Duv-CART cells conferred a survival advantage in mice bearing intermediate disease burden OSU-CLL. OSU-CLL was engrafted in NOG mice, and upon OSU-CLL (herein called CLL) reaching a mean of 1.2% (CD28/CD19) or 0.15% (41BB/CD19) of nucleated cell content, mice were treated with 1.0 × 10⁶ control CART or Duv-CART cells on days 15 to 18. Data from anti-CD19 CART with a CD28 costimulatory domain (CD28/CD19 CART) are in the left panels, and data from anti-CD19 CART with a 41BB costimulatory domain (41BB/CD19 CART) are shown in the right panels. (A) Kaplan-Meier survival analysis of CD28/CD19 control- and Duv-CART-cell–treated mice. (B) Frequency of CLL cells in peripheral blood, defined by flow cytometry as CD20⁺CD5⁺ over time after CLL engraftment. The in vivo expansion of total CART cells (C) and CD8⁺ CART cells (D) over time since infusion of CART. (CART cells were gated based on expression of GFP.) (E) Representative flow cytometry plots from peak expansion (day 18 after CART infusion). (F) Kaplan-Meier survival analysis of 41BB/CD19 control- and Duv-CART–treated mice. (G) Frequency of CLL in peripheral blood, defined by flow cytometry as CD20⁺CD5⁺ over time after CLL engraftment. The in vivo expansion of total human T cells (H) and CD8⁺ T cells (I) over time since infusion of CART. (The secondary antibody against the 41BB CAR failed to detect the CAR, and human CD3 was therefore used as a proxy for CAR-expressing cells.) (J) Representative flow cytometry plots from peak expansion (day 21 after CART infusion). In vivo immune checkpoint expression over time for CD28/CD19 CART within the CD4⁺ T-cell subset (K) and CD8⁺ T-cell subset (L). In vivo immune checkpoint expression over time for 41BB/CD19 CART within the CD4⁺ T-cell subset (M) and CD8⁺ T-cell subset (N). *P < .05; **P ≤ .01; ***P ≤ .001; ****P ≤ .0001.