Publications by authors named "George S Vassiliou"

77 Publications

Correction to: Analysis of T cell receptor clonotypes in tumor microenvironment identifies shared cancer-type-specific signatures.

Cancer Immunol Immunother 2021 Oct 22. Epub 2021 Oct 22.

Cancer Therapeutics Research Laboratory, National Cancer Centre Singapore, 11 Hospital Crescent, Singapore, 169610, Singapore.

View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1007/s00262-021-03086-0DOI Listing
October 2021

The CADM1 tumor suppressor gene is a major candidate gene in MDS with deletion of the long arm of chromosome 11.

Blood Adv 2021 Oct 12. Epub 2021 Oct 12.

Belgian Cancer Registry, Brussels, Belgium.

Myelodysplastic syndromes (MDS) represent a heterogeneous group of clonal hematopoietic stem-cell disorders characterized by ineffective hematopoiesis leading to peripheral cytopenias and in a substantial proportion of cases to acute myeloid leukemia. The deletion of the long arm of chromosome 11, del(11q), is a rare but recurrent clonal event in MDS. Here, we detail the largest series of 113 cases of MDS and myelodysplastic syndromes/myeloproliferative neoplasms (MDS/MPN) harboring a del(11q) analyzed at clinical, cytological, cytogenetic and molecular levels. Female predominance, a survival prognosis similar to other MDS, a low monocyte count and dysmegakaryopoiesis were the specific clinical and cytological features of del(11q) MDS. In most cases, del(11q) was isolated, primary and interstitial encompassing the 11q22-23 region containing ATM, KMT2A and CBL genes. The common deleted region at 11q23.2 is centered on an intergenic region between CADM1 (also known as TSLC1, Tumour Suppressor in Lung Cancer 1) and NXPE2. CADM1 was expressed in all myeloid cells analyzed in contrast to NXPE2. At the functional level, the deletion of Cadm1 in murine Lineage-Sca1+Kit+ cells modifies the lymphoid to myeloid ratio in bone marrow although not altering their multi-lineage hematopoietic reconstitution potential after syngenic transplantation. Together with the frequent simultaneous deletions of KMT2A, ATM and CBL and mutations of ASXL1, SF3B1 and CBL, we show that CADM1 may be important in the physiopathology of the del(11q) MDS, extending its role as tumor-suppressor gene from solid tumors to hematopoietic malignancies.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1182/bloodadvances.2021005311DOI Listing
October 2021

Analysis of T cell receptor clonotypes in tumor microenvironment identifies shared cancer-type-specific signatures.

Cancer Immunol Immunother 2021 Sep 27. Epub 2021 Sep 27.

Cancer Therapeutics Research Laboratory, National Cancer Centre Singapore, 11 Hospital Crescent, Singapore, 169610, Singapore.

Despite the conventional view that a truly random V(D)J recombination process should generate a highly diverse immune repertoire, emerging reports suggest that there is a certain bias toward the generation of shared/public immune receptor chains. These studies were performed in viral diseases where public T cell receptors (TCR) appear to confer better protective responses. Selective pressures generating common TCR clonotypes are currently not well understood, but it is believed that they confer a growth advantage. As very little is known about public TCR clonotypes in cancer, here we set out to determine the extent of shared TCR clonotypes in the intra-tumor microenvironments of virus- and non-virus-driven head and neck cancers using TCR sequencing. We report that tumor-infiltrating T cell clonotypes were indeed shared across individuals with the same cancer type, where the majority of shared sequences were specific to the cancer type (i.e., viral versus non-viral). These shared clonotypes were not particularly enriched in EBV-associated nasopharynx cancer but, in both cancers, exhibited distinct characteristics, namely shorter CDR3 lengths, restricted V- and J-gene usages, and also demonstrated convergent V(D)J recombination. Many of these shared TCRs were expressed in patients with a shared HLA background. Pattern recognition of CDR3 amino acid sequences revealed strong convergence to specific pattern motifs, and these motifs were uniquely found to each cancer type. This suggests that they may be enriched for specificity to common antigens found in the tumor microenvironment of different cancers. The identification of shared TCRs in infiltrating tumor T cells not only adds to our understanding of the tumor-adaptive immune recognition but could also serve as disease-specific biomarkers and guide the development of future immunotherapies.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1007/s00262-021-03047-7DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8476067PMC
September 2021

Mutational synergy during leukemia induction remodels chromatin accessibility, histone modifications and three-dimensional DNA topology to alter gene expression.

