Explore the vast range of titles available.

The treatment landscape of acute myeloid leukemia (AML) is evolving, with promising therapies entering clinical translation, yet patient responses remain heterogeneous, and biomarkers for tailoring treatment are lacking. To understand how disease heterogeneity links with therapy response, we determined the leukemia cell hierarchy makeup from bulk transcriptomes of more than 1,000 patients through deconvolution using single-cell reference profiles of leukemia stem, progenitor and mature cell types. Leukemia hierarchy composition was associated with functional, genomic and clinical properties and converged into four overall classes, spanning Primitive, Mature, GMP and Intermediate. Critically, variation in hierarchy composition along the Primitive versus GMP or Primitive versus Mature axes were associated with response to chemotherapy or drug sensitivity profiles of targeted therapies, respectively. A seven-gene biomarker derived from the Primitive versus Mature axis was associated with response to 105 investigational drugs. Cellular hierarchy composition constitutes a novel framework for understanding disease biology and advancing precision medicine in AML. A novel gene expression classifier of AML heterogeneity captures patient-specific variation in leukemia cell composition and predicts clinical responses to treatment.

The concept of differentiation therapy emerged from the fact that hormones or cytokines may promote differentiation ex vivo, thereby irreversibly changing the phenotype of cancer cells. Its hallmark success has been the treatment of acute promyelocytic leukaemia (APL), a condition that is now highly curable by the combination of retinoic acid (RA) and arsenic. Recently, drugs that trigger differentiation in a variety of primary tumour cells have been identified, suggesting that they are clinically useful. This Opinion article analyses the basis for the clinical successes of RA or arsenic in APL by assessing the respective roles of terminal maturation and loss of self-renewal. By reviewing other successful examples of drug-induced tumour cell differentiation, novel approaches to transform differentiating drugs into more efficient therapies are proposed.

N 6 -methyladenosine (m 6 A) is an abundant internal RNA modification 1 , 2 that is catalysed predominantly by the METTL3–METTL14 methyltransferase complex 3 , 4 . The m 6 A methyltransferase METTL3 has been linked to the initiation and maintenance of acute myeloid leukaemia (AML), but the potential of therapeutic applications targeting this enzyme remains unknown 5 – 7 . Here we present the identification and characterization of STM2457, a highly potent and selective first-in-class catalytic inhibitor of METTL3, and a crystal structure of STM2457 in complex with METTL3–METTL14. Treatment of tumours with STM2457 leads to reduced AML growth and an increase in differentiation and apoptosis. These cellular effects are accompanied by selective reduction of m 6 A levels on known leukaemogenic mRNAs and a decrease in their expression consistent with a translational defect. We demonstrate that pharmacological inhibition of METTL3 in vivo leads to impaired engraftment and prolonged survival in various mouse models of AML, specifically targeting key stem cell subpopulations of AML. Collectively, these results reveal the inhibition of METTL3 as a potential therapeutic strategy against AML, and provide proof of concept that the targeting of RNA-modifying enzymes represents a promising avenue for anticancer therapy. Treatment with a specific inhibitor of the N 6 -methyladenosine methyltransferase METTL3 leads to reduced growth of cancer cells, indicating the potential of approaches targeting RNA-modifying enzymes for anticancer therapy.

Key Points Myelodysplastic syndrome (MDS) is one of the most common haematological malignancies and is associated with increased age and exposure to previous chemotherapy and radiation. It is characterized by cytopenias, morphological dysplasia and a propensity to transform to acute myeloid leukaemia (AML). Clonal haematopoiesis of indeterminate potential (CHIP) is a condition in which a substantial percentage of haematopoietic cells bear a somatic mutation in a gene that is recurrently mutated in haematological malignancies, including MDS. CHIP is strongly associated with age and an increased risk of haematological malignancy. More than 50 recurrently mutated genes have been identified in MDS, many of which occur in genes encoding RNA splicing factors, epigenetic regulators, haematopoietic transcription factors and kinase signalling pathways. Individual mutations in MDS are associated with specific morphological findings, have independent prognostic significance and can predict response to therapy in some cases. AML that arises out of a pre-existing MDS can be distinguished from de novo AML by the presence of specific mutations, such as those in splicing factors and certain epigenetic regulators. Some mutations are associated with increased sensitivity or resistance to standard therapeutic interventions, providing new targets for the development of novel therapeutic agents. Currently, allogeneic haematopoietic stem cell transplantation is the only known curative treatment for MDS. This Review discusses the molecular processes and clonal evolution that lead to myelodysplastic syndrome (MDS) and secondary acute myeloid leukaemia, highlighting the ways in which these insights are shaping the clinical management of MDS. Myelodysplastic syndrome (MDS) is a clonal disease that arises from the expansion of mutated haematopoietic stem cells. In a spectrum of myeloid disorders ranging from clonal haematopoiesis of indeterminate potential (CHIP) to secondary acute myeloid leukaemia (sAML), MDS is distinguished by the presence of peripheral blood cytopenias, dysplastic haematopoietic differentiation and the absence of features that define acute leukaemia. More than 50 recurrently mutated genes are involved in the pathogenesis of MDS, including genes that encode proteins involved in pre-mRNA splicing, epigenetic regulation and transcription. In this Review we discuss the molecular processes that lead to CHIP and further clonal evolution to MDS and sAML. We also highlight the ways in which these insights are shaping the clinical management of MDS, including classification schemata, prognostic scoring systems and therapeutic approaches.

