Stem regenin 1

C/EBPA^N/C-8 cells were cultured at the concentration of 1 × 10⁶ cell/mL in StemSpanTM Serum-Free Expansion Medium II (SFEMII) plus 10% StemSpanTM CD34+ Expansion Supplement and 175 nM UM171 (STEMCELL Technologies). All other patient cells were grown in SFEMII supplemented with 100 ng/mL thrombopoietin, 10 ng/mL FMS-like tyrosine kinase 3 ligand, 750 nM Stem Regenin 1 (SRI), 10 ng/mL Interleukin 3, 10 ng/mL human granulocyte/macrophage colony stimulating factor, 150 ng/mL Stem Cell Factor (all Peprotech) and UM729 (STEMCELL Technologies). Patient CD34+ or CD117+ progenitors were isolated with CD34 MicroBead Kit, human or CD117 MicroBead Kit, human (Miltenyi-Biotech) as described in [4 (link)].

Primary AML and MDS cells were collected from patients who had consented to research protocols approved by the Institutional Review Board at The University of Texas MD Anderson Cancer Center for analysis of hematologic malignancies. BM aspirate mononuclear cells from patients with MDS were purified by Ficoll density centrifugation using standard procedures. Lymphocytes were isolated by using Lymphocyte Separation Medium (SIGMA).
Primary AML peripheral blood mononuclear cells (PBMCs) were cultured in serum-free Expansion Medium (Cat. 09650) supplemented with BIT 9500 Serum Substitute (Cat. 09500; both from STEMCELL Technologies Inc., Vancouver, BC, Canada) and cytokines including stem cell factor (SCF, 100 ng/mL, Cat. 300-07), Flt3 ligand (50 ng/mL, Cat. 300 – 19), IL3 (20 ng/mL, Cat. 200-03), and G-CSF (20 ng/mL, Cat. 300 – 23; all from Peprotech, Rocky Hill, NJ) and StemRegenin 1 (1 μM, Cat. S2858; Selleck Chemicals LLC, Houston, TX).