Issue 16, 2015, Pages 14005-14025.

The published version is available at: http://www.impactjournals.com/oncotarget/index.php?journal=oncotarget&page=article&op=view&path[]=4159

Copyright © 2015 The Authors

Abstract

DNA-PK is an enzyme that is required for proper DNA-repair and is thought to confer radio-resistance in cancer cells. As a consequence, it is a high-profile validated target for new pharmaceutical development. However, no FDA-approved DNA-PK inhibitors have emerged, despite many years of drug discovery and lead optimization. This is largely because existing DNA-PK inhibitors suffer from poor pharmacokinetics. They are not well absorbed and/or are unstable, with a short plasma half-life. Here, we identified the first FDA-approved DNA-PK inhibitor by "chemical proteomics". In an effort to understand how doxycycline targets cancer stem-like cells (CSCs), we serendipitously discovered that doxycycline reduces DNA-PK protein expression by nearly 15-fold (> 90%). In accordance with these observations, we show that doxycycline functionally radio-sensitizes breast CSCs, by up to 4.5-fold. Moreover, we demonstrate that DNA-PK is highly over-expressed in both MCF7- and T47D-derived mammospheres. Interestingly, genetic or pharmacological inhibition of DNA-PK in MCF7 cells is sufficient to functionally block mammosphere formation. Thus, it appears that active DNA-repair is required for the clonal expansion of CSCs. Mechanistically, doxycycline treatment dramatically reduced the oxidative mitochondrial capacity and the glycolytic activity of cancer cells, consistent with previous studies linking DNA-PK expression to the proper maintenance of mitochondrial DNA integrity and copy number. Using a luciferase-based assay, we observed that doxycycline treatment quantitatively reduces the anti-oxidant response (NRF1/2) and effectively blocks signaling along multiple independent pathways normally associated with stem cells, including STAT1/3, Sonic Hedgehog (Shh), Notch, WNT and TGF-beta signaling. In conclusion, we propose that the efficacy of doxycycline as a DNA-PK inhibitor should be tested in Phase-II clinical trials, in combination with radio-therapy. Doxycycline has excellent pharmacokinetics, with nearly 100% oral absorption and a long serum half-life (18-22 hours), at a standard dose of 200-mg per day. In further support of this idea, we show that doxycycline effectively inhibits the mammosphere-forming activity of primary breast cancer samples, derived from metastatic disease sites (pleural effusions or ascites fluid). Our results also have possible implications for the radio-therapy of brain tumors and/or brain metastases, as doxycycline is known to effectively cross the blood-brain barrier. Further studies will be needed to determine if other tetracycline family members also confer radio-sensitivity.

Recommended Citation

Lamb, Rebecca; Fiorillo, Marco; Chadwick, Amy; Ozsvari, Bela; Reeves, Kimberly J; Smith, Duncan L; Clarke, Robert B; Howell, Sacha J; Cappello, Anna Rita; Martinez-Outshoorn, MD, Ubaldo E.; Peiris-Pagès, Maria; Sotgia, Federica; and Lisanti, Michael P, "Doxycycline down-regulates DNA-PK and radiosensitizes tumor initiating cells: Implications for more effective radiation therapy." (2015). Kimmel Cancer Center Faculty Papers. Paper 51.
https://jdc.jefferson.edu/kimmelccfp/51

PubMed ID

26087309