The MLL gene at chromosome band 11q23 is commonly involved in reciprocal translocations detected in acute leukemias. A number of experiments show that the resulting MLL fusion genes directly contribute to leukemogenesis. Among the many known MLL fusion partners, AF4 is relatively common, particularly in acute lymphoblastic leukemia in infants. The AF4 protein interacts with the product of another gene, AF9, which is also fused to MLL in acute leukemias. Based on mapping studies of the AF9-binding domain of AF4, we have developed a peptide, designated PFWT, which disrupts the AF4-AF9 interaction in vitro and in vivo. We provide evidence that this peptide is able to inhibit the proliferation of leukemia cells with t(4;11) chromosomal translocations expressing MLL-AF4 fusion genes. Further, we show that this inhibition is mediated through apoptosis. Importantly, the peptide does not affect the proliferative capacity of hematopoietic progenitor cells. Our findings indicate that the AF4-AF9 protein complex is a promising new target for leukemia therapy and that the PFWT peptide may serve as a lead compound for drug development.