The most frequent chromosomal translocations in pediatric acute myeloid leukemia affect the 11q23 locus and give rise to mixed lineage leukemia (MLL) fusion genes, MLL-AF9 being the most prevalent. The MLL-AF9 fusion gene has been shown to induce leukemia in both mouse and human models. In this study, we demonstrate that leukemogenic activity of MLL-AF9 requires RUVBL2 (RuvB-like 2), an AAA þ ATPase family member that functions in a wide range of cellular processes, including chromatin remodeling and transcriptional regulation. Expression of RUVBL2 was dependent on MLL-AF9, as it increased upon immortalization of human cord blood-derived hematopoietic progenitor cells with the fusion gene and decreased following loss of fusion gene expression in conditionally immortalized mouse cells. Short hairpin RNA-mediated silencing experiments demonstrated that both the immortalized human cells and the MLL-AF9-expressing human leukemia cell line THP-1 required RUVBL2 expression for proliferation and survival. Furthermore, inhibition of RUVBL2 expression in THP-1 cells led to reduced telomerase activity and clonogenic potential. These data were confirmed with a dominant-negative Walker B-mutated RUVBL2 construct. Taken together, these data suggest the possibility of targeting RUVBL2 as a potential therapeutic strategy for MLL-AF9-associated leukemia.

Key Points• STAT3 activity is necessary for TEL-AML1 leukemia maintenance.• TEL-AML1 induces STAT3 activation via RAC1 and leading to induction of MYC expression.The t(12;21)(p13;q22) translocation is the most common chromosomal abnormality in pediatric leukemia. Although this rearrangement involves 2 well-characterized transcription factors, TEL and AML1, the molecular pathways affected by the result of the translocation remain largely unknown. Also in light of recent studies showing genetic and functional heterogeneities in cells responsible for cancer clone maintenance and propagation, targeting a single common deregulated pathway may be critical for the success of novel therapies. Here we describe a novel signaling pathway that is essential for oncogenic addiction in TEL-AML1 leukemia. Our data indicate a direct role for TEL-AML1, via increasing the activity of RAC1, in regulating the phosphorylation of signal transducer and activator of transcription 3 (STAT3), which results in transcriptional induction of MYC. We demonstrate that human leukemic cell lines carrying this translocation are highly sensitive to treatment with S3I-201, a specific STAT3 inhibitor, and, more interestingly, that primary human leukemic samples are also responsive to the drug in the same concentration range. Thus, STAT3 inhibition represents a promising possible therapeutic strategy for the treatment of TEL-AML1 leukemia. (Blood. 2013;122(4):542-549)

The molecular mechanisms underlying oncogenesis in leukemias associated with rearrangement of the Mixed Lineage Leukemia (MLL) gene have received a considerable amount of attention over the last two decades. In this review we will focus on recent studies, published over the past year, that reveal new insights into the multi-protein complexes formed by MLL and MLL fusion proteins, the role of epigenetic deregulation in MLL fusion function, downstream transcriptional target genes, the importance of the leukemia cell of origin, the role played by microRNAs, cooperating mutations and the implications that recent research has for the therapy of MLL-rearranged leukemia.

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IntroductionTranslocations affecting the mixed lineage leukemia (MLL) gene on chromosome band 11q23 are the most common chromosomal aberrations in pediatric acute myeloid leukemia (AML). 1 This cytogenetic subgroup is associated with an intermediate risk, 2 intensive chemotherapy having improved overall survival, although MLL-rearranged AML is still unfavorable in older patients. 3 Taken together, these data suggest that novel therapeutic interventions based on targeting MLL fusion function would benefit a significant number of patients.The MLL fusion genes resulting from these translocations are potent oncogenes. [4][5][6] They are sufficient to induce aberrant selfrenewal programs in mouse hematopoietic progenitor cells (HPCs) leading to transformation and leukemic progression. 7,8 Recent studies have demonstrated that this is also true for human HPCs. 9,10 We and others have previously shown that inhibition of MLL fusion expression reverses HPC immortalization [11][12][13] and abrogates established leukemia in vivo. 14,15 This suggests that the transcriptional and signaling pathways controlled by MLL fusions are necessary for maintenance as well as initiation of leukemia. Recent studies have demonstrated that progression of preleukemic to leukemic MLL fusion cells involves increased canonical Wnt signaling 16 and Rac GTPase activation 17 ; both of these signaling pathways play critical roles in leukemia progression. 10,16,[18][19][20] The sensitivity of MLL-rearranged AML cells to Rac inhibition suggests that targeting Rac activity or signaling pathways leading to Rac activation may represent novel therapeutic opportunities. In support of this, pharmacologic Rac inhibition was found to interfere with leukemic engraftment of MLL-rearranged human AML cell lines. 19 The available evidence indicates that MLL fusions can induce Rac activity. Thus, transformation of human cord blood-derived HPCs by MLL-AF9 was accompanied by increased Rac activation. 10,19 However, little is known about the molecular details of how this is achieved. In the present study, we have used conditionally immortalized preleukemic and leukemic cells to address the dependency of Rac GTPase activity on continued MLL fusion expression and to establish components of the signaling pathway linking the 2. These experiments identify multiple points potentially suitable for therapeutic targeting of MLL-rearranged AML. Methods MiceAll mice were maintained in the animal facilities of the UCL Institute of Child Health and experiments were performed according to United Kingdom Home Office regulations. The generation and characterization of Frat1/2/3 triple-knockout (TKO) mice have been reported previously. 21 These mice have been backcrossed onto the C57BL/6 background 20 times. Age-and sex-matched C57BL/6 J mice were used as controls. Retrovirus and lentivirus cloning and productionThe pMSCV-MLL-ENL-pgk-neo retroviral construct has been previously described. 14 Oligonucleotides for shRNAs targeting Frat1, Frat2, and Dvl1 were designed using RNAi centra...