title: Dual treatment with shikonin and temozolomide reduces glioblastoma tumor growth, migration and glial-to-mesenchymal transition.
abstract: OBJECTIVE
  Glioblastomas (GBM) comprise 17% of all primary brain tumors. These tumors are extremely aggressive due to their infiltrative capacity and chemoresistance, with glial-to-mesenchymal transition (GMT) proteins playing a prominent role in tumor invasion. One compound that has recently been used to reduce the expression of these proteins is shikonin (SHK), a naphthoquinone with anti-tumor properties. Temozolomide (TMZ), the most commonly used chemotherapeutic agent in GBM treatment, has so far not been studied in combination with SHK. Here, we investigated the combined effects of these two drugs on the proliferation and motility of GBM-derived cells.
  
  
  METHODS
  The cytotoxic and proliferative effects of SHK and TMZ on human GBM-derived cells were tested using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT), Ki67 staining and BrdU incorporation assays. The migration capacities of these cells were evaluated using a scratch wound assay. The expression levels of β3 integrin, metalloproteinases (MMPs) and GMT-associated proteins were determined by Western blotting and immunocytochemistry.
  
  
  RESULTS
  We found that GBM-derived cells treated with a combination of SHK and TMZ showed decreases in their proliferation and migration capacities. These decreases were followed by the suppression of GMT through a reduction of β3 integrin, MMP-2, MMP-9, Slug and vimentin expression via inactivation of PI3K/AKT signaling.
  
  
  CONCLUSIONS
  From our results we conclude that dual treatment with SHK and TMZ may constitute a powerful new tool for GBM treatment by reducing therapy resistance and tumor recurrence.
authors.affiliation: Brain's Biomedicine Laboratory, Paulo Niemeyer State Brain Institute, Secretaria de Estado de Saúde do Rio de Janeiro, Rua do Resende 156, Rio de Janeiro, 20231-092, Rio de Janeiro, Brazil.|Institute of Biomedical Sciences at Federal University of Rio de Janeiro (ICB/UFRJ), Rio de Janeiro, 21941-902, Brazil.
authors.forename: Diana
authors.initials: D
authors.lastname: Matias
authors.affiliation: Brain's Biomedicine Laboratory, Paulo Niemeyer State Brain Institute, Secretaria de Estado de Saúde do Rio de Janeiro, Rua do Resende 156, Rio de Janeiro, 20231-092, Rio de Janeiro, Brazil.|Center for Neuroscience and Cell Biology and Institute for Biomedical Imaging and Life Sciences (CNC.IBILI), Rua Larga Faculdade de Medicina, Pólo I, 1° andar, 3004-504, Coimbra, Portugal.|Faculty of Medicine at University of Coimbra (FMUC), Pólo III - Pólo das Ciências da Saúde, Azinhaga de Santa Comba, 3000-354, Coimbra, Portugal.
authors.forename: Joana
authors.initials: J
authors.lastname: Balça-Silva
authors.affiliation: Brain's Biomedicine Laboratory, Paulo Niemeyer State Brain Institute, Secretaria de Estado de Saúde do Rio de Janeiro, Rua do Resende 156, Rio de Janeiro, 20231-092, Rio de Janeiro, Brazil.
authors.forename: Luiz Gustavo
authors.initials: LG
authors.lastname: Dubois
authors.affiliation: Institute of Biomedical Sciences at Federal University of Rio de Janeiro (ICB/UFRJ), Rio de Janeiro, 21941-902, Brazil.
authors.forename: Bruno
authors.initials: B
authors.lastname: Pontes
authors.affiliation: Brain's Biomedicine Laboratory, Paulo Niemeyer State Brain Institute, Secretaria de Estado de Saúde do Rio de Janeiro, Rua do Resende 156, Rio de Janeiro, 20231-092, Rio de Janeiro, Brazil.
authors.forename: Valéria Pereira
authors.initials: VP
authors.lastname: Ferrer
authors.affiliation: Brain's Biomedicine Laboratory, Paulo Niemeyer State Brain Institute, Secretaria de Estado de Saúde do Rio de Janeiro, Rua do Resende 156, Rio de Janeiro, 20231-092, Rio de Janeiro, Brazil.
authors.forename: Luciane
authors.initials: L
authors.lastname: Rosário
authors.affiliation: Center for Neuroscience and Cell Biology and Institute for Biomedical Imaging and Life Sciences (CNC.IBILI), Rua Larga Faculdade de Medicina, Pólo I, 1° andar, 3004-504, Coimbra, Portugal.|Hospital Center and University of Coimbra (CHUC), Praceta Prof. Mota Pinto, 3000-075, Coimbra, Portugal.
authors.forename: Anália
authors.initials: A
authors.lastname: do Carmo
authors.affiliation: Paulo de Góes Institute of Microbiology, Federal University of Rio de Janeiro, Rio de Janeiro, 21941-902, Brazil.
authors.forename: Juliana
authors.initials: J
authors.lastname: Echevarria-Lima
authors.affiliation: Center for Neuroscience and Cell Biology and Institute for Biomedical Imaging and Life Sciences (CNC.IBILI), Rua Larga Faculdade de Medicina, Pólo I, 1° andar, 3004-504, Coimbra, Portugal.|Faculty of Medicine at University of Coimbra (FMUC), Pólo III - Pólo das Ciências da Saúde, Azinhaga de Santa Comba, 3000-354, Coimbra, Portugal.|Hospital Center and University of Coimbra (CHUC), Praceta Prof. Mota Pinto, 3000-075, Coimbra, Portugal.
authors.forename: Ana Bela
authors.initials: AB
authors.lastname: Sarmento-Ribeiro
authors.affiliation: Center for Neuroscience and Cell Biology and Institute for Biomedical Imaging and Life Sciences (CNC.IBILI), Rua Larga Faculdade de Medicina, Pólo I, 1° andar, 3004-504, Coimbra, Portugal.|Faculty of Pharmacy at University of Coimbra, Pólo das Ciências da Saúde, Azinhaga de Santa Comba, 3000-548, Coimbra, Portugal.
authors.forename: Maria Celeste
authors.initials: MC
authors.lastname: Lopes
authors.affiliation: Brain's Biomedicine Laboratory, Paulo Niemeyer State Brain Institute, Secretaria de Estado de Saúde do Rio de Janeiro, Rua do Resende 156, Rio de Janeiro, 20231-092, Rio de Janeiro, Brazil. vivaldomouraneto@gmail.com.
authors.forename: Vivaldo
authors.initials: V
authors.lastname: Moura-Neto
mesh_terms: D000971:Antineoplastic Combined Chemotherapy Protocols / Q000494:pharmacology*; D001932:Brain Neoplasms / Q000473:pathology*; D045744:Cell Line, Tumor; D002465:Cell Movement / Q000187:drug effects; D049109:Cell Proliferation / Q000187:drug effects; D003606:Dacarbazine / Q000031:analogs & derivatives* / Q000494:pharmacology; D019008:Drug Resistance, Neoplasm / Q000187:drug effects; D058750:Epithelial-Mesenchymal Transition / Q000187:drug effects; D005909:Glioblastoma / Q000473:pathology*; D006801:Humans; D009285:Naphthoquinones / Q000494:pharmacology*; D000077204:Temozolomide
chemical_list: D009285:Naphthoquinones; C016101:shikonin; D003606:Dacarbazine; D000077204:Temozolomide
keywords: Glial-to-mesenchymal transition; Glioblastoma; Migration; Shikonin; Temozolomide