@article{RydzekNerreterPengetal.2019,
  author    = {Rydzek, Julian and Nerreter, Thomas and Peng, Haiyong and Jutz, Sabrina and Leitner, Judith and Steinberger, Peter and Einsele, Hermann and Rader, Christoph and Hudecek, Michael},
  title     = {Chimeric Antigen Receptor Library Screening Using a Novel NF-kappa B/NFAT Reporter Cell Platform},
  series = {Molecular Therapy},
  volume    = {27},
  journal   = {Molecular Therapy},
  number    = {2},
  doi       = {10.1016/j.ymthe.2018.11.015},
  url       = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-227193},
  pages     = {287-299},
  year      = {2019},
  abstract  = {Chimeric antigen receptor (CAR)-T cell immunotherapy is under intense preclinical and clinical investigation, and it involves a rapidly increasing portfolio of novel target antigens and CAR designs. We established a platform that enables rapid and high-throughput CAR-screening campaigns with reporter cells derived from the T cell lymphoma line Jurkat. Reporter cells were equipped with nuclear factor kappa B (NF kappa B) and nuclear factor of activated T cells (NFAT) reporter genes that generate a duplex output of enhanced CFP (ECFP) and EGFP, respectively. As a proof of concept, we modified reporter cells with CD19-specific and ROR1-specific CARs, and we detected high-level reporter signals that allowed distinguishing functional from non-functional CAR constructs. The reporter data were highly reproducible, and the time required for completing each testing campaign was substantially shorter with reporter cells (6 days) compared to primary CAR-T cells (21 days). We challenged the reporter platform to a large-scale screening campaign on a ROR1-CAR library, and we showed that reporter cells retrieved a functional CAR variant that was present with a frequency of only 6 in 1.05 x 10(6). The data illustrate the potential to implement this reporter platform into the preclinical development path of novel CAR-T cell products and to inform and accelerate the selection of lead CAR candidates for clinical translation.},
  language  = {en}
}
@article{NerreterLetschertGoetzetal.2019,
  author    = {Nerreter, Thomas and Letschert, Sebastian and G{\"o}tz, Ralph and Doose, S{\"o}ren and Danhof, Sophia and Einsele, Hermann and Sauer, Markus and Hudecek, Michael},
  title     = {Super-resolution microscopy reveals ultra-low CD19 expression on myeloma cells that triggers elimination by CD19 CAR-T},
  series = {Nature Communications},
  volume    = {10},
  journal   = {Nature Communications},
  doi       = {10.1038/s41467-019-10948-w},
  url       = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-232258},
  year      = {2019},
  abstract  = {Immunotherapy with chimeric antigen receptor-engineered T-cells (CAR-T) is under investigation in multiple myeloma. There are reports of myeloma remission after CD19 CAR-T therapy, although CD19 is hardly detectable on myeloma cells by flow cytometry (FC). We apply single molecule-sensitive direct stochastic optical reconstruction microscopy (dSTORM), and demonstrate CD19 expression on a fraction of myeloma cells (10.3-80\%) in 10 out of 14 patients (density: 13-5,000 molecules per cell). In contrast, FC detects CD19 in only 2 of these 10 patients, on a smaller fraction of cells. Treatment with CD19 CAR-T in vitro results in elimination of CD19-positive myeloma cells, including those with <100 CD19 molecules per cell. Similar data are obtained by dSTORM analyses of CD20 expression on myeloma cells and CD20 CAR-T. These data establish a sensitivity threshold for CAR-T and illustrate how super-resolution microscopy can guide patient selection in immunotherapy to exploit ultra-low density antigens.},
  language  = {en}
}