Myeloid malignancies, including acute myeloid leukaemia (AML), arise from the expansion of haematopoietic stem and progenitor cells that acquire somatic mutations. Bulk molecular profiling has suggested that mutations are acquired in a stepwise fashion: mutant genes with high variant allele frequencies appear early in leukaemogenesis, and mutations with lower variant allele frequencies are thought to be acquired later 1 – 3 . Although bulk sequencing can provide information about leukaemia biology and prognosis, it cannot distinguish which mutations occur in the same clone(s), accurately measure clonal complexity, or definitively elucidate the order of mutations. To delineate the clonal framework of myeloid malignancies, we performed single-cell mutational profiling on 146 samples from 123 patients. Here we show that AML is dominated by a small number of clones, which frequently harbour co-occurring mutations in epigenetic regulators. Conversely, mutations in signalling genes often occur more than once in distinct subclones, consistent with increasing clonal diversity. We mapped clonal trajectories for each sample and uncovered combinations of mutations that synergized to promote clonal expansion and dominance. Finally, we combined protein expression with mutational analysis to map somatic genotype and clonal architecture with immunophenotype. Our findings provide insights into the pathogenesis of myeloid transformation and how clonal complexity evolves with disease progression. The evolution of myeloid malignancies is investigated using combined single-cell sequencing and immunophenotypic analysis.

Large-scale single-cell analyses are of fundamental importance in order to capture biological heterogeneity within complex cell systems, but have largely been limited to RNA-based technologies. Here we present a comprehensive benchmarked experimental and computational workflow, which establishes global single-cell mass spectrometry-based proteomics as a tool for large-scale single-cell analyses. By exploiting a primary leukemia model system, we demonstrate both through pre-enrichment of cell populations and through a non-enriched unbiased approach that our workflow enables the exploration of cellular heterogeneity within this aberrant developmental hierarchy. Our approach is capable of consistently quantifying ~1000 proteins per cell across thousands of individual cells using limited instrument time. Furthermore, we develop a computational workflow (SCeptre) that effectively normalizes the data, integrates available FACS data and facilitates downstream analysis. The approach presented here lays a foundation for implementing global single-cell proteomics studies across the world. Single-cell proteomics can provide insights into the molecular basis for cellular heterogeneity. Here, the authors develop a multiplexed single-cell proteomics and computational workflow, and show that their strategy captures the cellular hierarchies in an Acute Myeloid Leukemia culture model.

The pattern of somatic mutations observed at diagnosis of acute myeloid leukemia (AML) has been well-characterized. However, the premalignant mutational landscape of AML and its impact on risk and time to diagnosis is unknown. Here we identified 212 women from the Women’s Health Initiative who were healthy at study baseline, but eventually developed AML during follow-up (median time: 9.6 years). Deep sequencing was performed on peripheral blood DNA of these cases and compared to age-matched controls that did not develop AML. We discovered that mutations in IDH1 , IDH2 , TP53 , DNMT3A , TET2 and spliceosome genes significantly increased the odds of developing AML. All subjects with TP53 mutations ( n  = 21 out of 21 patients) and IDH1 and IDH2 ( n  = 15 out of 15 patients) mutations eventually developed AML in our study. The presence of detectable mutations years before diagnosis suggests that there is a period of latency that precedes AML during which early detection, monitoring and interventional studies should be considered. Somatic mutations detected years before diagnosis increase the odds of development of acute myeloid leukemia in women.

