To identify therapeutic targets in acute myeloid leukemia (AML), we chemically interrogated 200 sequenced primary specimens. Mubritinib, a known ERBB2 inhibitor, elicited strong anti-leukemic effects ...in vitro and in vivo. In the context of AML, mubritinib functions through ubiquinone-dependent inhibition of electron transport chain (ETC) complex I activity. Resistance to mubritinib characterized normal CD34+ hematopoietic cells and chemotherapy-sensitive AMLs, which displayed transcriptomic hallmarks of hypoxia. Conversely, sensitivity correlated with mitochondrial function-related gene expression levels and characterized a large subset of chemotherapy-resistant AMLs with oxidative phosphorylation (OXPHOS) hyperactivity. Altogether, our work thus identifies an ETC complex I inhibitor and reveals the genetic landscape of OXPHOS dependency in AML.
•Mubritinib exhibits strong in vitro and in vivo anti-AML activity•It is selectively toxic against a distinct and large subset of poor outcome AMLs•Mubritinib is a ubiquinone-dependent NADH dehydrogenase inhibitor•Mubritinib-sensitive primary leukemias exhibit OXPHOS hyperactivity
Through a chemical screen in sequenced primary AML specimens, Baccelli et al. identify mubritinib, a known ERBB2 inhibitor, to be selectively toxic against a large subset of poor outcome AML specimens that exhibit oxidative phosphorylation hyperactivity by inhibiting the electron transport chain complex I.
Using next-generation sequencing of primary acute myeloid leukemia (AML) specimens, we identified to our knowledge the first unifying genetic network common to the two subgroups of KMT2A ...(MLL)-rearranged leukemia, namely having MLL fusions or partial tandem duplications. Within this network, we experimentally confirmed upregulation of the gene with the most subtype-specific increase in expression, LOC100289656, and identified cryptic MLL fusions, including a new MLL-ENAH fusion. We also identified a subset of MLL fusion specimens carrying mutations in SPI1 accompanied by inactivation of its transcriptional network, as well as frequent RAS pathway mutations, which sensitized the leukemias to synthetic lethal interactions between MEK and receptor tyrosine kinase inhibitors. This transcriptomics-based characterization and chemical interrogation of human MLL-rearranged AML was a valuable approach for identifying complementary features that define this disease.
In this study, we analyzed RNA-sequencing data of 14 samples characterized by biallelic CEBPA (CEBPAbi) mutations included in the Leucegene collection of 415 primary acute myeloid leukemia (AML) ...specimens, and describe for the first time high frequency recurrent mutations in the granulocyte colony-stimulating factor receptor gene CSF3R, which signals through JAK-STAT proteins. Chemical interrogation of these primary human specimens revealed a uniform and specific sensitivity to all JAK inhibitors tested irrespective of their CSF3R mutation status, indicating a general sensitization of JAK-STAT signaling in this leukemia subset. Altogether, these results identified the co-occurrence of mutations in CSF3R and CEBPA in a well-defined AML subset, which uniformly responds to JAK inhibitors and paves the way to personalized clinical trials for this disease.
•CSF3R was the most frequently mutated gene identified in this CEBPAbi AML cohort analyzed by next-generation sequencing.•CEBPAbi AML that have a characteristic transcriptomic profile are more sensitive to JAK inhibitors than CEBPAwt AML.
Chromatin modulators are emerging as attractive drug targets, given their widespread implication in human cancers and susceptibility to pharmacological inhibition. Here we establish the histone ...methyltransferase G9a/EHMT2 as a selective regulator of fast proliferating myeloid progenitors with no discernible function in hematopoietic stem cells (HSCs). In mouse models of acute myeloid leukemia (AML), loss of G9a significantly delays disease progression and reduces leukemia stem cell (LSC) frequency. We connect this function of G9a to its methyltransferase activity and its interaction with the leukemogenic transcription factor HoxA9 and provide evidence that primary human AML cells are sensitive to G9A inhibition. Our results highlight a clinical potential of G9A inhibition as a means to counteract the proliferation and self-renewal of AML cells by attenuating HoxA9-dependent transcription.
-mutated (
) acute myeloid leukemia (AML) is associated with adverse outcome, highlighting the urgent need for a better genetic characterization of this AML subgroup and for the design of efficient ...therapeutic strategies for this disease. Toward this goal, we further dissected the mutational spectrum and gene expression profile of
AML and correlated these results to drug sensitivity to identify novel compounds targeting this AML subgroup.
RNA-sequencing of 47
primary AML specimens was performed and sequencing results were compared to those of
wild-type samples. Chemical screens were also conducted using
specimens to identify compounds selectively affecting the viability of
AML.
We show that samples with no remaining
wild-type allele are clinically and genetically distinct and display a more homogeneous gene expression profile. Chemical screening revealed that most
specimens are sensitive to glucocorticoids (GCs) and we confirmed that GCs inhibit AML cell proliferation through their interaction with the glucocorticoid receptor (GR). We observed that specimens harboring
mutations expected to result in low residual RUNX1 activity are most sensitive to GCs, and that coassociating mutations as well as GR levels contribute to GC sensitivity. Accordingly, acquired glucocorticoid sensitivity was achieved by negatively regulating
expression in human AML cells.
Our findings show the profound impact of
allele dosage on gene expression profile and glucocorticoid sensitivity in AML, thereby opening opportunities for preclinical testing which may lead to drug repurposing and improved disease characterization.
