A dual-targeting approach with anti-IL10R CAR-T cells engineered to release anti-CD33 bispecific antibody in enhancing killing effect on acute myeloid leukemia cells

Abstract Background The introduction of immunotherapies, such as chimeric antigen receptors (CAR) T cells and bispecific antibodies (BsAbs), has significantly revolutionized the treatment landscape for acute myeloid leukemia (AML). In this study, we developed a dual-targeting approach with anti-IL10R CAR-T cells engineered to release CD33-targeted bispecific antibody to address the major challenges in T cell-directed therapies, including antigen loss and tumor heterogeneity that contribute to relapse. Methods T cells were transduced with lentiviral supernatants containing IL10R CAR.CD33 BsAb (CAR.BsAb)-encoding sequence, which incorporated the CD33-targeted bsAb and a second-generation IL10R CAR. The efficacy of the CAR.BsAb-T therapy against AML was evaluated both in vitro by cocultures of CAR.BsAb-T cells with leukemia cell lines or primary AML samples, and in vivo using a xenograft leukemia mouse model. Results The study demonstrated the effectiveness of the dual-targeting strategy in eliminating AML cell lines and primary cells expressing varying levels of CD33 and/or IL10R. The secreted anti-CD33 bsAb by IL10R CAR-T cells could amplify the activation and cytotoxicity of both IL10R CAR-T cells and untransduced bystander T cells against CD33 positive leukemia cells. In vivo study further confirmed that CAR.BsAb-T cells could effectively redirect T cells, reduce tumor burden, prolong mice survival, and exhibit no obvious toxicity. This strategy of local bsAbs delivery directly to tumor sites mitigates the pharmacokinetic issues commonly associated with the rapid clearance of bsAbs. Conclusions Overall, the engineering of a single construct targeting IL10R CAR, which subsequently secretes CD33-targeted bsAb, addresses the issue of immune escape due to the heterogeneous expression of IL10R and CD33, and are expected to provide better therapeutic effects for AML treatment.