A humanized ossicle platform for real-time tracking and preclinical assessment of CAR-based immunotherapies in acute myeloid leukemia

Abstract

Preclinical evaluation of chimeric antigen receptor (CAR)-T therapies for acute myeloid leukemia (AML) is limited by the lack of models that faithfully recapitulate the human bone marrow (BM) niche. Here, we implemented a humanized ossicle-based AML model that enables simultaneous engraftment of leukemic blasts and longitudinal assessment of responses to CAR-based immunotherapies. Intravenous or intra-ossicle injection of AML blasts produced robust, reproducible disease mimicking features of human AML within its microenvironment. To monitor tumor burden and immune effector cells in real-time, we developed a dual bioluminescence system using distinct luciferases in AML and CAR-T cells. This approach allowed non-invasive longitudinal tracking of CAR-T cell localization, expansion, persistence, and leukemic clearance within the ossicle. Overall, our model provides a powerful platform to study CAR-T cell behavior within a human BM niche and, for the first time, allows simultaneous longitudinal visualization of leukemic burden and CAR-T cell dynamics in a physiologically relevant ossicle-based AML model. TeaserHumanized ossicles combined with dual BLI enable tracking of AML progression and CAR-T cell dynamics in a human stromal niche.