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NRF2 orchestrates the redox regulation induced by radiation therapy, sustaining embryonal and alveolar rhabdomyosarcoma cells radioresistance

  • Francesco MaramponEmail author
  • Silvia Codenotti
  • Francesca Megiorni
  • Andrea Del Fattore
  • Simona Camero
  • Giovanni Luca Gravina
  • Claudio Festuccia
  • Daniela Musio
  • Francesca De Felice
  • Valerio Nardone
  • Anna Natalizia Santoro
  • Carlo Dominici
  • Alessandro Fanzani
  • Luigi Pirtoli
  • Antonella Fioravanti
  • Vincenzo Tombolini
  • Sara Cheleschi
  • Paolo Tini
Original Article – Cancer Research
  • 55 Downloads

Abstract

Purpose

Tumor cells generally exhibit higher levels of reactive oxygen species (ROS), however, when stressed, tumor cells can undergo a process of ‘Redox Resetting’ to acquire a new redox balance with stronger antioxidant systems that enable cancer cells to become resistant to radiation therapy (RT). Here, we describe how RT affects the oxidant/antioxidant balance in human embryonal (RD) and alveolar (RH30) rhabdomyosarcoma (RMS) cell lines, investigating on the molecular mechanisms involved.

Methods

Radiations were delivered using an x-6 MV photon linear accelerator and their effects were assessed by vitality and clonogenic assays. The expression of specific antioxidant-enzymes, such as Superoxide Dismutases (SODs), Catalase (CAT) and Glutathione Peroxidases 4 (GPx4), miRNAs (miR-22, -126, -210, -375, -146a, -34a) and the transcription factor NRF2 was analyzed by quantitative polymerase chain reaction (q-PCR) and western blotting. RNA interference experiments were performed to evaluate the role of NRF2.

Results

Doses of RT higher than 2 Gy significantly affected RMS clonogenic ability by increasing ROS production. RMS rapidly and efficiently brought back ROS levels by up-regulating the gene expression of antioxidant enzymes, miRNAs as well as of NRF2. Silencing of NRF2 restrained the RMS ability to counteract RT-induced ROS accumulation, antioxidant enzyme and miRNA expression and was able to increase the abundance of γ-H2AX, a biomarker of DNA damage, in RT-treated cells.

Conclusions

Taken together, our data suggest the strategic role of oxidant/antioxidant balance in restraining the therapeutic efficiency of RT in RMS treatment and identify NRF2 as a new potential molecular target whose inhibition might represent a novel radiosensitizing therapeutic strategy for RMS clinical management.

Keywords

Rhabdomyosarcoma Radiotherapy Radioresistance Reactive oxygen species Anti-oxidant NRF2 

Notes

Acknowledgements

We are grateful to the Umberto Veronesi Foundation for awarding a post-doctoral fellowship to Francesco Marampon for the year 2018 and “FIVA Confcommercio” for supporting part of our work.

Author contributions

FMa, SC, FMe and ADF planned experiments; SC, GLG, CF, DM, FDF, VN and ANS performed experiments; CD, LP, AF VT and AF analyze data; SC and PT wrote the paper.

Compliance with ethical standards

Conflict of interest

The other authors declare that they have no competing interests.

Ethical approval

Studies on animal models were not performed. This article does not contain any studies with human participants performed by any of the authors.

Informed consent

This article does not contain any studies with human participants performed by any of the authors.

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Copyright information

© Springer-Verlag GmbH Germany, part of Springer Nature 2019

Authors and Affiliations

  • Francesco Marampon
    • 1
    Email author
  • Silvia Codenotti
    • 2
  • Francesca Megiorni
    • 3
  • Andrea Del Fattore
    • 4
  • Simona Camero
    • 3
  • Giovanni Luca Gravina
    • 5
  • Claudio Festuccia
    • 5
  • Daniela Musio
    • 1
  • Francesca De Felice
    • 1
  • Valerio Nardone
    • 6
  • Anna Natalizia Santoro
    • 7
  • Carlo Dominici
    • 1
  • Alessandro Fanzani
    • 2
  • Luigi Pirtoli
    • 7
    • 8
    • 9
    • 10
    • 11
    • 12
  • Antonella Fioravanti
    • 13
  • Vincenzo Tombolini
    • 1
  • Sara Cheleschi
    • 12
  • Paolo Tini
    • 8
    • 9
    • 11
    • 13
  1. 1.Department of RadiotherapyPoliclinico Umberto I, “Sapienza” University of RomeRomeItaly
  2. 2.Division of Biotechnology, Department of Molecular and Translational MedicineUniversity of BresciaBresciaItaly
  3. 3.Department of Pediatrics“Sapienza” University of RomeRomeItaly
  4. 4.Multi-Factorial Disease and Complex Phenotype Research Area, Bambino Gesù Children’s Hospital, IRCCSRomeItaly
  5. 5.Department of Biotechnological and Applied Clinical SciencesUniversity of L’AquilaL’AquilaItaly
  6. 6.Unit of Radiation TherapyOspedale del MareNaplesItaly
  7. 7.Azienda Ospedaliera Universitaria SeneseSienaItaly
  8. 8.Unit of Radiation OncologyAzienda Ospedaliera Universitaria SeneseSienaItaly
  9. 9.Istituto Toscano TumoriFlorenceItaly
  10. 10.Department of Medicine, Surgery and NeurosciencesUniversity of SienaSienaItaly
  11. 11.Department of Biology, College of Science and TechnologyTemple UniversityPhiladelphiaUSA
  12. 12.Department of Medicine, Surgery and Neuroscience, Rheumatology UnitUniversity of SienaSienaItaly
  13. 13.Sbarro Health Research OrganizationTemple UniversityPhiladelphiaUSA

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