Abstract Acute Myeloid Leukemia (AML) originates from the hematopoietic stem and progenitor cell compartment and is associated with an overall poor clinical outcome. T-cell engaging bispecific antibodies (TCEs), binding to a tumor-associated antigen and to CD3, redirect T-cells to the target-antigen expressing cells in an MHC/TCR-independent fashion. The development of TCEs for clinical application is facing several challenges. Firstly, it is difficult to identify a tumor-selective, non-MHC-presented tumor target, not expressed on the tissue of tumor origin, thus limiting specificity. Secondly, CD3-directed TCEs do not provide a second, T-cell activating signal, such as the stimulation of CD28 or 41BB, thus possibly limiting T-cell efficacy. To address both aspects, we generated a CD33xCD28 IgG4-scFv2 with CD28 agonistic activity and tested it in combination with a previously by us reported CD117xCD3 TCE in AML. Combining these two bispecific antibodies significantly improved T-cell mediated lysis of AML cell lines and primary AML samples by enhancing T-cell activation, proliferation and cytokine release in vitro. Furthermore, the addition of CD33xCD28 IgG4-scFv2 showed faster time to cell attachment, increased lytic events, and improved specificity towards double-target expressing cells. In summary, the data indicate that combining co-stimulation via a second tumor-associated antigen to CD3-TCEs enhances T-cell lytic activity and simultaneously increases specificity against double-target expressing AML cells.
Hofstetter et al. (2026) studied this question.