Cortés-López, MarielaSchulz, LauraEnculescu, MihaelaParet, ClaudiaSpiekermann, BeaQuesnel-Vallières, MathieuTorres-Diz, ManuelUnic, SebastianBusch, AnkeOrekhova, AnnaKuban, MonikaMesitov, MikhailMulorz, Miriam M.Shraim, RawanKielisch, FridolinFaber, JörgBarash, YosephThomas-Tikhonenko, AndreiZarnack, KathiLegewie, StefanKönig, Julian2025-04-1120222041-17231926576152http://nbn-resolving.de/urn:nbn:de:bsz:93-opus-ds-161860https://elib.uni-stuttgart.de/handle/11682/16186https://doi.org/10.18419/opus-16167Following CART-19 immunotherapy for B-cell acute lymphoblastic leukaemia (B-ALL), many patients relapse due to loss of the cognate CD19 epitope. Since epitope loss can be caused by aberrant CD19 exon 2 processing, we herein investigate the regulatory code that controls CD19 splicing. We combine high-throughput mutagenesis with mathematical modelling to quantitatively disentangle the effects of all mutations in the region comprising CD19 exons 1-3. Thereupon, we identify ~200 single point mutations that alter CD19 splicing and thus could predispose B-ALL patients to developing CART-19 resistance. Furthermore, we report almost 100 previously unknown splice isoforms that emerge from cryptic splice sites and likely encode non-functional CD19 proteins. We further identify cis-regulatory elements and trans-acting RNA-binding proteins that control CD19 splicing (e.g., PTBP1 and SF3B4) and validate that loss of these factors leads to pervasive CD19 mis-splicing. Our dataset represents a comprehensive resource for identifying predictive biomarkers for CART-19 therapy.enCC BYinfo:eu-repo/semantics/openAccesshttps://creativecommons.org/licenses/by/4.0/570article2024-11-25