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@ARTICLE{Selt:285655,
      author       = {F. Selt$^*$ and A. El Damaty and M. U. Schuhmann and R.
                      Sigaud$^*$ and J. Ecker$^*$ and P. Sievers$^*$ and D.
                      Kocher$^*$ and C. Herold-Mende and I. Oehme$^*$ and A. von
                      Deimling$^*$ and S. Pfister$^*$ and F. Sahm$^*$ and D.
                      Jones$^*$ and O. Witt$^*$ and T. Milde$^*$},
      title        = {{G}eneration of patient-derived pediatric pilocytic
                      astrocytoma in-vitro models using {SV}40 large {T}:
                      evaluation of a modeling workflow.},
      journal      = {Journal of neuro-oncology},
      volume       = {165},
      number       = {3},
      issn         = {0167-594X},
      address      = {Dordrecht [u.a.]},
      publisher    = {Springer Science + Business Media B.V},
      reportid     = {DKFZ-2023-02478},
      pages        = {467-478},
      year         = {2023},
      note         = {#EA:B310#LA:B310# / 2023 Dec;165(3):467-478},
      abstract     = {Although pediatric low-grade gliomas (pLGG) are the most
                      common pediatric brain tumors, patient-derived cell lines
                      reflecting pLGG biology in culture are scarce. This also
                      applies to the most common pLGG subtype pilocytic
                      astrocytoma (PA). Conventional cell culture approaches
                      adapted from higher-grade tumors fail in PA due to
                      oncogene-induced senescence (OIS) driving tumor cells into
                      arrest. Here, we describe a PA modeling workflow using the
                      Simian Virus large T antigen (SV40-TAg) to circumvent OIS.18
                      pLGG tissue samples (17 $(94\%)$ histological and/or
                      molecular diagnosis PA) were mechanically dissociated. Tumor
                      cell positive-selection using A2B5 was perfomed in 8/18
                      $(44\%)$ cases. All primary cell suspensions were seeded in
                      Neural Stem Cell Medium (NSM) and Astrocyte Basal Medium
                      (ABM). Resulting short-term cultures were infected with
                      SV40-TAg lentivirus. Detection of tumor specific alterations
                      (BRAF-duplication and BRAF V600E-mutation) by digital
                      droplet PCR (ddPCR) at defined time points allowed for
                      determination of tumor cell fraction (TCF) and evaluation of
                      the workflow. DNA-methylation profiling and gene-panel
                      sequencing were used for molecular profiling of primary
                      samples.Primary cell suspensions had a mean TCF of $55\%$
                      (+/- $23\%$ (SD)). No sample in NSM (0/18) and ten samples
                      in ABM (10/18) were successfully transduced. Three of these
                      ten $(30\%)$ converted into long-term pLGG cell lines (TCF
                      $100\%),$ while TCF declined to $0\%$ (outgrowth of
                      microenvironmental cells) in 7/10 $(70\%)$ cultures. Young
                      patient age was associated with successful model
                      establishment.A subset of primary PA cultures can be
                      converted into long-term cell lines using SV40-TAg depending
                      on sample intrinsic (patient age) and extrinsic
                      workflow-related (e.g. type of medium, successful
                      transduction) parameters. Careful monitoring of
                      sample-intrinsic and extrinsic factors optimizes the
                      process.},
      keywords     = {Circumvention of OIS (Other) / In-vitro models (Other) /
                      Inducible SV40 large T (Other) / Pediatric low-grade glioma
                      cell lines (Other) / Pilocytic astrocytoma (Other)},
      cin          = {B310 / B300 / B062 / B360 / HD01},
      ddc          = {610},
      cid          = {I:(DE-He78)B310-20160331 / I:(DE-He78)B300-20160331 /
                      I:(DE-He78)B062-20160331 / I:(DE-He78)B360-20160331 /
                      I:(DE-He78)HD01-20160331},
      pnm          = {312 - Funktionelle und strukturelle Genomforschung
                      (POF4-312)},
      pid          = {G:(DE-HGF)POF4-312},
      typ          = {PUB:(DE-HGF)16},
      pubmed       = {pmid:37999877},
      doi          = {10.1007/s11060-023-04500-6},
      url          = {https://inrepo02.dkfz.de/record/285655},
}