LigandCD70.CAR As a Platform for Dual-Targeting CAR T Cells for Acute Myeloid Leukemia

Chimeric antigen receptor (CAR) T cells have demonstrated remarkable potency in acute lymphoblastic leukemia (ALL), but similar success has yet to be seen in acute myeloid leukemia (AML). Due to the variable expression of targetable antigens on AML blasts and leukemic stem cells, treatment with single-antigen targeting immunotherapy may select for antigen-negative clones that ultimately cause disease progression or relapse. An appropriate combination of two different CAR-binders and co-stimulatory domains could both increase tumor control and reduce antigen escape. We utilized a binder derived from human CD27 protein, the only known ligand of CD70, to generate the LigandCD70.CAR, which demonstrated superior in vivo activity against AML compared to conventional antibody-derived CARs (Sauer et al 2021). As the LigandCD70.CAR consists of a full length CD27 fused with CD3-zeta, this design takes advantage of the physiologic receptor-ligand interaction of CD70/CD27 to bind with high affinity to CD70+ cells and provides CD27-mediated costimulation to the T cells. These characteristics combined with the high expression of CD70 on more than 80% of AML make the LigandCD70.CAR a promising candidate for combination with other CARs to target AML. We evaluated the LigandCD70.CAR in combination with a CLL-1.CAR and a CD123.CAR, as both have shown preclinical efficacy and are currently being tested in clinical trials. We co-transduced activated T cells with LigandD70.CAR and CLL-1 CAR to generate LigandCD70/CLL-1 bispecific CAR (CLL-1.BiCAR) T cells. CLL-1.BiCAR T cells had comparable in vitro expansion to single-CAR expressing T cells but with enhanced cytotoxicity against CLL-1+ AML cell lines compared to single CLL-1.CARs, even if CD70 expression on tumor cells was low. In xenograft AML models of CD70-low CLL-1-high tumors (HL60), CLL-1.BiCAR T cells enhanced tumor control, survival and T cell persistence compared to single CLL-1.CAR T Cells (Figure 1). In order to determine whether the LigandC70.CAR can enhance other CARs, we co-transduced ligandCD70.CAR with CD123.CAR (Riberdy et al 2020) to generate a Ligand.CD70/CD123 bispecific CAR (CD123.BiCAR). The in vitro expansion and cytotoxicity was comparable to single-CAR expressing T cells. Treatment with CD123.BiCAR T cells significantly improved in vivo survival and tumor control in a xenograft model of AML with CD70-low/CD123-high target cells. In summary, LigandCD70.CAR co-expressed with either CLL-1.CAR or CD123.CAR demonstrate enhanced anti-tumor efficacy compared to single-targeting CLL-1.CAR and CD123.CAR, even in CD70-low tumors. The simultaneous targeting of two AML antigens may thus mitigate the risk of antigen-negative relapse. These data indicate that the Ligand70.CAR combined with other AML targeting CARs could be an effective therapeutic option for AML. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal