Developing a membrane-proximal CD33-targeting CAR T cell.

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.
Competing Interests: Competing interests: AGK, ICL, RJB, and A Daniyan have filed provisional patent applications covering applications of membrane-proximal CD33 binders for cellular therapy. A Dunbar, YP, and RL are supported by National Cancer Institute P01 CA108671 11 (RL). DM and KKH are supported by an MSK Geoffrey Beene Cancer Research Award. RL is on the supervisory board of Qiagen and is a scientific advisor to Imago, Mission Bio, Zentalis, Ajax, Auron, Prelude, C4 Therapeutics and Isoplexis. He receives research support from and consulted for Celgene and Roche and has consulted for Incyte, Janssen, Astellas, Morphosys and Novartis, and has received honoraria from AstraZeneca, Roche, Lilly and Amgen for invited lectures and from Gilead for grant reviews. JJB has consulted for Avrobio, Sobi, BlueRock, Sanofi, Omeros, Advanced Clinical, MERCK, Bluebird Bio and is on the Data Safety Monitoring Board/Advisory Board for Advanced Clinical. RJB has licensed intellectual property to and collects royalties from BMS, Caribou, and Sanofi. RJB received research funding from BMS. RJB is a consultant to BMS, Atara Biotherapeutics Inc, Cargo Tx, Triumvira and was a consultant for CoImmune but ended in the past 3 months and Gracell Biotechnologies Inc but ended employment in the past 24 months. RJB is a member of the scientific advisory board for Triumvira, Cargo Tx and was a member of the scientific advisory board for CoImmune, but that ended in the past 3 months.
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