Anti cxcr2

Leukemic cells were generated as described in Figs. 2a and 3a. MSCs were collected from the endosteal surface of the BM as described before, cultured in low oxygen concentration until the formation of CFU-F colonies, and then plated at the same number for the co-cultures. Once MSCs reached 80% confluency, leukemic cells were plated on top of them and the co-cultured started. Leukemic cells were analyzed after 4 days in co-culture. For serial co-cultures, leukemic cells were collected from the previous co-culture, counted and re-plated onto new MSCs after normalization considering the percentage of GFP⁺ cells. In selected experiments neutralizing antibodies anti-CXCR2 (R&D) (250 ng/ml) and anti-IL6R (Biolegend) (100 ng/ml), or the recombinant proteins CXCL1 and IL6 (both from Biolegend and added at the concentration of 100 ng/ml) were added to the co-cultures. In selected experiments, leukemic cells were derived from the co-cultures and plated in equal number (1000 cells/plate) in the methyl-cellulose. The number of colonies containing more than 50 cells were then counted after 5 days from the plating. In other experiments, total leukemic cells from the co-cultures have been transplanted into wild-type mice. Specifically, we transplanted 20,000 HoxA9–Meis1 GFP⁺ leukemic cells, and 1000 or 20,000 MLL/AF9 GFP⁺ leukemic cells.