BN-CD38 displays a strong antileukemic efficacy in vivo. (A) Schematic representation of the experimental design and treatment for in vivo studies in NSG mice xenografted with 1 million GFP^posLuc^posTHP-1 cells. On day 18, engraftment was confirmed with bioluminescence imaging (BLI), and mice were randomized. Treatment was started on day 19, and mice were administered control human IgG or BN-CD38 (2.5 mg/kg per mouse), together with 3 million healthy donor–derived human T cells per mouse weekly by IV. Mice in the treatment group were treated once a week for 4 weeks with BN-CD38 plus T cells, whereas mice in the control group received only 2 treatments, because all control mice had died by day 32, before the next planned treatment (day 33). Mice were monitored weekly by BLI for tumor burden assessment. (B) BLI images showing tumor engraftment in the different treatment groups. (C) Bar graph showing the BLI average radiance for control human IgG– and BN-CD38–treated mice on days 25 and 29. Error bars are represented as mean ± SEM. Unpaired Student t test was used for statistical significance calculation. (D) Kaplan-Meier survival curve showing increased survival in BN-CD38–treated mice (n = 8) compared with control IgG–treated mice (n = 10). Log-rank (Mantel-Cox) test was used to determine statistical significance. (E) Kaplan-Meier survival curve showing increased survival in PDX-1 (AML-15) mice transplanted with the BM of BN-CD38–treated animals as reported in the schematic representation and treatment design in supplemental Figure 5G-J. Briefly, 1 million MNCs from a patient with AML with complex karyotype (PDX-1, AML-15) were injected by IV into irradiated NSG, and once engraftment was confirmed in the PB, mice were randomized and weekly cotreated with BN-CD38 (A, 2.5 mg/kg per mouse; n = 5), BN-CD38Mut (n = 5), or control IgG (n = 5), and 3 million healthy donor purified T cells by IV. A total of 3 treatments were administered, and mice BM cells were collected on day 42 and subjected to subsequent secondary and tertiary transplantations. Log-rank (Mantel-Cox) test was used to determine statistical significance; ∗∗∗P < .001. (F) Kaplan-Meier survival curve showing increased survival in PDX-2 (AML-22) mice engrafted with 1 million of total AML MNCs and treated with BN-CD38. Briefly, once engraftment was confirmed in week 8, mice were randomized into 4 groups: control human IgG (n = 5), BN-CD38Mut (n = 5), CD38 NB (n = 4), and BN-CD38 (n = 6). Each group received 2.5 mg/kg of respective treatment and 3 million healthy donor–derived T cells weekly at week 8, 9, 10, and 11 by IV. In week 12 and 13, mice of each group received only respective treatment and no T cells. Survival was monitored, and on week 20, BN-CD38 (A) treated mice were rechallenged with 1 million paired AML blasts (AML-22). In week 29, mice remained healthy and were humanely euthanized to assess tumor burden. Log-rank (Mantel-Cox) test was used to determine statistical significance. (G) Schematic representation of the experimental design and treatment for PDX-3 (AML-4); this model was administered with autologous T cells. Once engraftment was confirmed by flow cytometry on day 14, mice were randomized into 2 groups: BN-CD38Mut (n = 4) and BN-CD38 (n = 5). Each week mice were treated with 2.5 mg/kg of respective BIONICs and 1 million autologous T cells enriched fraction by IV. A total of 4 treatments (once a week) were administered. Mice were humanely euthanized on day 71 and assessed for tumor engraftment and T cells. (H-I) Contour plots of 1 representative mouse for each treatment group illustrate percentages of human CD33^pos AML cells and human CD3^pos T cells in the total BM and spleen (SP) cellular population. AML-to–T cells ratio was calculated for each mouse in 2 treatment groups, and violin plots show that BN-CD38Mut–treated mice had higher tumor burden than levels in BN-CD38–treated mice that showed T-cell expansion. Unpaired Student t test was used to calculate statistical significance; ∗∗P < .01 and ∗∗∗∗P < .0001.