Enhanced mobilization and competitive repopulation with triple combination of SLU-2609, plerixafor, and tGro-β. (A-B) DBA/2J mice were injected with every combination of tGro-β (2.5 mg/kg), CXCR4 inhibitor plerixafor (5 mg/kg), and/or VLA4 inhibitor SLU-2609 (6 mg/kg). tGro-β was always administered as a separate SC injection. Mice treated with the triple combination of a VLA-4 inhibitor, plerixafor, and tGro-β were first injected SC with a mixture of the VLA-4 inhibitor plus plerixafor followed immediately thereafter by a SC injection of tGro-β. Peripheral blood samples were collected 30, 120, and 240 minutes after injection. Numbers of circulating CFUs (A; n = 10) and LSK cells (B; n = 5) were analyzed. Data are mean ± SD. (C-D) Wild-type and splenectomized DBA/2J mice were treated with G-CSF (9 doses every 12 hours; 125 μg/kg per dose) or the triple combination of tGro-β (2.5 mg/kg), plerixafor (5 mg/kg), and SLU-2609 (9 mg/kg). Treatment in splenectomized mice began 7 days after splenectomy. Numbers of circulating CFUs (A) and LSK cells (B) were analyzed from peripheral blood taken after completion of the 5-day G-CSF regimen or 4 hours after injection of the triple combination. Data are mean ± SD of 2 independent experiments; n = 10 mice per cohort. (E-F) C57BL/6J mice were injected with G-CSF (9 doses every 12 hours; 125 μg/kg per dose), tGro-β (2.5 mg/kg), plerixafor (5 mg/kg), and/or SLU-2609 (9 mg/kg). Numbers of circulating CFUs (E; n = 9-10) and LSK cells (F; n = 4-5) were analyzed from peripheral blood taken at 4 hours after injection. Data are mean ± SD. (G) Competitive repopulation assay. Blood (10 μL) from CD45.1⁺ donors (BALB/c; n = 3 per cohort) mobilized with G-CSF (9 doses every 12 hours; 125 μg/kg per dose), tGro-β (2.5 mg/kg), plerixafor (5 mg/kg), and/or SLU-2609 (9 mg/kg) was mixed with CD45.2⁺ competitor BM cells (BALB/cJ; n = 2 donors; 2.5 × 10⁵ cells per recipient) and transplanted into lethally irradiated primary CD45.2⁺ hosts (BALB/cJ; n = 8-10 recipients). The contribution of mobilized cell populations to hematopoiesis was determined by flow cytometry for CD45.1⁺ donor cells within the CD45⁺CD3^– compartment monthly after transplantation. Data are mean ± SD. ∗P < .05; ∗∗P < .01; ∗∗∗P < .001; ∗∗∗∗P < .0001. BM, bone marrow; SD, standard deviation.