Comparative small molecule screening of primary human acute leukemias, engineered human leukemia and leukemia cell lines.

Targeted therapeutics for high-risk cancers remain an unmet medical need. Here we report the results of a large-scale screen of over 11,000 molecules for their ability to inhibit the survival and growth in vitro of human leukemic cells from multiple sources including patient samples, de novo generated human leukemia models, and established human leukemic cell lines. The responses of cells from de novo models were most similar to those of patient samples, both of which showed striking differences from the cell-line responses.

Mathematical modelling of clonal reduction therapeutic strategies in acute myeloid leukemia.

Over the years, the overall survival of older patients diagnosed with acute myeloid leukemia (AML) has not significantly increased. Although standard cytotoxic therapies that rapidly eliminate dividing myeloblasts are used to induce remission, relapse can occur due to surviving therapy-resistant leukemic stem cells (LSCs). Hence, anti-LSC strategies have become a key target to cure AML.

CD200 expression marks leukemia stem cells in human AML.

The leukemia stem cell (LSC) populations of acute myeloid leukemia (AML) exhibit phenotypic, genetic, and functional heterogeneity that contribute to therapy failure and relapse. Progress toward understanding the mechanistic basis for therapy resistance in LSCs has been hampered by difficulties in isolating cell fractions that enrich for the entire heterogeneous population of LSCs within individual AML samples. We previously reported that CD200 gene expression is upregulated in LSC-containing AML fractions.

Mechanisms and Antitumor Activity of a Binary EGFR/DNA-Targeting Strategy Overcomes Resistance of Glioblastoma Stem Cells to Temozolomide.

Purpose: Glioblastoma (GBM) is a fatal primary malignant brain tumor. GBM stem cells (GSC) contribute to resistance to the DNA-damaging chemotherapy, temozolomide. The epidermal growth factor receptor (EGFR) displays genomic alterations enabling DNA repair mechanisms in half of GBMs.

Mutant H3 histones drive human pre-leukemic hematopoietic stem cell expansion and promote leukemic aggressiveness.

Our ability to manage acute myeloid leukemia (AML) is limited by our incomplete understanding of the epigenetic disruption central to leukemogenesis, including improper histone methylation. Here we examine 16 histone H3 genes in 434 primary AML samples and identify Q69H, A26P, R2Q, R8H and K27M/I mutations (1.6%), with higher incidence in secondary AML (9%).

Heart failure drug proscillaridin A targets MYC overexpressing leukemia through global loss of lysine acetylation.

Background: Cardiac glycosides are approved for the treatment of heart failure as Na/K pump inhibitors. Their repurposing in oncology is currently investigated in preclinical and clinical studies. However, the identification of a specific cancer type defined by a molecular signature to design targeted clinical trials with cardiac glycosides remains to be characterized.

Leukemic stem cell signatures identify novel therapeutics targeting acute myeloid leukemia.

Therapy for acute myeloid leukemia (AML) involves intense cytotoxic treatment and yet approximately 70% of AML are refractory to initial therapy or eventually relapse. This is at least partially driven by the chemo-resistant nature of the leukemic stem cells (LSCs) that sustain the disease, and therefore novel anti-LSC therapies could decrease relapses and improve survival. We performed in silico analysis of highly prognostic human AML LSC gene expression signatures using existing datasets of drug-gene interactions to identify compounds predicted to target LSC gene programs.

Gene expression and mutation-guided synthetic lethality eradicates proliferating and quiescent leukemia cells.

Quiescent and proliferating leukemia cells accumulate highly lethal DNA double-strand breaks that are repaired by 2 major mechanisms: BRCA-dependent homologous recombination and DNA-dependent protein kinase-mediated (DNA-PK-mediated) nonhomologous end-joining, whereas DNA repair pathways mediated by poly(ADP)ribose polymerase 1 (PARP1) serve as backups. Here we have designed a personalized medicine approach called gene expression and mutation analysis (GEMA) to identify BRCA- and DNA-PK-deficient leukemias either directly, using reverse transcription-quantitative PCR, microarrays, and flow cytometry, or indirectly, by the presence of oncogenes such as BCR-ABL1. DNA-PK-deficient quiescent leukemia cells and BRCA/DNA-PK-deficient proliferating leukemia cells were sensitive to PARP1 inhibitors that were administered alone or in combination with current antileukemic drugs.

