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. 2014 Dec 17;3(Suppl 1):S25–S26. doi: 10.1038/leusup.2014.14

Myeloid differentiation and the leukemia-initiating cell

D G Tenen 1,2,*
PMCID: PMC4851280  PMID: 27175269

Acute myeloblastic leukemia (AML) is defined by a block in the differentiation of multipotent proliferating progenitor cells into mature granulocytes. C/EBPα (CCAAT enhancer binding protein alpha) is a basic leucine zipper transcription factor and one of the central factors controlling differentiation. C/EBPα is required for adult hematopoetic stem cell (HSC) quiescence, and low levels of C/EBPα lead to enhanced HSC function. C/EBPα is a crucial regulator of granulopoiesis, and its absence, as shown in C/EBPα knockout (KO) mice, results in a block in the differentiation of common myeloid progenitor cells into granulocyte–macrophage progenitor cells accompanied by an absence of mature granulocytes. C/EBPα induces cell cycle arrest and has the capacity to inhibit proliferation by inducing p21, inhibiting cyclin-dependent kinases, mediating Calpain cleavage of cyclin A, interacting with and inhibiting E2F, and downregulating C-myc. We have shown that CEBPA is mutated in 16% of AML-M2 patients. Mutations that occur at the N-terminus result in a frameshift and loss of the full-length 42-kDa C/EBPα protein. A nonfunctional 30-kDa C/EBPα form that is initiated downstream of the frameshift is still produced. The N-terminal mutation acts as a dominant negative inhibitor of full-length CEBPA.1 To generate a mouse model that recapitulates the dominant negative function of C/EBPα, which results in AML, a combination of C-terminal mutation and N-terminal mutation was used (C/EBPα K/L).2 The central question in the molecular etiology of AML is: which cell is transformed in AML, HSC or committed progenitor? Is myeloid differentiation required for the formation of the ‘leukemic stem cell' in AML? C/EBPα KOs mimic human AML and although the mice have, by definition, human AML (blasts in bone marrow and blood), the mice do not develop a malignant disease.3, 4, 5 It is known that C/EBPα suppresses targets such as Bmi-1 and N-myc.3, 4, 5 Our efforts focused on identifying other targets that may contribute to the phenotype. A short hairpin RNA (shRNA) screen done on Lin−kit+Sca1+ (LSK) cells isolated from C/EBPα KO bone marrow identified Sox4 as a C/EBPα target-mediating serial replating (‘self-renewal') and inhibiting differentiation. Sox4 is a member of the SOX transcription factor family with a highly conserved high mobility group domain. Sox4 is a frequent target of insertional mutagenesis in retrovirally induced myeloid leukemias. Sox4 is also required for self-renewal and transformation pathways during leukemic evolution of multiple murine AML models (MOZ-TIF2, AML1-ETO and NUP98-HOXA9).6, 7, 8, 9 It is known that Sox4 expression is reciprocal to that of C/EBPα in wild-type mice, and C/EBPα binds to the Sox4 promoter directly to represses its expression (like Bmi-1 and N-myc). Therefore, we were interested in comparing the phenotypes of C/EBPα and Sox4 KOs to better understand the relationship between these two transcriptional factors in the context of myeloid differentiation. We have observed that Sox4 was upregulated in C/EBPα KO LSK and that Sox4 KO rescued myeloid progenitor expansion and increased proliferation of C/EBPα KO LSK (and rescued suppression of Bmi1 and N-myc). Importantly, in humans, enhanced SOX4 levels are observed in AML patients with dysfunctional CEBPA.10 Moreover SOX4 knockdown was confirmed to rescue myeloid differentiation in human CEBPA-mutant AML, and human C/EBPα mutants derived from patients did not repress Sox4 RNA levels in C/EBPα KO LSK and did not repress Sox4 promoter activity.11 In summary, our studies showed that Sox4 mediates serial replating (‘self-renewal') of both C/EBPα KO LSK and C/EBPα-mutant AML (K/L). In addition, Sox4 is upregulated in human primary blasts with C/EBPα mutation (modeled by K/L) or silenced (modeled by C/EBPa KO). shRNA downregulation of Sox4 induces differentiation of human primary blasts with C/EBPα dysfunction.

Acknowledgments

The symposium and publication of this supplement were sponsored by the Division of Hematology/Oncology at the Warren Alpert Medical School of Brown University and NIH Center of Biomedical Research Excellence (COBRE) for Stem Cells Biology at Rhode Island Hospital.

The author declares no conflict of interest.

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