To relate clinical outcomes to molecular-level differences between CD28 and 4-1BB CAR T cells, Gad et al. investigated the CAR immune synapse (CARIS). Compared to 4-1BB CARs, CD28 CARs associated with membrane lipid rafts and clustered at the CARIS more rapidly upon target encounter, resulting in stronger lytic granule polarization, shorter contact time, faster cytotoxicity, enhanced serial killing, and effector CD8+ bias. 4-1BB CAR T cells were more proliferative, had higher adhesion molecule (LFA-1, ICAM-1) expression, maintained stronger attachments to target cells, and killed (more slowly) via FasL.
Contributed by Alex Najibi
ABSTRACT: Chimeric antigen receptor T cells (CART) targeting CD19 through CD28.ζ signaling induce rapid lysis of leukemic blasts, contrasting with persistent tumor control exhibited by 4-1BB.ζ-CART. We reasoned that molecular dynamics at the CART immune synapse (CARIS) could explain differences in their tumor rejection kinetics. We observed that CD28.ζ-CART engaged in brief highly lethal CARIS and mastered serial killing, whereas 4-1BB.ζ-CART formed lengthy CARIS and relied on robust expansion and cooperative killing. We analyzed CARIS membrane lipid rafts (mLRs) and found that, upon tumor engagement, CD28.ζ-CAR molecules rapidly but transiently translocated into mLRs, mobilizing the microtubular organizing center and lytic granules to the CARIS. This enabled fast CART recovery and sensitivity to low target site density. In contrast, gradual accumulation of 4-1BB.ζ-CAR and LFA-1 molecules at mLRs built mechanically tonic CARIS mediating chronic Fas ligand-based killing. The differences in CD28.ζ- and 4-1BB.ζ-CARIS dynamics explain the distinct cytolytic behavior of CART and can guide engineering of more adaptive effective cellular products.
Author Info: (1) Interdepartmental Translational Biology and Molecular Medicine Graduate Program, Baylor College of Medicine, Houston, TX 77030, USA. Texas Children's Cancer Center, Texas Child
Author Info: (1) Interdepartmental Translational Biology and Molecular Medicine Graduate Program, Baylor College of Medicine, Houston, TX 77030, USA. Texas Children's Cancer Center, Texas Children's Hospital, Baylor College of Medicine, Houston, TX 77030, USA. Center for Cell and Gene Therapy, Texas Children's Hospital, Houston Methodist Hospital, Baylor College of Medicine, Houston, TX 77030, USA. Department of Pediatrics, Baylor College of Medicine, Houston, TX 77030, USA. Dan L. Duncan Comprehensive Cancer Center, Baylor College of Medicine, Houston, TX 77030, USA. (2) Interdepartmental Translational Biology and Molecular Medicine Graduate Program, Baylor College of Medicine, Houston, TX 77030, USA. Texas Children's Cancer Center, Texas Children's Hospital, Baylor College of Medicine, Houston, TX 77030, USA. Center for Cell and Gene Therapy, Texas Children's Hospital, Houston Methodist Hospital, Baylor College of Medicine, Houston, TX 77030, USA. Department of Pediatrics, Baylor College of Medicine, Houston, TX 77030, USA. Dan L. Duncan Comprehensive Cancer Center, Baylor College of Medicine, Houston, TX 77030, USA. (3) Texas Children's Cancer Center, Texas Children's Hospital, Baylor College of Medicine, Houston, TX 77030, USA. Center for Cell and Gene Therapy, Texas Children's Hospital, Houston Methodist Hospital, Baylor College of Medicine, Houston, TX 77030, USA. Department of Pediatrics, Baylor College of Medicine, Houston, TX 77030, USA. Dan L. Duncan Comprehensive Cancer Center, Baylor College of Medicine, Houston, TX 77030, USA. (4) William A. Brookshire Department of Chemical and Biomolecular Engineering, University of Houston, Houston, TX 77204, USA. (5) Texas Children's Cancer Center, Texas Children's Hospital, Baylor College of Medicine, Houston, TX 77030, USA. Center for Cell and Gene Therapy, Texas