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@ARTICLE{Usta:156979,
      author       = {D. Usta$^*$ and R. Sigaud$^*$ and J. L. Buhl$^*$ and F.
                      Selt$^*$ and V. Marquardt$^*$ and D. Pauck$^*$ and J. Jansen
                      and S. Pusch$^*$ and J. Ecker$^*$ and T. Hielscher$^*$ and
                      J. Vollmer$^*$ and A. C. Sommerkamp$^*$ and T. Rubner$^*$
                      and D. Hargrave and C. M. van Tilburg$^*$ and S. M.
                      Pfister$^*$ and D. T. W. Jones$^*$ and M. Remke$^*$ and T.
                      Brummer and O. Witt$^*$ and T. Milde$^*$},
      title        = {{A} cell-based {MAPK} reporter assay reveals synergistic
                      {MAPK} pathway activity suppression by {MAPK} inhibitor
                      combination in {BRAF}-driven pediatric low-grade glioma
                      cells.},
      journal      = {Molecular cancer therapeutics},
      volume       = {19},
      number       = {8},
      issn         = {1538-8514},
      address      = {Philadelphia, Pa.},
      publisher    = {AACR},
      reportid     = {DKFZ-2020-01284},
      pages        = {1736-1750},
      year         = {2020},
      note         = {2020 Aug;19(8):1736-1750#EA:B310#LA:B310#},
      abstract     = {Pilocytic astrocytomas (PAs) as well as other pediatric
                      low-grade gliomas (pLGGs) exhibit genetic events leading to
                      aberrant activation of the MAPK pathway. The most common
                      alterations are KIAA1549:BRAF fusions, BRAFV600E and NF1
                      mutations. Novel drugs targeting the MAPK pathway (MAPKi)
                      are prime candidates for the treatment of these
                      single-pathway diseases. We aimed to develop an assay
                      suitable for pre-clinical testing of MAPKi in pLGGs with the
                      goal to identify novel MAPK pathway suppressing synergistic
                      drug combinations. A reporter plasmid (pDIPZ) with a
                      MAPK-responsive ELK-1-binding element driving the expression
                      of destabilized firefly luciferase was generated and
                      packaged using a lentiviral vector system. Pediatric glioma
                      cell lines with a BRAF fusion (DKFZ-BT66) and a BRAFV600E
                      mutation (BT-40) background, respectively, were stably
                      transfected. Modulation of the MAPK pathway activity by
                      MAPKi was measured using the luciferase reporter and
                      validated by detection of phosphorylated protein levels. A
                      screen of a MAPKi library was performed and synergy of
                      selected combinations was calculated. Screening of a MAPKi
                      library revealed MEK inhibitors as the class inhibiting the
                      pathway with the lowest IC50s, followed by ERK and
                      next-generation RAF inhibitors. Combination treatments with
                      different MAPKi classes showed synergistic effects in BRAF
                      fusion as well as BRAFV600E mutation backgrounds. We here
                      report a novel reporter assay for medium- to high-throughput
                      pre-clinical drug testing in pLGG cell lines. The assay
                      confirmed MEK, ERK and next-generation RAF inhibitors as
                      potential treatment approaches for KIAA1549:BRAF and
                      BRAFV600E mutated pLGGs. In addition, the assay revealed
                      that combination treatments synergistically suppressed MAPK
                      pathway activity.},
      cin          = {B310 / HD01 / ED01 / FR01 / B300 / C060 / B360 / W220 /
                      B062},
      ddc          = {570},
      cid          = {I:(DE-He78)B310-20160331 / I:(DE-He78)HD01-20160331 /
                      I:(DE-He78)ED01-20160331 / I:(DE-He78)FR01-20160331 /
                      I:(DE-He78)B300-20160331 / I:(DE-He78)C060-20160331 /
                      I:(DE-He78)B360-20160331 / I:(DE-He78)W220-20160331 /
                      I:(DE-He78)B062-20160331},
      pnm          = {312 - Functional and structural genomics (POF3-312)},
      pid          = {G:(DE-HGF)POF3-312},
      typ          = {PUB:(DE-HGF)16},
      pubmed       = {pmid:32451331},
      doi          = {10.1158/1535-7163.MCT-19-1021},
      url          = {https://inrepo02.dkfz.de/record/156979},
}