1:Josep Carreras Leukemia Research Institute; 2:Cancer Immunotherapy Unit (UNICA), Department of Immunology, Hospital Universitario 12 de Octubre, 28041 Madrid, Spain; 3:Immuno-Oncology and Immunotherapy Group, Instituto de Investigación Sanitaria 12 de Octubre (imas12), 28041 Madrid, Spain; 4:Centro de Biología Molecular Severo Ochoa CSIC-UAM, Madrid, Spain; 5:Red Española de Terapias Avanzadas (TERAV) - Instituto de Salud Carlos III (ISCII) (RICORS, RD21/0017/0029-RD21; RD21/0017/0030); 6:Centro de Investigacio´n Biome´dica en Red-Oncologi´a (CIBERONC), Instituto de Salud Carlos III, Madrid, Spain; 7:Department of Biomedicine, School of Medicine, Universitat de Barcelona, Spain; 8:Institucio´ Catalana de Recerca i Estudis Avanc¸ats (ICREA), Barcelona, Spain.

The dismal clinical outcome of relapsed/refractory (R/R) T-cell acute lymphoblastic leukemia (T-ALL) highlights the need for effective targeted therapeutic strategies. Despite chimeric antigen receptor (CAR)-engineered T cells have revolutionized the treatment of B-cell malignancies, their clinical implementation in T-ALL is on its infancy. In this context, Antigen-A (AgA) represents a safe target for cortical T-ALL (coT-ALL) patients. T-ALL relapses are commonly very aggressive and hyperleukocytic, posing a challenge to recover sufficient non-leukemic mature T cells from leukapheresis in R/R T-ALL patients. Here, we report a comprehensive study comparing the efficacy of engineered T cells either expressing a second-generation AgA-CAR or Secreting AgAxCD3 T cell-engaging Antibodies (AgA-STAb). AgA-T cell engagers bind to cell surface expressed AgA and CD3 and specific T cell activation. We show that AgA-STAb recruitment of bystander T cells provides AgA-STAbs with enhanced in vitro cytotoxic efficiency than AgA-CAR T cells at much lower effector:target ratios. AgA-STAb cells are as effective as AgA-CAR T cells in cuttin-edge in vivo T-ALL patient-derived xenograft models. Our data suggests that AgA-STAb-T cells could be an alternative to AgA-CAR T-cells in coT-ALL patients, especially for those with aggressive and hyperleukocytic relapses with limited numbers of non-leukemic effector T cells.