Nat Genet 2021 Oct 23;53(10):1443-1455. Epub 2021 Sep 23.

Wellcome - MRC Cambridge Stem Cell Institute, Cambridge, UK.

Altered transcription is a cardinal feature of acute myeloid leukemia (AML); however, exactly how mutations synergize to remodel the epigenetic landscape and rewire three-dimensional DNA topology is unknown. Here, we apply an integrated genomic approach to a murine allelic series that models the two most common mutations in AML: Flt3-ITD and Npm1c. We then deconvolute the contribution of each mutation to alterations of the epigenetic landscape and genome organization, and infer how mutations synergize in the induction of AML. Our studies demonstrate that Flt3-ITD signals to chromatin to alter the epigenetic environment and synergizes with mutations in Npm1c to alter gene expression and drive leukemia induction. These analyses also allow the identification of long-range cis-regulatory circuits, including a previously unknown superenhancer of Hoxa locus, as well as larger and more detailed gene-regulatory networks, driven by transcription factors including PU.1 and IRF8, whose importance we demonstrate through perturbation of network members.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/s41588-021-00925-9DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7611829PMC
October 2021

METTL1-mediated mG modification of Arg-TCT tRNA drives oncogenic transformation.

Mol Cell 2021 08 4;81(16):3323-3338.e14. Epub 2021 Aug 4.

Stem Cell Program, Division of Hematology/Oncology, Boston Children's Hospital, Boston, MA 02115, USA; Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, Boston, MA 02115, USA; Division of Hematology/Oncology, Boston Children's Hospital, Boston, MA 02115, USA; Department of Pediatrics, Harvard Medical School, Boston, MA 02115, USA; Harvard Stem Cell Institute, Cambridge, MA 02138, USA; Harvard Initiative for RNA Medicine, Boston, MA 02115, USA. Electronic address:

The emerging "epitranscriptomics" field is providing insights into the biological and pathological roles of different RNA modifications. The RNA methyltransferase METTL1 catalyzes N7-methylguanosine (mG) modification of tRNAs. Here we find METTL1 is frequently amplified and overexpressed in cancers and is associated with poor patient survival. METTL1 depletion causes decreased abundance of mG-modified tRNAs and altered cell cycle and inhibits oncogenicity. Conversely, METTL1 overexpression induces oncogenic cell transformation and cancer. Mechanistically, we find increased abundance of mG-modified tRNAs, in particular Arg-TCT-4-1, and increased translation of mRNAs, including cell cycle regulators that are enriched in the corresponding AGA codon. Accordingly, Arg-TCT expression is elevated in many tumor types and is associated with patient survival, and strikingly, overexpression of this individual tRNA induces oncogenic transformation. Thus, METTL1-mediated tRNA modification drives oncogenic transformation through a remodeling of the mRNA "translatome" to increase expression of growth-promoting proteins and represents a promising anti-cancer target.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.molcel.2021.06.031DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8380730PMC
August 2021

and Mutations Exert Divergent Effects on DNA Repair and Sensitivity of Leukemia Cells to PARP Inhibitors.

Cancer Res 2021 Oct 2;81(19):5089-5101. Epub 2021 Jul 2.

Wellcome-MRC Cambridge Stem Cell Institute, University of Cambridge, Cambridge, United Kingdom.

