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Colon
Original article
CHK1-targeted therapy to deplete DNA replication-stressed, p53-deficient, hyperdiploid colorectal cancer stem cells
  1. Gwenola Manic1,
  2. Michele Signore2,
  3. Antonella Sistigu3,
  4. Giorgio Russo3,4,
  5. Francesca Corradi1,
  6. Silvia Siteni3,5,
  7. Martina Musella3,6,
  8. Sara Vitale4,
  9. Maria Laura De Angelis2,
  10. Matteo Pallocca7,
  11. Carla Azzurra Amoreo8,
  12. Francesca Sperati9,
  13. Simone Di Franco10,
  14. Sabina Barresi11,
  15. Eleonora Policicchio2,12,
  16. Gabriele De Luca2,
  17. Francesca De Nicola7,
  18. Marcella Mottolese8,
  19. Ann Zeuner2,
  20. Maurizio Fanciulli7,
  21. Giorgio Stassi10,
  22. Marcello Maugeri-Saccà13,
  23. Marta Baiocchi2,
  24. Marco Tartaglia11,
  25. Ilio Vitale1,3,
  26. Ruggero De Maria4
  1. 1Department of Biology, University of Rome “Tor Vergata”, Rome, Italy
  2. 2Department of Oncology and Molecular Medicine, Istituto Superiore di Sanità, Rome, Italy
  3. 3Department of Research, Advanced Diagnostics and Technological Innovation, Regina Elena National Cancer Institute, Rome, Italy
  4. 4Institute of General Pathology, Catholic University and A. Gemelli Polyclinic, Rome, Italy
  5. 5Department of Science, University “Roma Tre”, Rome, Italy
  6. 6Department of Molecular Medicine, University “La Sapienza”, Rome, Italy
  7. 7SAFU, Department of Research, Advanced Diagnostics, and Technological Innovation, Translational Research Area, Regina Elena National Cancer Institute, Rome Italy
  8. 8Department of Pathology, Regina Elena National Cancer Institute, Rome, Italy
  9. 9Biostatistical Unit, Regina Elena National Cancer Institute, Rome, Italy
  10. 10Department of Surgical Oncological and Stomatological Sciences, University of Palermo, Palermo, Italy
  11. 11Genetics and Rare Diseases Research Division, Ospedale Pediatrico “Bambino Gesù”, Rome, Italy
  12. 12Department of Experimental Medicine, University “La Sapienza”, Rome, Italy
  13. 13Division of Medical Oncology 2, Regina Elena National Cancer Institute, Rome, Italy
  1. Correspondence to Dr Ilio Vitale, Department of Biology, University of Rome “Tor Vergata”, via della Ricerca Scientifica 1, Rome 00133, Italy; iliovit{at}gmail.com Professor Ruggero De Maria, Catholic University and Gemelli Polyclinic, Largo Francesco Vito 1, Rome 00168, Italy; ruggero.demaria{at}unicatt.it

Abstract

Objective Cancer stem cells (CSCs) are responsible for tumour formation and spreading, and their targeting is required for tumour eradication. There are limited therapeutic options for advanced colorectal cancer (CRC), particularly for tumours carrying RAS-activating mutations. The aim of this study was to identify novel CSC-targeting strategies.

Design To discover potential therapeutics to be clinically investigated as single agent, we performed a screening with a panel of FDA-approved or investigational drugs on primary CRC cells enriched for CSCs (CRC-SCs) isolated from 27 patients. Candidate predictive biomarkers of efficacy were identified by integrating genomic, reverse-phase protein microarray (RPPA) and cytogenetic analyses, and validated by immunostainings. DNA replication stress (RS) was increased by employing DNA replication-perturbing or polyploidising agents.

Results The drug-library screening led to the identification of LY2606368 as a potent anti-CSC agent acting in vitro and in vivo in tumour cells from a considerable number of patients (∼36%). By inhibiting checkpoint kinase (CHK)1, LY2606368 affected DNA replication in most CRC-SCs, including RAS-mutated ones, forcing them into premature, lethal mitoses. Parallel genomic, RPPA and cytogenetic analyses indicated that CRC-SCs sensitive to LY2606368 displayed signs of ongoing RS response, including the phosphorylation of RPA32 and ataxia telangiectasia mutated serine/threonine kinase (ATM). This was associated with mutation(s) in TP53 and hyperdiploidy, and made these CRC-SCs exquisitely dependent on CHK1 function. Accordingly, experimental increase of RS sensitised resistant CRC-SCs to LY2606368.

Conclusions LY2606368 selectively eliminates replication-stressed, p53-deficient and hyperdiploid CRC-SCs independently of RAS mutational status. These results provide a strong rationale for biomarker-driven clinical trials with LY2606368 in patients with CRC.

  • CELL CYCLE CONTROL
  • CHEMOTHERAPY
  • COLORECTAL CANCER
  • DNA DAMAGE
  • DRUG DEVELOPMENT

This is an Open Access article distributed in accordance with the Creative Commons Attribution Non Commercial (CC BY-NC 4.0) license, which permits others to distribute, remix, adapt, build upon this work non-commercially, and license their derivative works on different terms, provided the original work is properly cited and the use is non-commercial. See: http://creativecommons.org/licenses/by-nc/4.0/

https://doi.org/10.1136/gutjnl-2016-312623

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    Footnotes

    • GM, MS and AS contributed equally. IV and RDM are senior coauthors.

    • Contributors GM, MS, AS, GR designed and performed experiments, analysed and interpreted data; FC, SS, MM, SV, MLDA, CAA, SDF, SB, GDL performed experiments; MP, EP, FDN, MF analysed data and performed bioinformatic studies; FS, MM-S analysed data and performed statistical analysis; MM, AZ, GS, MB, MT designed experiments; IV obtained funding, supervised the project, designed and performed experiments, analysed and interpreted data and wrote the manuscript; RDM obtained funding, supervised the project, designed experiments and wrote the manuscript.

    • Funding This work was supported by the Associazione Italiana per la Ricerca sul Cancro (AIRC, MFAG 2013 grant number 14641 to IV, 5 per Mille grant number 9979 to GS, MT and RDM), Ministero Italiano della Salute (grant number RF_GR-2011-02351355 to IV), Ministero Italiano dell'Istruzione, dell'Universita e della Ricerca (MIUR, Programma per i Giovani Ricercatori ‘Rita Levi Montalcini’ 2010 to IV and Fondo per gli Investimenti della Ricerca di Base, FIRB, grant number RBAP11WCRZ-005 U54 2010 to RDM). AS is supported by AIRC (Start-Up 2016 #18418). GM is supported by AIRC (Triennial Fellowship ‘Antonietta Latronico’ 2014). AZ is supported by AIRC (grant number 15749). SDF is an AIRC fellowship recipient for 2016.

    • Competing interests None declared.

    • Provenance and peer review Not commissioned; externally peer reviewed.