Complement cascade gene expression defines novel prognostic subgroups of acute myeloid leukemia.

The involvement of the complement pathway in cancer is supported by a growing body of evidence, and yet its role in acute myeloid leukemia (AML) has not been extensively studied. We examined the expression of 87 genes in the complement, coagulation, and fibrinolysis-proteolytic pathways in 374 cytogenetically normal AML samples and observed that these samples can be divided into subgroups on the basis of complement gene expression. Three complement regulatory genes were linked to poor outcome as individual factors in a multivariate analysis (CFH, CFD, and SERPING1) in multiple cohorts.

Barriers to horizontal cell transformation by extracellular vesicles containing oncogenic H-ras.

Extracellular vesicles (EVs) enable the exit of regulatory, mutant and oncogenic macromolecules (proteins, RNA and DNA) from their parental tumor cells and uptake of this material by unrelated cellular populations. Among the resulting biological effects of interest is the notion that cancer-derived EVs may mediate horizontal transformation of normal cells through transfer of mutant genes, including mutant ras. Here, we report that H-ras-mediated transformation of intestinal epithelial cells (IEC-18) results in the emission of exosome-like EVs containing genomic DNA, HRAS oncoprotein and transcript.

GPR56 identifies primary human acute myeloid leukemia cells with high repopulating potential in vivo.

Acute myeloid leukemia (AML) is a genetically heterogeneous hematologic malignancy, which is initiated and driven by a rare fraction of leukemia stem cells (LSCs). Despite the difficulties of identifying a common LSC phenotype, there is increasing evidence that high expression of stem cell gene signatures is associated with poor clinical outcome. Identification of functionally distinct subpopulations in this disease is therefore crucial to dissecting the molecular machinery underlying LSC self-renewal.

miR-126 Regulates Distinct Self-Renewal Outcomes in Normal and Malignant Hematopoietic Stem Cells.

To investigate miRNA function in human acute myeloid leukemia (AML) stem cells (LSC), we generated a prognostic LSC-associated miRNA signature derived from functionally validated subpopulations of AML samples. For one signature miRNA, miR-126, high bioactivity aggregated all in vivo patient sample LSC activity into a single sorted population, tightly coupling miR-126 expression to LSC function. Through functional studies, miR-126 was found to restrain cell cycle progression, prevent differentiation, and increase self-renewal of primary LSC in vivo.

The unfolded protein response governs integrity of the haematopoietic stem-cell pool during stress.

The blood system is sustained by a pool of haematopoietic stem cells (HSCs) that are long-lived due to their capacity for self-renewal. A consequence of longevity is exposure to stress stimuli including reactive oxygen species (ROS), nutrient fluctuation and DNA damage. Damage that occurs within stressed HSCs must be tightly controlled to prevent either loss of function or the clonal persistence of oncogenic mutations that increase the risk of leukaemogenesis.

Reduced lymphoid lineage priming promotes human hematopoietic stem cell expansion.

The hematopoietic system sustains regeneration throughout life by balancing self-renewal and differentiation. To stay poised for mature blood production, hematopoietic stem cells (HSCs) maintain low-level expression of lineage-associated genes, a process termed lineage priming. Here, we modulated expression levels of Inhibitor of DNA binding (ID) proteins to ask whether lineage priming affects self-renewal of human HSCs.

Lysosomal disruption preferentially targets acute myeloid leukemia cells and progenitors.

Despite efforts to understand and treat acute myeloid leukemia (AML), there remains a need for more comprehensive therapies to prevent AML-associated relapses. To identify new therapeutic strategies for AML, we screened a library of on- and off-patent drugs and identified the antimalarial agent mefloquine as a compound that selectively kills AML cells and AML stem cells in a panel of leukemia cell lines and in mice. Using a yeast genome-wide functional screen for mefloquine sensitizers, we identified genes associated with the yeast vacuole, the homolog of the mammalian lysosome.

The genetic basis of early T-cell precursor acute lymphoblastic leukaemia.

Early T-cell precursor acute lymphoblastic leukaemia (ETP ALL) is an aggressive malignancy of unknown genetic basis. We performed whole-genome sequencing of 12 ETP ALL cases and assessed the frequency of the identified somatic mutations in 94 T-cell acute lymphoblastic leukaemia cases. ETP ALL was characterized by activating mutations in genes regulating cytokine receptor and RAS signalling (67% of cases; NRAS, KRAS, FLT3, IL7R, JAK3, JAK1, SH2B3 and BRAF), inactivating lesions disrupting haematopoietic development (58%; GATA3, ETV6, RUNX1, IKZF1 and EP300) and histone-modifying genes (48%; EZH2, EED, SUZ12, SETD2 and EP300).

