The two subunits of core binding factor (Runx1 and CBFβ) play critical roles in hematopoiesis and are frequent targets of chromosomal translocations found in leukemia. The binding of the CBFβ-smooth muscle myosin heavy chain (SMMHC) fusion protein to Runx1 is essential for leukemogenesis, making this a viable target for treatment. We have developed inhibitors with low micromolar affinity which effectively block binding of Runx1 to CBFβ. NMR-based docking shows that these compounds bind to CBFβ at a site displaced from the binding interface for Runx1, that is, these compounds function as allosteric inhibitors of this protein-protein interaction, a potentially generalizable approach. Treatment of the human leukemia cell line ME-1 with these compounds shows decreased proliferation, indicating these are good candidates for further development.