Somatic variants in and are founding mutations in hematological malignancies that affect the epigenetic regulation of DNA methylation. Mutations in both genes often co-occur with activating mutations in genes encoding oncogenic tyrosine kinases such as , and , or with mutations affecting related signaling pathways such as and . Here, we show that and mutations exert divergent roles in regulating DNA repair activities in leukemia cells expressing these oncogenes. Malignant TET2-deficient cells displayed downregulation of BRCA1 and LIG4, resulting in reduced activity of BRCA1/2-mediated homologous recombination (HR) and DNA-PK-mediated non-homologous end-joining (D-NHEJ), respectively. TET2-deficient cells relied on PARP1-mediated alternative NHEJ (Alt-NHEJ) for protection from the toxic effects of spontaneous and drug-induced DNA double-strand breaks. Conversely, DNMT3A-deficient cells favored HR/D-NHEJ owing to downregulation of PARP1 and reduction of Alt-NHEJ. Consequently, malignant TET2-deficient cells were sensitive to PARP inhibitor (PARPi) treatment and , whereas DNMT3A-deficient cells were resistant. Disruption of TET2 dioxygenase activity or TET2-Wilms' tumor 1 (WT1)-binding ability was responsible for DNA repair defects and sensitivity to PARPi associated with TET2 deficiency. Moreover, mutation or deletion of mimicked the effect of mutation on DSB repair activity and sensitivity to PARPi. Collectively, these findings reveal that and mutations may serve as biomarkers of synthetic lethality triggered by PARPi, which should be explored therapeutically. SIGNIFICANCE: and mutations affect distinct DNA repair mechanisms and govern the differential sensitivities of oncogenic tyrosine kinase-positive malignant hematopoietic cells to PARP inhibitors.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1158/0008-5472.CAN-20-3761DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8487956PMC
October 2021

SETBP1 overexpression acts in the place of class-defining mutations to drive FLT3-ITD-mutant AML.

Blood Adv 2021 05;5(9):2412-2425

Wellcome Sanger Institute, Wellcome Genome Campus, Cambridge, United Kingdom.

Advances in cancer genomics have revealed genomic classes of acute myeloid leukemia (AML) characterized by class-defining mutations, such as chimeric fusion genes or in genes such as NPM1, MLL, and CEBPA. These class-defining mutations frequently synergize with internal tandem duplications in FLT3 (FLT3-ITDs) to drive leukemogenesis. However, ∼20% of FLT3-ITD-positive AMLs bare no class-defining mutations, and mechanisms of leukemic transformation in these cases are unknown. To identify pathways that drive FLT3-ITD mutant AML in the absence of class-defining mutations, we performed an insertional mutagenesis (IM) screening in Flt3-ITD mice, using Sleeping Beauty transposons. All mice developed acute leukemia (predominantly AML) after a median of 73 days. Analysis of transposon insertions in 38 samples from Flt3-ITD/IM leukemic mice identified recurrent integrations at 22 loci, including Setbp1 (20/38), Ets1 (11/38), Ash1l (8/38), Notch1 (8/38), Erg (7/38), and Runx1 (5/38). Insertions at Setbp1 led exclusively to AML and activated a transcriptional program similar, but not identical, to those of NPM1-mutant and MLL-rearranged AMLs. Guide RNA targeting of Setbp1 was highly detrimental to Flt3ITD/+/Setbp1IM+, but not to Flt3ITD/+/Npm1cA/+, AMLs. Also, analysis of RNA-sequencing data from hundreds of human AMLs revealed that SETBP1 expression is significantly higher in FLT3-ITD AMLs lacking class-defining mutations. These findings propose that SETBP1 overexpression collaborates with FLT3-ITD to drive a subtype of human AML. To identify genetic vulnerabilities of these AMLs, we performed genome-wide CRISPR-Cas9 screening in Flt3ITD/+/Setbp1IM+ AMLs and identified potential therapeutic targets, including Kdm1a, Brd3, Ezh2, and Hmgcr. Our study gives new insights into epigenetic pathways that can drive AMLs lacking class-defining mutations and proposes therapeutic approaches against such cases.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1182/bloodadvances.2020003443DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8114559PMC
May 2021

Small-molecule inhibition of METTL3 as a strategy against myeloid leukaemia.

