title: The MEK1/2 Inhibitor Pimasertib Enhances Gemcitabine Efficacy in Pancreatic Cancer Models by Altering Ribonucleotide Reductase Subunit-1 (RRM1).
abstract: OBJECTIVE
  Gemcitabine, a nucleoside analogue, is an important treatment for locally advanced and metastatic pancreatic ductal adenocarcinoma (PDAC) but provides only modest survival benefit. Targeting downstream effectors of the RAS/ERK signaling pathway by direct inhibition of MEK1/2 proteins is a promising therapeutic strategy, as aberrant activation of this pathway occurs frequently in PDAC. In this study, the ability of pimasertib, a selective allosteric MEK1/2 inhibitor, to enhance gemcitabine efficacy was tested and the molecular mechanism of their interaction was investigated.
  
  
  METHODS
  Cell survival and apoptosis were assessed by MTT and Caspase 3/7 Glo assays in human pancreatic cancer cell lines. Protein expression was detected by immunoblotting. The in vivo sensitivity of gemcitabine with pimasertib was evaluated in an orthotopic model of pancreatic tumor.
  
  
  RESULTS
  Synergistic activity was observed when gemcitabine was combined sequentially with pimasertib, in human pancreatic cancer cells. In particular, pimasertib reduced ribonucleotide reductase subunit 1 (RRM1) protein, and this was associated with sensitivity to gemcitabine. Pretreatment with MG132 impaired reduction of RRM1 protein induced by pimasertib, suggesting that RRM1 is degraded posttranslationally. Immunoprecipitation indicated enhanced MDM2-mediated polyubiquitination of RRM1 through Lys-48-mediated linkage following pimasertib treatment, an effect mediated, in part, by AKT. Finally, the combination treatment with pimasertib and gemcitabine caused significant tumor growth delays in an orthotopic pancreatic cancer model, with RRM1 downregulation in pimasertib-treated mice.
  
  
  CONCLUSIONS
  These results confirm an important role of RRM1 in gemcitabine response and indicate MEK as a potential target to sensitize gemcitabine therapy for PDAC. Clin Cancer Res; 21(24); 5563-77. ©2015 AACR.
authors.affiliation: Cancer Research UK Drug-DNA Interactions Research Group, UCL Cancer Institute, University College London, London, United Kingdom.
authors.forename: Francesca
authors.initials: F
authors.lastname: Vena
authors.affiliation: Barts Cancer Institute, University of London, London, United Kingdom.
authors.forename: Eleonora
authors.initials: E
authors.lastname: Li Causi
authors.affiliation: Department of Research Pathology, UCL Cancer Institute, London, United Kingdom.
authors.forename: Manuel
authors.initials: M
authors.lastname: Rodriguez-Justo
authors.affiliation: EMD Serono Research and Development Institute, Billerica, Massachusetts.
authors.forename: Samantha
authors.initials: S
authors.lastname: Goodstal
authors.affiliation: Barts Cancer Institute, University of London, London, United Kingdom.
authors.forename: Thorsten
authors.initials: T
authors.lastname: Hagemann
authors.affiliation: Cancer Research UK Drug-DNA Interactions Research Group, UCL Cancer Institute, University College London, London, United Kingdom.
authors.forename: John A
authors.initials: JA
authors.lastname: Hartley
authors.affiliation: Cancer Research UK Drug-DNA Interactions Research Group, UCL Cancer Institute, University College London, London, United Kingdom. d.hochhauser@ucl.ac.uk.
authors.forename: Daniel
authors.initials: D
authors.lastname: Hochhauser
mesh_terms: D000818:Animals; D000970:Antineoplastic Agents / Q000494:pharmacology*; D017209:Apoptosis / Q000187:drug effects; D020169:Caspases / Q000378:metabolism; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects; D003841:Deoxycytidine / Q000031:analogs & derivatives* / Q000494:pharmacology; D004195:Disease Models, Animal; D019008:Drug Resistance, Neoplasm / Q000235:genetics; D004357:Drug Synergism; D005260:Female; D055785:Gene Knockdown Techniques; D006801:Humans; D048369:MAP Kinase Kinase 1 / Q000037:antagonists & inhibitors; D048370:MAP Kinase Kinase 2 / Q000037:antagonists & inhibitors; D051379:Mice; D009536:Niacinamide / Q000031:analogs & derivatives* / Q000494:pharmacology; D010190:Pancreatic Neoplasms / Q000188:drug therapy / Q000235:genetics / Q000378:metabolism* / Q000473:pathology*; D047428:Protein Kinase Inhibitors / Q000494:pharmacology*; D059748:Proteolysis; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism; D051736:Proto-Oncogene Proteins c-mdm2 / Q000378:metabolism; D034741:RNA, Small Interfering / Q000235:genetics; D012262:Ribonucleoside Diphosphate Reductase; D014185:Transplantation, Isogeneic; D047368:Tumor Burden / Q000187:drug effects; D025521:Tumor Suppressor Proteins / Q000235:genetics / Q000378:metabolism*; D054875:Ubiquitination; D000093542:Gemcitabine
chemical_list: D000970:Antineoplastic Agents; C550600:N-(2,3-dihydroxypropyl)-1-((2-fluoro-4-iodophenyl)amino)isonicotinamide; D047428:Protein Kinase Inhibitors; D034741:RNA, Small Interfering; D025521:Tumor Suppressor Proteins; D003841:Deoxycytidine; D009536:Niacinamide; C475286:RRM1 protein, human; D012262:Ribonucleoside Diphosphate Reductase; D051736:Proto-Oncogene Proteins c-mdm2; C482128:MAP2K2 protein, human; D051057:Proto-Oncogene Proteins c-akt; D048369:MAP Kinase Kinase 1; D048370:MAP Kinase Kinase 2; C482125:MAP2K1 protein, human; D020169:Caspases; D000093542:Gemcitabine