title: Dual blockade of the PI3K/AKT/mTOR (AZD8055) and RAS/MEK/ERK (AZD6244) pathways synergistically inhibits rhabdomyosarcoma cell growth in vitro and in vivo.
abstract: OBJECTIVE
  To provide rationale for using phosphoinositide 3-kinase (PI3K) and/or mitogen-activated protein kinase (MAPK) pathway inhibitors to treat rhabdomyosarcomas, a major cause of pediatric and adolescent cancer deaths.
  
  
  METHODS
  The prevalence of PI3K/MAPK pathway activation in rhabdomyosarcoma clinical samples was assessed using immunohistochemistry. Compensatory signaling and cross-talk between PI3K/MAPK pathways was determined in rhabdomyosarcoma cell lines following p110α short hairpin RNA-mediated depletion. Pharmacologic inhibition of reprogrammed signaling in stable p110α knockdown lines was used to determine the target-inhibition profile inducing maximal growth inhibition. The in vitro and in vivo efficacy of inhibitors of TORC1/2 (AZD8055), MEK (AZD6244), and P13K/mTOR (NVP-BEZ235) was evaluated alone and in pairwise combinations.
  
  
  RESULTS
  PI3K pathway activation was seen in 82.5% rhabdomyosarcomas with coactivated MAPK in 36% and 46% of alveolar and embryonal subtypes, respectively. p110α knockdown in cell lines over the short and long term was associated with compensatory expression of other p110 isoforms, activation of the MAPK pathway, and cross-talk to reactivate the PI3K pathway. Combinations of PI3K pathway and MAP-ERK kinase (MEK) inhibitors synergistically inhibited cell growth in vitro. Treatment of RD cells with AZD8055 plus AZD6244 blocked reciprocal pathway activation, as evidenced by reduced AKT/ERK/S6 phosphorylation. In vivo, the synergistic effect on growth and changes in pharmacodynamic biomarkers was recapitulated using the AZD8055/AZD6244 combination but not NVP-BEZ235/AZD6244. Pharmacokinetic analysis provided evidence of drug-drug interaction with both combinations.
  
  
  CONCLUSIONS
  Dual PI3K/MAPK pathway activation and compensatory signaling in both rhabdomyosarcoma subtypes predict a lack of clinical efficacy for single agents targeting either pathway, supporting a therapeutic strategy combining a TORC1/2 with a MEK inhibitor.
authors.affiliation: Authors' Affiliations: Divisions of Clinical Studies, Cancer Therapeutics, and Molecular Pathology, The Institute of Cancer Research, Sutton, Surrey; Histopathology Department, The Royal Marsden Hospital NHS Foundation Trust, London, United Kingdom; and Department of Neuropathology, University of Bonn, Bonn, Germany.
authors.forename: Jane
authors.initials: J
authors.lastname: Renshaw
authors.forename: Kathryn R
authors.initials: KR
authors.lastname: Taylor
authors.forename: Ryan
authors.initials: R
authors.lastname: Bishop
authors.forename: Melanie
authors.initials: M
authors.lastname: Valenti
authors.forename: Alexis
authors.initials: A
authors.lastname: De Haven Brandon
authors.forename: Sharon
authors.initials: S
authors.lastname: Gowan
authors.forename: Suzanne A
authors.initials: SA
authors.lastname: Eccles
authors.forename: Ruth R
authors.initials: RR
authors.lastname: Ruddle
authors.forename: Louise D
authors.initials: LD
authors.lastname: Johnson
authors.forename: Florence I
authors.initials: FI
authors.lastname: Raynaud
authors.forename: Joanna L
authors.initials: JL
authors.lastname: Selfe
authors.forename: Khin
authors.initials: K
authors.lastname: Thway
authors.forename: Torsten
authors.initials: T
authors.lastname: Pietsch
authors.forename: Andrew D
authors.initials: AD
authors.lastname: Pearson
authors.forename: Janet
authors.initials: J
authors.lastname: Shipley
mesh_terms: D000818:Animals; D000970:Antineoplastic Agents / Q000008:administration & dosage / Q000494:pharmacology; D001562:Benzimidazoles / Q000008:administration & dosage / Q000494:pharmacology; D045744:Cell Line, Tumor; D058534:Class I Phosphatidylinositol 3-Kinases; D004195:Disease Models, Animal; D004357:Drug Synergism; D004789:Enzyme Activation / Q000187:drug effects; D005260:Female; D055785:Gene Knockdown Techniques; D006801:Humans; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism*; D009025:Morpholines / Q000008:administration & dosage / Q000494:pharmacology; D019869:Phosphatidylinositol 3-Kinases / Q000235:genetics / Q000378:metabolism*; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism*; D016283:Proto-Oncogene Proteins p21(ras) / Q000378:metabolism*; D012208:Rhabdomyosarcoma / Q000188:drug therapy / Q000378:metabolism* / Q000473:pathology*; D015398:Signal Transduction / Q000187:drug effects; D058570:TOR Serine-Threonine Kinases / Q000378:metabolism*; D023041:Xenograft Model Antitumor Assays
chemical_list: D000970:Antineoplastic Agents; D001562:Benzimidazoles; D058534:Class I Phosphatidylinositol 3-Kinases; D020928:Mitogen-Activated Protein Kinases; D009025:Morpholines; D019869:Phosphatidylinositol 3-Kinases; D051057:Proto-Oncogene Proteins c-akt; D016283:Proto-Oncogene Proteins p21(ras); D058570:TOR Serine-Threonine Kinases; C517975:AZD 6244; C546624:(5-(2,4-bis((3S)-3-methylmorpholin-4-yl)pyrido(2,3-d)pyrimidin-7-yl)-2-methoxyphenyl)methanol; C484760:PIK3CA protein, human
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