Ibrutinib increases the engraftment and antitumor activity of CTL019 cells when administered concurrently. (A) Protocol schema depicting the engraftment of NOD scid gamma (NSG) mice with 10⁶ luciferase-expressing Nalm-6 cells or 5 to 10 × 10⁶ OSU-CLL cells. At day 7 or 9, animals (n = 5-12 per group) were randomized according to tumor burden to receive vehicle alone, 25 mg/kg per day of ibrutinib, or 10⁶ CTL019 cells per mouse. Ibrutinib or empty vehicle was continuously administered for the entire duration of animal experiments. (B) Absolute numbers of adoptively transferred peripheral blood T cells were monitored weekly by retro-orbital bleeding and flow cytometric detection. (C) Bioluminescence images of 5 representative mice in the CTL019 and CTL019 + Ibr. treatment groups are shown at day 20 post–CAR T-cell infusion. (D) Peak bioluminescent (BLI) tumor burden depicted for each experimental group. (E) PD-1 expression on adoptively transferred CTL019 cells in the spleens of ibrutinib-naive or ibrutinib-treated mice at the time of euthanasia. (F) Overall survival of untreated mice (Nalm-6 alone [No Rx]) or animals treated with ibrutinib alone, CTL019 cells, or a combination of CTL019 and ibrutinib shown over time. Two separate experiments with similar results were performed. (G) In vivo expansion kinetics of CTL019 cells in mice with or without continuous ibrutinib treatment. Error bars show the standard error. (H) Survival of OSU-CLL-bearing NSG mice treated on day 7 after tumor injection with 10⁶ CTL019 cells (n = 12 mice per group). Overall survival curves were plotted using the Kaplan-Meier method and compared using the log-rank (Mantel-Cox) test. Statistical significance between groups with differences in CTL019 expansion and tumor burden was determined using a 2-tailed Student t test. *P < .05, **P < .01, ***P < .001, ****P < .0001. CBG, click beetle green; n.s., not statistically significant.