Sfemii media

For testing drug cytotoxic/cytostatic effects, BCP‐ALL primografts were cocultured with BM‐MSC (Pal et al., 2016). To this end, BM‐MSC were seeded onto 48‐well plate at a density of 2.5–5.0 × 10⁴ cells per well, in DMEM (low glucose) supplemented with 20% FBS (Gibco), 2 mm of l‐glutamine (Sigma‐Aldrich), and 1% penicillin/streptomycin solution (Sigma‐Aldrich). The next day, BCP‐ALL primograft cells were added at a density of 0.5 × 10⁶ cells·mL⁻¹ in SFEM II media (STEMCELL Technologies) supplemented with 20% FBS (Gibco), 20 ng·mL⁻¹ of recombinant IL‐3 (R&D Systems, Minneapolis, MN, USA), 10 ng·mL⁻¹ of recombinant IL‐7 (R&D Systems), and penicillin/streptomycin solution (Sigma‐Aldrich). After 24 h of coculture, cells were treated with auranofin (AUR) or adenanthin (ADE) at indicated concentration ranges, in a final volume of 700 μL. For each drug concentration, samples were run in duplicate. After 5 days, cells in suspension (BCP‐ALL cells) and adherent cells (BCP‐ALL+MSC) were collected and the number of viable cells was assessed in a hemocytometer. The numbers of viable BCP‐ALL cells in control groups after 5‐day‐long coculture exceeded the number of seeded cells, confirming the growth‐supporting role of BM‐MSC. Primograft samples used in drug‐testing experiments are listed in Table S7.

A custom optimized cytokine cocktail was developed using commercially available cytokines (Peprotech, New Jersey) to facilitate simultaneous multi-lineage differentiation and self-renewal of freeze-thawed primary human bone marrow-derived CD34+ cells in SFEM II media (cells and media from StemCell Technologies, Vancouver). Cultures were carried out in ultra-low attachment 96-well plates (Corning, New York) with test article added on day 0 and cultured at 37°C, 85% relative humidity, and 5% CO2 for six days before interrogating drug impact via 17-parameter flow cytometry.