Children's Hospital, Houston Methodist Hospital, Baylor College of Medicine, Houston, TX 77030, USA. Department of Pediatrics, Baylor College of Medicine, Houston, TX 77030, USA. Dan L. Duncan Comprehensive Cancer Center, Baylor College of Medicine, Houston, TX 77030, USA. Immunology & Microbiology Graduate Program, Baylor College of Medicine, Houston, TX 77030, USA. (6) Texas Children's Cancer Center, Texas Children's Hospital, Baylor College of Medicine, Houston, TX 77030, USA. Center for Cell and Gene Therapy, Texas Children's Hospital, Houston Methodist Hospital, Baylor College of Medicine, Houston, TX 77030, USA. Department of Pediatrics, Baylor College of Medicine, Houston, TX 77030, USA. Dan L. Duncan Comprehensive Cancer Center, Baylor College of Medicine, Houston, TX 77030, USA. (7) Department of Pediatrics, Baylor College of Medicine, Houston, TX 77030, USA. Texas Children's Hospital William T. Shearer Center for Human Immunobiology, Houston, TX 77030, USA. (8) Texas Children's Cancer Center, Texas Children's Hospital, Baylor College of Medicine, Houston, TX 77030, USA. Department of Pediatrics, Baylor College of Medicine, Houston, TX 77030, USA. Development, Disease Models & Therapeutics Graduate Program, Baylor College of Medicine, Houston, TX 77030, USA. (9) Texas Children's Cancer Center, Texas Children's Hospital, Baylor College of Medicine, Houston, TX 77030, USA. Center for Cell and Gene Therapy, Texas Children's Hospital, Houston Methodist Hospital, Baylor College of Medicine, Houston, TX 77030, USA. Department of Pediatrics, Baylor College of Medicine, Houston, TX 77030, USA. Dan L. Duncan Comprehensive Cancer Center, Baylor College of Medicine, Houston, TX 77030, USA. (10) Texas Children's Cancer Center, Texas Children's Hospital, Baylor College of Medicine, Houston, TX 77030, USA. Center for Cell and Gene Therapy, Texas Children's Hospital, Houston Methodist Hospital, Baylor College of Medicine, Houston, TX 77030, USA. Department of Pediatrics, Baylor College of Medicine, Houston, TX 77030, USA. Dan L. Duncan Comprehensive Cancer Center, Baylor College of Medicine, Houston, TX 77030, USA. (11) Texas Children's Cancer Center, Texas Children's Hospital, Baylor College of Medicine, Houston, TX 77030, USA. Center for Cell and Gene Therapy, Texas Children's Hospital, Houston Methodist Hospital, Baylor College of Medicine, Houston, TX 77030, USA. Dan L. Duncan Comprehensive Cancer Center, Baylor College of Medicine, Houston, TX 77030, USA. (12) Texas Children's Cancer Center, Texas Children's Hospital, Baylor College of Medicine, Houston, TX 77030, USA. Center for Cell and Gene Therapy, Texas Children's Hospital, Houston Methodist Hospital, Baylor College of Medicine, Houston, TX 77030, USA. Department of Pediatrics, Baylor College of Medicine, Houston, TX 77030, USA. Dan L. Duncan Comprehensive Cancer Center, Baylor College of Medicine, Houston, TX 77030, USA. (13) Interdepartmental Translational Biology and Molecular Medicine Graduate Program, Baylor College of Medicine, Houston, TX 77030, USA. Texas Children's Cancer Center, Texas Children's Hospital, Baylor College of Medicine, Houston, TX 77030, USA. Center for Cell and Gene Therapy, Texas Children's Hospital, Houston Methodist Hospital, Baylor College of Medicine, Houston, TX 77030, USA. Department of Pediatrics, Baylor College of Medicine, Houston, TX 77030, USA. Dan L. Duncan Comprehensive Cancer Center, Baylor College of Medicine, Houston, TX 77030, USA. (14) Texas Children's Cancer Center, Texas Children's Hospital, Baylor College of Medicine, Houston, TX 77030, USA. Center for Cell and Gene Therapy, Texas Children's Hospital, Houston Methodist Hospital, Baylor College of Medicine, Houston, TX 77030, USA. Department of Pediatrics, Baylor College of Medicine, Houston, TX 77030, USA. Dan L. Duncan