A Distinct CAR-T Cell Phenotype Mediates Therapeutic Response at Limited Doses

Chimeric antigen receptor (CAR) T cells are typically administered at high doses to ensure enduring therapeutic responses. However, achieving high cell numbers to meet end-product specifications is often challenging, posing substantial medical and socioeconomic barriers. Lower doses may be administered, but response rates are rather low, and the molecular and cellular mechanisms underlying therapeutic efficacy at limited doses remain unknown. To address this gap, we conducted deep phenotyping of anti-CD19 CAR-T cell products and their starting leukapheresis materials from a phase I/II clinical dose-escalation trial targeting B-cell malignancies. Patients responding to CAR-T cells despite low doses exhibited a dose-dependent enrichment of CAR-T cells with functional effector or effector memory-like states. The absolute number of these cells emerged as a robust biomarker of therapeutic response across varying dose levels. Notably, the manufacturing of CAR-T cells capable of mediating therapeutic responses at low doses was determined by T cell levels and T cell-supportive myeloid cells present in the starting material. Conversely, regulatory myeloid cell states contributed to the production of dysfunctional CAR-T cells and subsequent therapy failure. Our study provides critical insights into the phenotype, mechanisms, and biomarkers of CAR-T cells that drive therapeutic responses at low doses, with significant medical and socioeconomic implications.