title: CHK1-targeted therapy to deplete DNA replication-stressed, p53-deficient, hyperdiploid colorectal cancer stem cells.
abstract: 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.
  
  
  
  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.
  
  
  
  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) inTP53and hyperdiploidy, and made these CRC-SCs exquisitely dependent on CHK1 function. Accordingly, experimental increase of RS sensitised resistant CRC-SCs to LY2606368.
  
  
  
  LY2606368 selectively eliminates replication-stressed, p53-deficient and hyperdiploid CRC-SCs independently ofRASmutational status. These results provide a strong rationale for biomarker-driven clinical trials with LY2606368 in patients with CRC.
authors.affiliation: Department of Biology, University of Rome "Tor Vergata", Rome, Italy.
authors.forename: Gwenola
authors.initials: G
authors.lastname: Manic
authors.affiliation: Department of Oncology and Molecular Medicine, Istituto Superiore di Sanità, Rome, Italy.
authors.forename: Michele
authors.initials: M
authors.lastname: Signore
authors.affiliation: Department of Research, Advanced Diagnostics and Technological Innovation, Regina Elena National Cancer Institute, Rome, Italy.
authors.forename: Antonella
authors.initials: A
authors.lastname: Sistigu
authors.affiliation: Department of Research, Advanced Diagnostics and Technological Innovation, Regina Elena National Cancer Institute, Rome, Italy.|Institute of General Pathology, Catholic University and A. Gemelli Polyclinic, Rome, Italy.
authors.forename: Giorgio
authors.initials: G
authors.lastname: Russo
authors.affiliation: Department of Biology, University of Rome "Tor Vergata", Rome, Italy.
authors.forename: Francesca
authors.initials: F
authors.lastname: Corradi
authors.affiliation: Department of Research, Advanced Diagnostics and Technological Innovation, Regina Elena National Cancer Institute, Rome, Italy.|Department of Science, University "Roma Tre", Rome, Italy.
authors.forename: Silvia
authors.initials: S
authors.lastname: Siteni
authors.affiliation: Department of Research, Advanced Diagnostics and Technological Innovation, Regina Elena National Cancer Institute, Rome, Italy.|Department of Molecular Medicine, University "La Sapienza", Rome, Italy.
authors.forename: Martina
authors.initials: M
authors.lastname: Musella
authors.affiliation: Institute of General Pathology, Catholic University and A. Gemelli Polyclinic, Rome, Italy.
authors.forename: Sara
authors.initials: S
authors.lastname: Vitale
authors.affiliation: Department of Oncology and Molecular Medicine, Istituto Superiore di Sanità, Rome, Italy.
authors.forename: Maria Laura
authors.initials: ML
authors.lastname: De Angelis
authors.affiliation: SAFU, Department of Research, Advanced Diagnostics, and Technological Innovation, Translational Research Area, Regina Elena National Cancer Institute, Rome Italy.
authors.forename: Matteo
authors.initials: M
authors.lastname: Pallocca
authors.affiliation: Department of Pathology, Regina Elena National Cancer Institute, Rome, Italy.
authors.forename: Carla Azzurra
authors.initials: CA
authors.lastname: Amoreo
authors.affiliation: Biostatistical Unit, Regina Elena National Cancer Institute, Rome, Italy.
authors.forename: Francesca
authors.initials: F
authors.lastname: Sperati
authors.affiliation: Department of Surgical Oncological and Stomatological Sciences, University of Palermo, Palermo, Italy.
authors.forename: Simone
authors.initials: S
authors.lastname: Di Franco
authors.affiliation: Genetics and Rare Diseases Research Division, Ospedale Pediatrico "Bambino Gesù", Rome, Italy.
authors.forename: Sabina
authors.initials: S
authors.lastname: Barresi
authors.affiliation: Department of Oncology and Molecular Medicine, Istituto Superiore di Sanità, Rome, Italy.|Department of Experimental Medicine, University "La Sapienza", Rome, Italy.
authors.forename: Eleonora
authors.initials: E
authors.lastname: Policicchio
authors.affiliation: Department of Oncology and Molecular Medicine, Istituto Superiore di Sanità, Rome, Italy.
authors.forename: Gabriele
authors.initials: G
authors.lastname: De Luca
authors.affiliation: SAFU, Department of Research, Advanced Diagnostics, and Technological Innovation, Translational Research Area, Regina Elena National Cancer Institute, Rome Italy.
authors.forename: Francesca
authors.initials: F
authors.lastname: De Nicola
authors.affiliation: Department of Pathology, Regina Elena National Cancer Institute, Rome, Italy.
authors.forename: Marcella
authors.initials: M
authors.lastname: Mottolese
authors.affiliation: Department of Oncology and Molecular Medicine, Istituto Superiore di Sanità, Rome, Italy.
authors.forename: Ann
authors.initials: A
authors.lastname: Zeuner
authors.affiliation: SAFU, Department of Research, Advanced Diagnostics, and Technological Innovation, Translational Research Area, Regina Elena National Cancer Institute, Rome Italy.
authors.forename: Maurizio
authors.initials: M
authors.lastname: Fanciulli
authors.affiliation: Department of Surgical Oncological and Stomatological Sciences, University of Palermo, Palermo, Italy.
authors.forename: Giorgio
authors.initials: G
authors.lastname: Stassi
authors.affiliation: Division of Medical Oncology 2, Regina Elena National Cancer Institute, Rome, Italy.
authors.forename: Marcello
authors.initials: M
authors.lastname: Maugeri-Saccà
authors.affiliation: Department of Oncology and Molecular Medicine, Istituto Superiore di Sanità, Rome, Italy.
authors.forename: Marta
authors.initials: M
authors.lastname: Baiocchi
authors.affiliation: Genetics and Rare Diseases Research Division, Ospedale Pediatrico "Bambino Gesù", Rome, Italy.
authors.forename: Marco
authors.initials: M
authors.lastname: Tartaglia
authors.affiliation: Department of Biology, University of Rome "Tor Vergata", Rome, Italy.|Department of Research, Advanced Diagnostics and Technological Innovation, Regina Elena National Cancer Institute, Rome, Italy.
authors.forename: Ilio
authors.initials: I
authors.lastname: Vitale
authors.affiliation: Institute of General Pathology, Catholic University and A. Gemelli Polyclinic, Rome, Italy.
authors.forename: Ruggero
authors.initials: R
authors.lastname: De Maria
mesh_terms: D000970:Antineoplastic Agents / Q000494:pharmacology*; D045744:Cell Line, Tumor; D000071877:Checkpoint Kinase 1 / Q000187:drug effects* / Q000235:genetics; D015179:Colorectal Neoplasms / Q000188:drug therapy* / Q000235:genetics; D004261:DNA Replication / Q000187:drug effects; D006801:Humans; D007150:Immunohistochemistry; D009154:Mutation; D014411:Neoplastic Stem Cells / Q000187:drug effects* / Q000378:metabolism; D020411:Oligonucleotide Array Sequence Analysis; D011719:Pyrazines / Q000494:pharmacology*; D011720:Pyrazoles / Q000494:pharmacology*; D016159:Tumor Suppressor Protein p53 / Q000235:genetics
chemical_list: D000970:Antineoplastic Agents; D011719:Pyrazines; D011720:Pyrazoles; D016159:Tumor Suppressor Protein p53; C000608121:prexasertib; C000605867:CHEK1 protein, human; D000071877:Checkpoint Kinase 1
keywords: CELL CYCLE CONTROL; CHEMOTHERAPY; COLORECTAL CANCER; DNA DAMAGE; DRUG DEVELOPMENT