Comprehensive Cancer Center, Baylor College of Medicine, Houston, TX 77030, USA. (15) Texas Children's Cancer Center, Texas Children's Hospital, Baylor College of Medicine, Houston, TX 77030, USA. Center for Cell and Gene Therapy, Texas Children's Hospital, Houston Methodist Hospital, Baylor College of Medicine, Houston, TX 77030, USA. Dan L. Duncan Comprehensive Cancer Center, Baylor College of Medicine, Houston, TX 77030, USA. (16) William A. Brookshire Department of Chemical and Biomolecular Engineering, University of Houston, Houston, TX 77204, USA. (17) William A. Brookshire Department of Chemical and Biomolecular Engineering, University of Houston, Houston, TX 77204, USA. (18) Department of Molecular Physiology and Biological Physics, Center for Molecular and Cell Physiology, University of Virginia, Charlottesville, VA 22903, USA. (19) Texas Children's Cancer Center, Texas Children's Hospital, Baylor College of Medicine, Houston, TX 77030, USA. Center for Cell and Gene Therapy, Texas Children's Hospital, Houston Methodist Hospital, Baylor College of Medicine, Houston, TX 77030, USA. Department of Pediatrics, Baylor College of Medicine, Houston, TX 77030, USA. Dan L. Duncan Comprehensive Cancer Center, Baylor College of Medicine, Houston, TX 77030, USA. (20) Texas Children's Cancer Center, Texas Children's Hospital, Baylor College of Medicine, Houston, TX 77030, USA. Center for Cell and Gene Therapy, Texas Children's Hospital, Houston Methodist Hospital, Baylor College of Medicine, Houston, TX 77030, USA. Dan L. Duncan Comprehensive Cancer Center, Baylor College of Medicine, Houston, TX 77030, USA. Department of Medicine, Baylor College of Medicine, Houston, TX 77030, USA. Department of Pathology and Immunology, Baylor College of Medicine, Houston, TX 77030, USA. (21) Department of Pediatrics, Baylor College of Medicine, Houston, TX 77030, USA. Texas Children's Hospital William T. Shearer Center for Human Immunobiology, Houston, TX 77030, USA. (22) William A. Brookshire Department of Chemical and Biomolecular Engineering, University of Houston, Houston, TX 77204, USA. (23) Texas Children's Cancer Center, Texas Children's Hospital, Baylor College of Medicine, Houston, TX 77030, USA. Center for Cell and Gene Therapy, Texas Children's Hospital, Houston Methodist Hospital, Baylor College of Medicine, Houston, TX 77030, USA. Department of Pediatrics, Baylor College of Medicine, Houston, TX 77030, USA. Dan L. Duncan Comprehensive Cancer Center, Baylor College of Medicine, Houston, TX 77030, USA. (24) Texas Children's Cancer Center, Texas Children's Hospital, Baylor College of Medicine, Houston, TX 77030, USA. Center for Cell and Gene Therapy, Texas Children's Hospital, Houston Methodist Hospital, Baylor College of Medicine, Houston, TX 77030, USA. Department of Pediatrics, Baylor College of Medicine, Houston, TX 77030, USA. Dan L. Duncan Comprehensive Cancer Center, Baylor College of Medicine, Houston, TX 77030, USA. (25) Department of Molecular Physiology and Biological Physics, Center for Molecular and Cell Physiology, University of Virginia, Charlottesville, VA 22903, USA. (26) Department of Pediatrics, Baylor College of Medicine, Houston, TX 77030, USA. Texas Children's Hospital William T. Shearer Center for Human Immunobiology, Houston, TX 77030, USA. (27) Texas Children's Cancer Center, Texas Children's Hospital, Baylor College of Medicine, Houston, TX 77030, USA. Center for Cell and Gene Therapy, Texas Children's Hospital, Houston Methodist Hospital, Baylor College of Medicine, Houston, TX 77030, USA. Department of Pediatrics, Baylor College of Medicine, Houston, TX 77030, USA. Dan L. Duncan Comprehensive Cancer Center, Baylor College of Medicine, Houston, TX 77030, USA. Department of Pathology and Immunology, Baylor College of Medicine, Houston, TX 77030, USA.
