Developing a membrane-proximal CD33-targeting CAR T cell
Ruby Freeman, Sanam Shahid, Abdul G. Khan, Serena C Mathew, Sydney Souness, Erin R. Burns, Jasmine S. Um, Kento Tanaka, Winson Cai, Sarah Yoo, Andrew Dunbar, Young Joo Park, Devin McAvoy, Kinga Hosszu, Ross L. Levine, Jaap Jan Boelens, Ivo C. Lorenz, Renier J. Brentjens, Anthony F. Daniyan
Abstract
BACKGROUND: CD33 is a tractable target in acute myeloid leukemia (AML) for chimeric antigen receptor (CAR) T cell therapy, but clinical success is lacking. METHODS: We developed 3P14HLh28Z, a novel CD33-directed CD28/CD3Z-based CAR T cell derived from a high-affinity binder obtained through membrane-proximal fragment immunization in humanized mice. RESULTS: We found that immunization exclusively with the membrane-proximal domain of CD33 is necessary for identification of membrane-proximal binders in humanized mice. Compared with clinically validated lintuzumab-based CAR T cells targeting distal CD33 epitopes, 3P14HLh28Z showed enhanced in vitro functionality as well as superior tumor control and increased overall survival in both low antigen density and clinically relevant patient-derived xenograft models. Increased activation and enhanced polyfunctionality led to enhanced efficacy. CONCLUSIONS: Showing for the first time that a membrane-proximal CAR is superior to a membrane-distal one in the setting of CD33 targeting, our results demonstrate the rationale for targeting membrane-proximal epitopes with high-affinity binders. We also demonstrate the importance of optimizing CAR T cells for functionality in settings of both low antigen density and clinically relevant patient-derived models.