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Histone methylation is a dynamic and reversible process proposed to directly impact on stem cell fate. The Jumonji (JmjC) domain–containing family of demethylases comprises 27 members that target ...mono-, di-, and trimethylated lysine residues of histone (or nonhistone) proteins. To evaluate their role in regulation of hematopoietic stem cell (HSC) behavior, we performed an in vivo RNAi-based functional screen and demonstrated that Jarid1b and Jhdm1f play opposing roles in regulation of HSC activity. Decrease in Jarid1b levels correlated with an in vitro expansion of HSCs with preserved long-term in vivo lymphomyeloid differentiation potential. Through RNA sequencing analysis, Jarid1b knockdown was associated with increased expression levels of several HSC regulators (Hoxa7, Hoxa9, Hoxa10, Hes1, Gata2) and reduced levels of differentiation-associated genes. shRNA against Jhdmlf, in contrast, impaired hematopoietic reconstitution of bone marrow cells. Together, our studies identified Jarid1b as a negative regulator of HSC activity and Jhdmlf as a positive regulator of HSC activity.
•Jarid1b knockdown promotes enhanced HSC activity.
Hematopoietic stem cells (HSCs) can self-renew extensively after transplantation. The conditions supporting their in vitro expansion are still being defined. Retroviral overexpression of the human ...homeobox B4 (HOXB4) gene in mouse bone marrow cells enables over 40-fold expansion of HSCs in vitro. To circumvent the requirement for retroviral infection, we used recombinant human TAT-HOXB4 protein carrying the protein transduction domain of the HIV transactivating protein (TAT) as a potential growth factor for stem cells. HSCs exposed to TAT-HOXB4 for 4 d expanded by about four- to sixfold and were 8-20 times more numerous than HSCs in control cultures, indicating that HSC expansion induced by TAT-HOXB4 was comparable to that induced by the human HOXB4 retrovirus during a similar period of observation. Our results also show that TAT-HOXB4-expanded HSC populations retain their normal in vivo potential for differentiation and long-term repopulation. It is thus feasible to exploit recombinant HOXB4 protein for rapid and significant ex vivo expansion of normal HSCs.
While the role of canonical (β-catenin-mediated) Wnt signaling in hematolymphopoiesis has been studied extensively, little is known of the potential importance of non-canonical Wnt signals in ...hematopoietic cells. Wnt4 is one of the Wnt proteins that can elicit non-canonical pathways. We have previously shown that retroviral overexpression of Wnt4 by hematopoietic cells increased thymic cellularity as well as the frequency of early thymic progenitors and bone marrow hematopoietic progenitor cells (HPCs). However, the molecular pathways responsible for its effect in HPCs are not known.
Here we report that Wnt4 stimulation resulted in the activation of the small GTPase Rac1 as well as Jnk kinases in an HPC cell line. Jnk activity was necessary, while β-catenin was dispensable, for the Wnt4-mediated expansion of primary fetal liver HPCs in culture. Furthermore, Jnk2-deficient and Wnt4 hemizygous mice presented lower numbers of HPCs in their bone marrow, and Jnk2-deficient HPCs showed increased rates of apoptosis. Wnt4 also improved HPC activity in a competitive reconstitution model in a cell-autonomous, Jnk2-dependent manner. Lastly, we identified Fz6 as a receptor for Wnt4 in immature HPCs and showed that the absence of Wnt4 led to a decreased expression of four polarity complex genes.
Our results establish a functional role for non-canonical Wnt signaling in hematopoiesis through a pathway involving Wnt4, Fz6, Rac1 and Jnk kinases.
The incidence of refractory acute myeloid leukemia (AML) is on the increase due in part to an aging population that fails to respond to traditional therapies. High throughput genomic analysis ...promises better diagnosis, prognosis, and therapeutic intervention based on improved patient stratification. Relevant preclinical models are urgently required to advance drug development in this area. The collaborating oncogenes, HOXA9 and MEIS1, are frequently co-overexpressed in cytogenetically normal AML (CN-AML), and a conditional transplantation mouse model was developed that demonstrated oncogene dependency and expression levels comparable to CN-AML patients. Integration of gene signatures obtained from the mouse model and a cohort of CN-AML patients using statistically significant connectivity map analysis identified Entinostat as a drug with the potential to alter the leukemic condition toward the normal state. Ex vivo treatment of leukemic cells, but not age-matched normal bone marrow controls, with Entinostat validated the gene signature and resulted in reduced viability in liquid culture, impaired colony formation, and loss of the leukemia initiating cell. Furthermore, in vivo treatment with Entinostat resulted in prolonged survival of leukemic mice. This study demonstrates that the HDAC inhibitor Entinostat inhibits disease maintenance and prolongs survival in a clinically relevant murine model of cytogenetically normal AML.
Hox gene products, initially characterized as master regulators of embryonic patterning, are also required for proper functioning of adult tissues. There is also a growing body of evidence that links ...Hox proteins to regulation of cellular proliferation/transformation. However, the underlying molecular mechanisms of Hox-associated transformation and tissue growth have yet to be elucidated. Using a well established model system for studying changes in cellular proliferation induced by Hoxb4, we show that AP-1 activity is markedly increased in Hoxb4-transduced cells due to significant upregulation of Jun-B and Fra-1 protein levels. Furthermore, we also show that the specific changes in AP-1 protein expression are necessary for the proliferation effects induced by Hoxb4, and that these changes converge to increase levels of cyclin D1, a known integrator of proliferation signals. Our observations thus link Hox gene products with key elements of the cell cycle machinery.
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