Lentiviral gene therapy reverts GPIX expression and phenotype in Bernard-Soulier syndrome type C Martínez Navajas, Gonzalo Ceron-Hernandez, Jorge Simon, Iris Lupiañez, Pablo Diaz-McLynn, Sofia Perales, Sonia Modlich, Ute Guerrero, José Antonio Martin, Francisco Sevivas, Teresa Lozano, María Luisa Rivera, Jose Ramos Mejía, Verónica Tersteeg, Claudia Real Luna, Pedro José RNA/DNA editing Bernard-Soulier Syndrome Gene therapy GP9 Hematopoietic stem cells Disease models Induced pluripotent stem cells Lentiviral vector CRISPR-Cas9 GPIb-V-IX receptor von Willebrand factor Edición génica Síndrome de Bernard-Soulier Terapia génica Células madre hematopoyéticas Modelos de enfermedad Células madre pluripotentes Vector lentiviral Bernard-Soulier syndrome (BSS) is a rare congenital disease characterized by macrothrombocytopenia and frequent bleeding. It is caused by pathogenic variants in three genes (GP1BA, GP1BB, or GP9) that encode for the GPIbα, GPIbβ, and GPIX subunits of the GPIb-V-IX complex, the main platelet surface receptor for von Willebrand factor, being essential for platelet adhesion and aggregation. According to the affected gene, we distinguish BSS type A1 (GP1BA), type B (GP1BB), or type C (GP9). Pathogenic variants in these genes cause absent, incomplete, or dysfunctional GPIb-V-IX receptor and, consequently, a hemorrhagic phenotype. Using gene-editing tools, we generated knockout (KO) human cellular models that helped us to better understand GPIb-V-IX complex assembly. Furthermore, we developed novel lentiviral vectors capable of correcting GPIX expression, localization, and functionality in human GP9-KO megakaryoblastic cell lines. Generated GP9-KO induced pluripotent stem cells produced platelets that recapitulated the BSS phenotype: absence of GPIX on the membrane surface and large size. Importantly, gene therapy tools reverted both characteristics. Finally, hematopoietic stem cells from two unrelated BSS type C patients were transduced with the gene therapy vectors and differentiated to produce GPIX-expressing megakaryocytes and platelets with a reduced size. These results demonstrate the potential of lentiviral-based gene therapy to rescue BSS type C. 2025-01-21T11:46:46Z 2025-01-21T11:46:46Z 2023-06-12 journal article https://hdl.handle.net/10481/99837 10.1016/j.omtn.2023.06.008 eng http://creativecommons.org/licenses/by-nc-nd/4.0/ open access Attribution-NonCommercial-NoDerivatives 4.0 Internacional