CD38-directed, single-chain T cell-engager targets leukemia stem cells through IFNγ-induced CD38 expression.

Treatment resistance of leukemia stem cells (LSCs) and suppression of the autologous immune system represent major challenges to achieve a cure in acute myeloid leukemia (AML). Although AML blasts generally retain high levels of surface CD38 (CD38pos), LSCs are frequently enriched in the CD34posCD38neg blast fraction. Here, we report that IFNγ reduces LSCs clonogenic activity and induces CD38 upregulation in both CD38pos and CD38neg LSC-enriched blasts. IFNγ-induced CD38 upregulation depends on IRF-1 transcriptional activation of the CD38 promoter. To leverage this observation, we created a novel compact, single-chain CD38-CD3 T cell engager (BN-CD38) designed to promote an effective immunological synapse between both CD8 and CD4 T cells and CD38pos AML cells. We demonstrate that BN-CD38 engages autologous CD4pos and CD8pos T cells and CD38pos AML blasts leading to T cell activation and expansion and to the elimination of leukemia cells in an autologous setting. Importantly, BN-CD38 engagement induces the release of high levels of IFNγ, driving the expression of CD38 on CD34posCD38neg LSC-enriched blasts and their subsequent elimination. Critically, while BN-CD38 showed significant in vivo efficacy across multiple disseminated AML cell lines and patient derived xenograft models, it did not affect normal hematopoietic stem cell clonogenicity and the development of multi-lineage human immune cells in CD34pos humanized mice. Taken together, this study provides important insights to target and eliminate AML LSCs.