Persistent CAR-T donor cells contribute to mixed chimerism with cytopenia after CAR-T-bridged allo-HSCT: rescue by CAR-target-knockout DLI in three cases.

Persistent CAR-T donor cells after CAR-T-bridged allogeneic hematopoietic stem cell transplantation (allo-HSCT) may complicate hematopoietic recovery. When CAR-T and HSCT cells share the same donor, standard donor chimerism cannot distinguish their contributions. We report three patients with relapsed/refractory T-cell malignancies who received CAR-T and allo-HSCT from distinct donors, enabling three-party short tandem repeat (STR) analysis to resolve CAR-T donor, HSCT donor, and recipient compartments. Patient 2, with detectable residual CAR transgene at transplantation (240 copies/µg gDNA), developed mixed chimerism with cytopenia (MCC) and a CAR-T donor contribution of 47.72%. Patient 3, also with detectable residual CAR transgene (22,162 copies/µg gDNA), developed mixed chimerism, with a CAR-T donor contribution of 14.23%. In contrast, Patient 1, with undetectable CAR transgene at transplantation, showed stable complete donor chimerism (CDC) and hematopoietic recovery. Guided by the two-step reconstitution pattern observed in Patient 1, we administered CAR-target-knockout donor lymphocyte infusion (KO-DLI) as rescue treatment in Patient 2 and preemptive intervention in Patient 3. Both eventually became CAR-undetectable, with restoration of CDC and hematologic recovery, although the independent effect of KO-DLI could not be isolated. All three remain in complete remission (follow-up + 713, +335, + 201 days). Residual CAR transgene at transplantation may serve as an early risk signal for CAR-T donor cell persistence, and KO-DLI warrants prospective validation as a transplantation-sparing salvage approach.