ERG and c-MYC regulate a critical gene network in BCR::ABL1-driven B cell acute lymphoblastic leukemia.

Philadelphia chromosome-positive B cell acute lymphoblastic leukemia (B-ALL), characterized by the BCR::ABL1 fusion gene, remains a poor prognosis cancer needing new therapeutic approaches. Transcriptomic profiling identified up-regulation of oncogenic transcription factors ERG and c-MYC in BCR::ABL1 B-ALL with ERG and c-MYC required for BCR::ABL1 B-ALL in murine and human models. Profiling of ERG- and c-MYC-dependent gene expression and analysis of ChIP-seq data established ERG and c-MYC coordinate a regulatory network in BCR::ABL1 B-ALL that controls expression of genes involved in several biological processes. Prominent was control of ribosome biogenesis, including expression of RNA polymerase I (POL I) subunits, the importance of which was validated by inhibition of BCR::ABL1 cells by POL I inhibitors, including CX-5461, that prevents promoter recruitment and transcription initiation by POL I. Our results reveal an essential ERG- and c-MYC-dependent transcriptional network involved in regulation of metabolic and ribosome biogenesis pathways in BCR::ABL1 B-ALL, from which previously unidentified vulnerabilities and therapeutic targets may emerge.

Differential in vivo roles of Mpl cytoplasmic tyrosine residues in murine hematopoiesis and myeloproliferative disease.

Thrombopoietin (Tpo), which binds to its specific receptor, the Mpl protein, is the major cytokine regulator of megakaryopoiesis and circulating platelet number. Tpo binding to Mpl triggers activation of Janus kinase 2 (Jak2) and phosphorylation of the receptor, as well as activation of several intracellular signalling cascades that mediate cellular responses. Three tyrosine (Y) residues in the C-terminal region of the Mpl intracellular domain have been implicated as sites of phosphorylation required for regulation of major Tpo-stimulated signalling pathways: Mpl-Y565, Mpl-Y599 and Mpl-Y604. Here, we have introduced mutations in the mouse germline and report a consistent physiological requirement for Mpl-Y599, mutation of which resulted in thrombocytopenia, deficient megakaryopoiesis, low hematopoietic stem cell (HSC) number and function, and attenuated responses to myelosuppression. We further show that in models of myeloproliferative neoplasms (MPN), where Mpl is required for pathogenesis, thrombocytosis was dependent on intact Mpl-Y599. In contrast, Mpl-Y565 was required for negative regulation of Tpo responses; mutation of this residue resulted in excess megakaryopoiesis at steady-state and in response to myelosuppression, and exacerbated thrombocytosis associated with MPN.

T-ALL can evolve to oncogene independence.

The majority of cases of T-cell acute lymphoblastic leukemia (T-ALL) contain chromosomal abnormalities that drive overexpression of oncogenic transcription factors. However, whether these initiating oncogenes are required for leukemia maintenance is poorly understood. To address this, we developed a tetracycline-regulated mouse model of T-ALL driven by the oncogenic transcription factor Lmo2. This revealed that whilst thymus-resident pre-Leukemic Stem Cells (pre-LSCs) required continuous Lmo2 expression, the majority of leukemias relapsed despite Lmo2 withdrawal. Relapse was associated with a mature phenotype and frequent mutation or loss of tumor suppressor genes including Ikzf1 (Ikaros), with targeted deletion Ikzf1 being sufficient to transform Lmo2-dependent leukemias to Lmo2-independence. Moreover, we found that the related transcription factor TAL1 was dispensable in several human T-ALL cell lines that contain SIL-TAL1 chromosomal deletions driving its overexpression, indicating that evolution to oncogene independence can also occur in human T-ALL. Together these results indicate an evolution of oncogene addiction in murine and human T-ALL and show that loss of Ikaros is a mechanism that can promote self-renewal of T-ALL lymphoblasts in the absence of an initiating oncogenic transcription factor.

An Erg-driven transcriptional program controls B cell lymphopoiesis.

B lymphoid development is initiated by the differentiation of hematopoietic stem cells into lineage committed progenitors, ultimately generating mature B cells. This highly regulated process generates clonal immunological diversity via recombination of immunoglobulin V, D and J gene segments. While several transcription factors that control B cell development and V(D)J recombination have been defined, how these processes are initiated and coordinated into a precise regulatory network remains poorly understood. Here, we show that the transcription factor ETS Related Gene (Erg) is essential for early B lymphoid differentiation. Erg initiates a transcriptional network involving the B cell lineage defining genes, Ebf1 and Pax5, which directly promotes expression of key genes involved in V(D)J recombination and formation of the B cell receptor. Complementation of Erg deficiency with a productively rearranged immunoglobulin gene rescued B lineage development, demonstrating that Erg is an essential and stage-specific regulator of the gene regulatory network controlling B lymphopoiesis.

Deficiency of 5-hydroxyisourate hydrolase causes hepatomegaly and hepatocellular carcinoma in mice.

With the notable exception of humans, uric acid is degraded to (S)-allantoin in a biochemical pathway catalyzed by urate oxidase, 5-hydroxyisourate (HIU) hydrolase, and 2-oxo-4-hydroxy-4-carboxy-5-ureidoimidazoline decarboxylase in most vertebrate species. A point mutation in the gene encoding mouse HIU hydrolase, Urah, that perturbed uric acid metabolism within the liver was discovered during a mutagenesis screen in mice. The predicted substitution of cysteine for tyrosine in a conserved helical region of the mutant-encoded HIU hydrolase resulted in undetectable protein expression. Mice homozygous for this mutation developed elevated platelet counts secondary to excess thrombopoietin production and hepatomegaly. The majority of homozygous mutant mice also developed hepatocellular carcinoma, and tumor development was accelerated by exposure to radiation. The development of hepatomegaly and liver tumors in mice lacking Urah suggests that uric acid metabolites may be toxic and that urate oxidase activity without HIU hydrolase function may affect liver growth and transformation. The absence of HIU hydrolase in humans predicts slowed metabolism of HIU after clinical administration of exogenous urate oxidase in conditions of uric acid-related pathology. The data suggest that prolonged urate oxidase therapy should be combined with careful assessment of toxicity associated with extrahepatic production of uric acid metabolites.