Stemspam

HEK293T cells were obtained from ATCC and grown in DMEM (Thermo Fisher Scientific) supplemented with 10% Hyclone (GE Healthcare) and 1% penicillin/streptomycin solution (Gibco).
Umbilical cord blood samples (CB) from healthy donors were provided by Centro de Transfusión de la Comunidad de Madrid. All samples were collected under written consent and Centro de Transfusión de la Comunidad de Madrid ‘s institutional review board agreement (number PKDEFIN [SAF2017-84248-P]). Mononuclear cells were obtained by fractionation in Ficoll-hypaque according to manufacturer’s recommendations (GE Healthcare). Purified CD34⁺ cells were obtained using a MACS CD34 Micro-Bead kit (Miltenyi Biotec) and were kept frozen or used fresh in further experiments. Cells were viably frozen in 10% dimethyl sulfoxide solution and stored at–liquid nitrogen. Cells were grown in StemSpam (StemCell Technologies) supplemented with 0.5% penicillin/streptomycin solution (Gibco), SCF (100 ng/ml), TPO (100 ng/ml), Flt3 ligand (100 ng/ml) (all of them from EuroBiosciences). Cells were cultured at 37°C, 5% CO₂ and 20% O₂.

Primary Human CD34⁺ cells from Yale Cooperative Center of Excellence in Hematology (YCCEH, Yale University) were immunomagnetically isolated for CD34⁺ as described^{55 (link)} from granulocyte colony stimulating factor–mobilized peripheral blood mononuclear cells from healthy donors. Primary human T-cells were received from UNC Chapel Hill; StemSpam, Methocult Media, MesenCult MSC Basal Media, ACF, MyeloCult (H5100) media, and BIT 9500 serum substitute were purchased from stem cell technologies (Vancouver, Canada). Growth factors IL-3, IL-6, FLT3L, SCF, IL-2, IL-7, and IL-15 were all obtained from ConnStem (CT, USA). Gilteritinib (MW 552.7) was purchased from Medchemexpress (NJ, USA), azacytidine (MW 244.2) from Selleck Chemical (TX, USA), arsenic trioxide (MW 197.8) from Alfa Aesar (MA, USA), cytarabine (MW 243.2) from Acros Organics (NJ, USA), and imatinib (MW 589.7) and U0126 (MW 380.5) from MilliporeSigma (MA, USA). Vibrant CM-DiI was purchased from Invitrogen (CA, USA).