Tisa-cel and axi-cel CAR structure influences the development of resistance to CD19-CAR-T therapy

CD19-CAR-T cells are a major therapy for relapsed/refractory B-cell malignancies, yet ~50% of patients relapse after infusion. Clinical data suggest that tisa-cel treatment is associated with higher rates of CD19 mutation than axi-cel, though direct comparisons have been lacking. Although both products target the FMC63 epitope, they employ different structures of hinge/transmembrane/costimulatory domains (CD8α/CD8α/4-1BB in tisa-cel, CD28/CD28/CD28 in axi-cel). Here, we show that CAR structure is a critical determinant of CD19 loss in tumor B cells. Specifically, repeated exposure to CD19-4-1BB-based CAR-T cells, but not CD19-CD28-based counterparts, drives FMC63-epitope and total CD19 protein loss. Consistent with clinical observations, resistance to CD19-4-1BB-based treatment is associated with aberrant splicing, loss of heterozygosity, and frameshift/missense mutations in CD19 in exons encoding the FMC63 epitope. Mathematical modeling indicates that the failure of CD19-4-1BB-based CAR-T cells to eliminate CD19^low cells, unlike CD28-based CARs, promotes the expansion of resistant populations. These results suggest that optimizing recognition of CD19^low cells could prevent the emergence of resistant clones and reduce relapse rates. We also show that commonly used diagnostic anti-CD19 monoclonal antibodies detect epitopes distinct from FMC63 and may therefore give misleading CD19-positive results, underscoring the need for FMC63-specific assessment to guide diagnostic and therapeutic decisions.