title: mTORC1 and DNA-PKcs as novel molecular determinants of sensitivity to Chk1 inhibition.
abstract: BACKGROUND
  Chk1 inhibitors are currently under clinical evaluation as single agents and in combination with cytotoxic chemotherapy. Understanding determinants of sensitivity and novel combinations is critical for further clinical development.
  
  
  METHODS
  Potentiation of mTOR inhibitor cytotoxicity by the Chk1 inhibitor V158411 was determined in p53 mutant colon cancer cells. DNA damage response, expression levels of repair proteins, cell cycle effects and the contribution of alternative DSB repair pathways were further evaluated by western blotting and high content analysis.
  
  
  RESULTS
  mTOR inhibitors AZD8055, RAD-001, rapamycin and BEZ235 induced synergistic cytotoxicity with the Chk1 inhibitor V158411 in p53 mutant colon cancer cells. Reduced FANCD2, RAD51 and RPA70, core proteins in homologous recombination repair (HRR) and interstrand crosslink repair (ICLR), following inhibition of mTOR was associated with increased V158411 induced DSBs and caspase 3-independent cell death. Dual mTOR and Chk1 inhibition activated DNA-PKcs. Cells defective in DNA-PKcs exhibited increased resistance to V158411 with Chk1 expression closely correlated to DNA-PKcs expression in various types of cancer.
  
  
  CONCLUSIONS
  Down regulation of proteins involved in HRR or ICLR by mTOR inhibitors is associated with increased sensitivity of human tumours to Chk1 inhibitors such as V158411. High levels of DNA-PKcs may be a potential biomarker to stratify patients to Chk1 inhibitor therapy alone or in combination with mTOR inhibitors.
authors.affiliation: Vernalis Research, Granta Park, Cambridge, CB21 6GB, UK. Electronic address: a.massey@vernalis.com.
authors.forename: Andrew J
authors.initials: AJ
authors.lastname: Massey
authors.affiliation: Newcastle University, Northern Institute for Cancer Research, Newcastle upon Tyne, NE2 4HH, UK.
authors.forename: Peter
authors.initials: P
authors.lastname: Stephens
authors.affiliation: Vernalis Research, Granta Park, Cambridge, CB21 6GB, UK.
authors.forename: Rebecca
authors.initials: R
authors.lastname: Rawlinson
authors.affiliation: Newcastle University, Northern Institute for Cancer Research, Newcastle upon Tyne, NE2 4HH, UK.
authors.forename: Lauren
authors.initials: L
authors.lastname: McGurk
authors.affiliation: Newcastle University, Northern Institute for Cancer Research, Newcastle upon Tyne, NE2 4HH, UK.
authors.forename: Ruth
authors.initials: R
authors.lastname: Plummer
authors.affiliation: Newcastle University, Northern Institute for Cancer Research, Newcastle upon Tyne, NE2 4HH, UK. Electronic address: nicola.curtin@ncl.ac.uk.
authors.forename: Nicola J
authors.initials: NJ
authors.lastname: Curtin
mesh_terms: D045744:Cell Line, Tumor; D000071877:Checkpoint Kinase 1; D003110:Colonic Neoplasms / Q000235:genetics / Q000378:metabolism / Q000473:pathology; D004249:DNA Damage; D059766:DNA End-Joining Repair; D051747:DNA-Activated Protein Kinase / Q000378:metabolism*; D006801:Humans; D000076222:Mechanistic Target of Rapamycin Complex 1; D046912:Multiprotein Complexes / Q000037:antagonists & inhibitors / Q000378:metabolism*; D047428:Protein Kinase Inhibitors / Q000494:pharmacology*; D011494:Protein Kinases / Q000187:drug effects*; D015398:Signal Transduction; D058570:TOR Serine-Threonine Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism*
chemical_list: D046912:Multiprotein Complexes; D047428:Protein Kinase Inhibitors; D011494:Protein Kinases; C000605867:CHEK1 protein, human; D000071877:Checkpoint Kinase 1; D051747:DNA-Activated Protein Kinase; D000076222:Mechanistic Target of Rapamycin Complex 1; D058570:TOR Serine-Threonine Kinases
keywords: Chk1; DNA-PKcs; Double strand break repair; V158411; mTOR