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Blood, Vol. 113, Issue 24, 6215-6224, June 11, 2009
Targeting the leukemia microenvironment by CXCR4 inhibition overcomes resistance to kinase inhibitors and chemotherapy in AML
Blood Zeng et al.
113: 6215
Supplemental materials for: Zeng et al
Files in this Data Supplement:
- Figure S1. AMD3465 enhances anti-tumor effects of ara-C in vivo (JPG, 69.3 KB)
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A20-luc/YFP cells were injected IV into BALB/c mice, and mice were treated with ara-C, AMD3465, or ara-C plus AMD3465. On day 14, mice from control group and from AMD3465 and ara-C–treated group were sacrificed. Histologic sections of bone, liver, and spleen of mice were stained with H & E in control or AMD3465 + ara-C (100mg/kg) treated mice. Red arrows indicate the infiltrating leukemia cells.
- Figure S2 (JPG, 84.5 KB)
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(A) Left panel, CB.17 severe combined immunodeficient (SCID) female mice (5–6 weeks old) were injected intravenously with 0.5 × 106 of Ba/F3-ITD/Luc/GFP cells. 2 weeks post cell injection, AMD3465 was administered subcutaneously at a dose of 20 mg/kg/day for 4 days. The percentage of mobilized Ba/F3-ITD/Luc/GFP cells was detected in peripheral blood on day 4 by GFP flow cytometry. Results represent average percentage of mobilized GFP (+) cells from 3 mice. Right panel, cells from a primary AML sample carrying FLT3-ITD mutation were intravenously injected (20 × 106/mouse) into 7 NOD/SICD mice 8 hours post irradiation (300 rads). AMD3465 was injected subcutaneously at a single dose of 10 mg/kg after leukemia cell engraftment was confirmed by sacrificing two randomly selected mice at week 4. Mobilized human AML cells in the peripheral blood were detected by CD45 flow cytometry one hour after AMD3465 administration. Four of the five mice demonstrated an increased percentage of CD45+ human cells (bar graph). (B) Immunophenotypic analysis of mobilized AML cells. Peripheral blood from mouse #1 was collected prior to and 1 hour post injection of AMD3465 and stained with anti-human antibodies against CD45-APC (top panel); or CD34-FITC, CD38-APC, and CD123-PE. Top panel, SSC (Y-axis) vs CD45 (X-axis) dot plot; middle panel, CD38 (Y-axis) vs CD34 (X-axis); and lower panel, CD123 (Y-axis) vs CD34 (X-axis) dot plot.
- Figure S3. The percentages of circulating Ba/F3-ITD/Luc/GFP cells from mice in sorafenib-treated group or sorafenib combined with AMD3465/G-CSF–treated group were detected by GFP (+) flow cytometry on day 12 (JPG, 53.3 KB)
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The numbers represent percentage of GFP(+) cells.
- Figure S4. Combined CXCR4 and VLA-4 inhibition sensitize blast cells from a primary AML sample to chemotherapy-induced apoptosis (JPG, 46.8 KB)
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(A) Histogram demonstrate surface expression levels of CD34, CXCR4, and CD49d (VLA-4) on a primary AML sample (white: isotypic control; gray: CD34-FITC, CXCR4-PE or CD49d-APC VLA-4 antibody conjugated with APC). (B) The primary AML cells were treated with ara-C alone or in combination with CXCR4 and VLA-4 blocking antibody for 96 hours in the presence or absence of stromal cells. Induction of apoptotic cell death was analyzed by annexin V flow cytometry.
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