The development of cancer is intimately associated with genetic abnormalities that target proteins with intrinsically disordered regions (IDRs). In human haematological malignancies, recurrent chromosomal translocation of nucleoporin (NUP98 or NUP214) generates an aberrant chimera that invariably retains the nucleoporin IDR—tandemly dispersed repeats of phenylalanine and glycine residues 1 , 2 . However, how unstructured IDRs contribute to oncogenesis remains unclear. Here we show that IDRs contained within NUP98–HOXA9, a homeodomain-containing transcription factor chimera recurrently detected in leukaemias 1 , 2 , are essential for establishing liquid–liquid phase separation (LLPS) puncta of chimera and for inducing leukaemic transformation. Notably, LLPS of NUP98–HOXA9 not only promotes chromatin occupancy of chimera transcription factors, but also is required for the formation of a broad ‘super-enhancer’-like binding pattern typically seen at leukaemogenic genes, which potentiates transcriptional activation. An artificial HOX chimera, created by replacing the phenylalanine and glycine repeats of NUP98 with an unrelated LLPS-forming IDR of the FUS protein 3 , 4 , had similar enhancing effects on the genome-wide binding and target gene activation of the chimera. Deeply sequenced Hi-C revealed that phase-separated NUP98–HOXA9 induces CTCF-independent chromatin loops that are enriched at proto-oncogenes. Together, this report describes a proof-of-principle example in which cancer acquires mutation to establish oncogenic transcription factor condensates via phase separation, which simultaneously enhances their genomic targeting and induces organization of aberrant three-dimensional chromatin structure during tumourous transformation. As LLPS-competent molecules are frequently implicated in diseases 1 , 2 , 4 – 7 , this mechanism can potentially be generalized to many malignant and pathological settings. The NUP98–HOXA9 oncogenic fusion protein found in leukaemia undergoes phase separation in the nucleus, which helps to promote activation of leukaemic genes and to establish aberrant chromatin looping.

Howard Chang, Ravindra Majeti and colleagues define the chromatin accessibility and transcriptional landscapes in 13 human primary blood cell types and in acute myeloid leukemia cells. They identify potential regulators governing hematopoietic differentiation and genetic elements linked to regulatory evolution in cancer cells. We define the chromatin accessibility and transcriptional landscapes in 13 human primary blood cell types that span the hematopoietic hierarchy. Exploiting the finding that the enhancer landscape better reflects cell identity than mRNA levels, we enable 'enhancer cytometry' for enumeration of pure cell types from complex populations. We identify regulators governing hematopoietic differentiation and further show the lineage ontogeny of genetic elements linked to diverse human diseases. In acute myeloid leukemia (AML), chromatin accessibility uncovers unique regulatory evolution in cancer cells with a progressively increasing mutation burden. Single AML cells exhibit distinctive mixed regulome profiles corresponding to disparate developmental stages. A method to account for this regulatory heterogeneity identified cancer-specific deviations and implicated HOX factors as key regulators of preleukemic hematopoietic stem cell characteristics. Thus, regulome dynamics can provide diverse insights into hematopoietic development and disease.

The N 6 -methyladenosine (m 6 A) modification in mRNAs, generated by the enzyme METTL3, controls normal human hematopoietic stem/progenitor cell differentiation and maintains the undifferentiated leukemic phenotype of human acute myeloid leukemia cells. N 6 -methyladenosine (m 6 A) is an abundant nucleotide modification in mRNA that is required for the differentiation of mouse embryonic stem cells. However, it remains unknown whether the m 6 A modification controls the differentiation of normal and/or malignant myeloid hematopoietic cells. Here we show that shRNA-mediated depletion of the m 6 A-forming enzyme METTL3 in human hematopoietic stem/progenitor cells (HSPCs) promotes cell differentiation, coupled with reduced cell proliferation. Conversely, overexpression of wild-type METTL3, but not of a catalytically inactive form of METTL3, inhibits cell differentiation and increases cell growth. METTL3 mRNA and protein are expressed more abundantly in acute myeloid leukemia (AML) cells than in healthy HSPCs or other types of tumor cells. Furthermore, METTL3 depletion in human myeloid leukemia cell lines induces cell differentiation and apoptosis and delays leukemia progression in recipient mice in vivo . Single-nucleotide-resolution mapping of m 6 A coupled with ribosome profiling reveals that m 6 A promotes the translation of c-MYC , BCL2 and PTEN mRNAs in the human acute myeloid leukemia MOLM-13 cell line. Moreover, loss of METTL3 leads to increased levels of phosphorylated AKT, which contributes to the differentiation-promoting effects of METTL3 depletion. Overall, these results provide a rationale for the therapeutic targeting of METTL3 in myeloid leukemia.