The homeobox transcription factor HB9 induces senescence and blocks differentiation in hematopoietic stem and progenitor cells.

The homeobox gene HLXB9 encodes for the transcription factor HB9, which is essential for pancreatic as well as motor neuronal development. Beside its physiological expression pattern, aberrant HB9 expression has been observed in several neoplasias. Especially in infant translocation t(7;12) acute myeloid leukemia, aberrant HB9 expression is the only known molecular hallmark and is assumed to be a key factor in leukemic transformation. However, so far, only poor functional data exist addressing the oncogenic potential of HB9 or its influence on hematopoiesis. We investigated the influence of HB9 on cell proliferation and cell cycle in vitro, as well as on hematopoietic stem cell differentiation in vivo using murine and human model systems. In vitro, HB9 expression led to premature senescence in human HT1080 and murine NIH3T3 cells, providing for the first time evidence for an oncogenic potential of HB9. Onset of senescence was characterized by induction of the p53-p21 tumor suppressor network, resulting in growth arrest, accompanied by morphological transformation and expression of senescence-associated ß-galactosidase. In vivo, HB9-transduced primary murine hematopoietic stem and progenitor cells underwent a profound differentiation arrest and accumulated at the megakaryocyte/erythrocyte progenitor stage. In line, gene expression analyses revealed de novo expression of erythropoiesis-related genes in human CD34+hematopoietic stem and progenitor cells upon HB9 expression. In summary, the novel findings of HB9-dependent premature senescence and myeloid-biased perturbed hematopoietic differentiation, for the first time shed light on the oncogenic properties of HB9 in translocation t(7;12) acute myeloid leukemia.

Tumoral stem cell reprogramming as a driver of cancer: Theory, biological models, implications in cancer therapy.

Cancer is a clonal malignant disease originated in a single cell and characterized by the accumulation of partially differentiated cells that are phenotypically reminiscent of normal stages of differentiation. According to current models, therapeutic strategies that block oncogene activity are likely to selectively target tumor cells. However, recent evidences have revealed that cancer stem cells could arise through a tumor stem cell reprogramming mechanism, suggesting that genetic lesions that initiate the cancer process might be dispensable for tumor progression and maintenance. This review addresses the impact of these results toward a better understanding of cancer development and proposes new approaches to treat cancer in the future.

Homeobox protein HB9 binds to the prostaglandin E receptor 2 promoter and inhibits intracellular cAMP mobilization in leukemic cells.

BACKGROUND: HB9 is highly expressed in translocation t(7;12) positive infant AML. RESULTS: HB9 binds to the PTGER2 promoter, down-regulates PTGER2 gene expression and subsequently represses cAMP synthesis in hematopoietic cells. CONCLUSION: Expression of HLXB9 represses PTGER2 mediated signaling. SIGNIFICANCE: First molecular report of HB9-dependent target gene regulation in hematopoietic cells. The transcription factor HB9, encoded by the homeobox gene B9 (HLXB9), is involved in the development of pancreatic beta- and motor neuronal cells. In addition, HLXB9 is recurrently rearranged in young children with acute myeloid leukemia characterized by a chromosomal translocation t(7;12)-HLXB9/TEL and concomitant high expression of the unrearranged, wild-type HLXB9 allele. However, target genes of HB9 in hematopoietic cells are not known to date. In this study, we used ChIP-on-chip analysis together with expression profiling and identified PTGER2 (prostaglandin E receptor 2) as a target gene of HB9 in a hematopoietic cell line. The functional HB9 homeodomain as well as the HB9 binding domain within the PTGER2 promoter are essential for binding of HB9 to the PTGER2 promoter region and down-regulation of PTGER2 expression. Functionally, HB9 conducted down-regulation of PTGER2 results in a reduced content of intracellular cAMP mobilization and furthermore the decreased PTGER2 gene expression is valid in bone marrow cells from translocation t(7;12) positive patients. Among the primary and secondary target genes of HB9 in the myeloid cell line HL60, 78% of significantly regulated genes are down-regulated, indicating an overall repressive function of HB9. Differentially regulated genes were preferentially confined to pathways involved in cell-adhesion and cell-cell interactions, similar to the gene expression footprint of HLXB9-expressing cells from t(7;12) positive patients.

Activation-induced cytidine deaminase prevents pro-B cell acute lymphoblastic leukemia by functioning as a negative regulator in Rag1 deficient pro-B cells.

Activation-induced cytidine deaminase (AID) is essential for somatic hypermutation and class switch recombination in mature B-cells, while AID was also shown to play a role in developing pre-BCR/BCR-positive B-cells of the bone marrow. To further elucidate a potential function of Aid in the bone marrow prior to V(D)J-recombination, we utilized an in vivo model which exerts a B-cell developmental arrest at the pro-B cell stage with low frequencies of pro-B cell acute lymphoblastic leukemia (pro-B ALL) development. Therefore, p19Arf-/-Rag1-/- (AR) mice were crossed with Aid-deficient mice (ARA). Surprisingly, loss of Aid expression in pro-B cells accelerated pro-B ALL incidence from 30% (AR) to 98% (ARA). This effect was Aid dose dependent, since Aid+/- animals of the same background displayed a significantly lower incidence (83%). Furthermore, B-cell-specific Aid up-regulation was observed in Aid-competent pro-B ALLs. Additional whole exome/sanger sequencing of murine pro-B ALLs revealed an accumulation of recurrent somatic Jak3 (p.R653H, p.V670A) and Dnm2 (p.G397R) mutations, which highlights the importance of active IL7R signaling in the pro-B ALL blast cells. These findings were further supported by an enhanced proliferative potential of ARA pro-B cells compared to Aid-competent cells from the same genetic background. In summary, we show that both Aid and Rag1 act as a negative regulators in pro-B cells, preventing pro-B ALL.

GEMMs addressing Pax5 loss-of-function in childhood pB-ALL.

Germline mutations in transcription factors, which are implicated in hematopoiesis in general or specifically in B-cell differentiation have recently been described to confer an inherited risk to pB-ALL with often reduced penetrance. Predicting leukemia development, therapy response and long term follow up of mutation carriers is challenging because experience from large patient cohorts and their long term follow up are not available. Genetically Engineered Murine Models (GEMMs) represent a promising approach to create individualized and precise models reproducing the molecular makeup of the human disease. This review focuses on PAX5 loss-of-function and summarizes techniques of murine model generation, available GEMMs, which mimic Pax5 loss-of-function in leukemia development and discusses the challenges and drawbacks of these models. These aspects are discussed in the context of creating a robust model, which serves not only for validation of the relevance of a genomic alteration in pB-ALL but at the same time as a valid preclinical model.