A small molecule screening strategy with validation on human leukemia stem cells uncovers the therapeutic efficacy of kinetin riboside.

Gene regulatory networks that govern hematopoietic stem cells (HSCs) and leukemia-initiating cells (L-ICs) are deeply entangled. Thus, the discovery of compounds that target L-ICs while sparing HSC is an attractive but difficult endeavor. Presently, most screening approaches fail to counter-screen compounds against normal hematopoietic stem/progenitor cells (HSPCs).

Stem cell gene expression programs influence clinical outcome in human leukemia.

Xenograft studies indicate that some solid tumors and leukemias are organized as cellular hierarchies sustained by cancer stem cells (CSCs). Despite the promise of the CSC model, its relevance in humans remains uncertain. Here we show that acute myeloid leukemia (AML) follows a CSC model on the basis of sorting multiple populations from each of 16 primary human AML samples and identifying which contain leukemia stem cells (LSCs) using a sensitive xenograft assay.

A distinctive DNA damage response in human hematopoietic stem cells reveals an apoptosis-independent role for p53 in self-renewal.

Highly regenerative tissues such as blood must possess effective DNA damage responses (DDR) that balance long-term regeneration with protection from leukemogenesis. Hematopoietic stem cells (HSCs) sustain life-long blood production, yet their response to DNA damage remains largely unexplored. We report that human HSCs exhibit delayed DNA double-strand break rejoining, persistent gammaH2AX foci, and enhanced p53- and ASPP1-dependent apoptosis after gamma-radiation compared to progenitors.

Revised map of the human progenitor hierarchy shows the origin of macrophages and dendritic cells in early lymphoid development.

The classical model of hematopoiesis posits the segregation of lymphoid and myeloid lineages as the earliest fate decision. The validity of this model in the mouse has been questioned; however, little is known about the lineage potential of human progenitors. Here we provide a comprehensive analysis of the human hematopoietic hierarchy by clonally mapping the developmental potential of seven progenitor classes from neonatal cord blood and adult bone marrow.

Comparison of human cord blood engraftment between immunocompromised mouse strains.

The nonobese diabetic/severe combined immune deficiency (NOD-scid) xenotransplantation model is the "gold standard" for assaying human hematopoietic stem cell activity. Systematic advancements, such as depletion of natural killer cell activity with anti-CD122 antibody, direct intrafemoral injection, and deletion or truncation of IL2Rgamma, have improved human cell engraftment; however, questions remain whether these mouse models are equivalent or, if not, which model is superior for assaying hematopoietic stem cell activity. To address this, we compared overall engraftment and multilineage differentiation of near-limiting doses of lineage-depleted human umbilical cord blood cells by direct intrafemoral injection into NOD/Lt-scid, NOD/Shi-scid, NOD/Lt-scid/IL2Rgamma(null) (NSG), and NOD/Shi-scid/IL2Rgamma(null) mice.

Altered expression and deletion of RMO1 in osteosarcoma.

In order to increase our understanding of the molecular events underlying osteosarcoma progression, the expression of approximately 950 genes was examined in 24 primary and metastatic osteosarcoma tumor specimens. A gene, RMO1, was isolated with decreased expression in metastatic samples. Real-Time PCR corroborated this pattern, revealing lower expression in the primary sample in 6 of 7 cases for which both primary and metastatic osteosarcoma samples were available from the same patient (p = 0.

von Willebrand factor expression in osteosarcoma metastasis.

A number of genes are implicated in the initiation and progression of osteosarcoma; however, cytogenetic and comparative genomic hybridization studies indicate the involvement of additional unidentified genes. An examination of gene expression profiles in 22 high-grade osteosarcoma tumor specimens from 15 patients (including paired primary and metastatic samples from five patients) indicated that von Willebrand factor (vWF) mRNA expression may increase during tumor progression. vWF, a large glycoprotein previously considered to be expressed exclusively by endothelial cells and megakaryocytes, is involved in platelet aggregation and adhesion to the subendothelial matrix, processes critical to hematogenous tumor cell metastasis to the lung.