Nature 2021 May 26;593(7860):597-601. Epub 2021 Apr 26.

Milner Therapeutics Institute, University of Cambridge, Cambridge, UK.

N-methyladenosine (mA) is an abundant internal RNA modification that is catalysed predominantly by the METTL3-METTL14 methyltransferase complex. The mA 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. 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 mA 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.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/s41586-021-03536-wDOI Listing
May 2021

The PML-RARA fusion is not detectable in historical blood samples of acute promyelocytic leukaemia patients.

Ann Hematol 2021 Mar 1. Epub 2021 Mar 1.

Wellcome-MRC Cambridge Stem Cell Institute, Jeffrey Cheah Biomedical Centre, Puddicombe Way, Cambridge, CB2 0AW, UK.

View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1007/s00277-021-04472-5DOI Listing
March 2021

meCLICK-Seq, a Substrate-Hijacking and RNA Degradation Strategy for the Study of RNA Methylation.

ACS Cent Sci 2020 Dec 29;6(12):2196-2208. Epub 2020 Oct 29.

Department of Chemistry, University of Cambridge, Lensfield Road, Cambridge CB2 1EW, U.K.

The fates of RNA species in a cell are controlled by ribonucleases, which degrade them by exploiting the universal structural 2'-OH group. This phenomenon plays a key role in numerous transformative technologies, for example, RNA interference and CRISPR/Cas13-based RNA editing systems. These approaches, however, are genetic or oligomer-based and so have inherent limitations. This has led to interest in the development of small molecules capable of degrading nucleic acids in a targeted manner. Here we describe click-degraders, small molecules that can be covalently attached to RNA species through click-chemistry and can degrade them, that are akin to ribonucleases. By using these molecules, we have developed the meCLICK-Seq (methylation CLICK-degradation Sequencing) a method to identify RNA modification substrates with high resolution at intronic and intergenic regions. The method hijacks RNA methyltransferase activity to introduce an alkyne, instead of a methyl, moiety on RNA. Subsequent copper(I)-catalyzed azide-alkyne cycloaddition reaction with the click-degrader leads to RNA cleavage and degradation exploiting a mechanism used by endogenous ribonucleases. Focusing on N-methyladenosine (mA), meCLICK-Seq identifies methylated transcripts, determines RNA methylase specificity, and reliably maps modification sites in intronic and intergenic regions. Importantly, we show that METTL16 deposits mA to intronic polyadenylation (IPA) sites, which suggests a potential role for METTL16 in IPA and, in turn, splicing. Unlike other methods, the readout of meCLICK-Seq is depletion, not enrichment, of modified RNA species, which allows a comprehensive and dynamic study of RNA modifications throughout the transcriptome, including regions of low abundance. The click-degraders are highly modular and so may be exploited to study any RNA modification and design new technologies that rely on RNA degradation.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1021/acscentsci.0c01094DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7760485PMC
December 2020

Counteracts Anti-Leukemic and Stem Cell Inhibitory Effects of Retinoic Acid on -ITD/-Driven Acute Myeloid Leukemia Cells.

Biomedicines 2020 Sep 28;8(10). Epub 2020 Sep 28.

Division of Oncology, Department of Medicine I, Medical University of Vienna, 1090 Vienna, Austria.

retinoic acid (atRA) has a dramatic impact on the survival of patients with acute promyelocytic leukemia, but its therapeutic value in other types of acute myeloid leukemia (AML) has so far remained unclear. Given that AML is a stem cell-driven disease, recent studies have addressed the effects of atRA on leukemic stem cells (LSCs). atRA promoted stemness of -driven AML in an -dependent manner but had the opposite effect in -ITD/-driven AML. Overexpression of the stem cell-associated transcription factor predicts a poor prognosis in AML, and is observed in different genetic subtypes, including cytogenetically normal AML. Here, we therefore investigated the effects of in a mouse model for cytogenetically normal AML, which rests on the combined activity of -ITD and mutations. Experimental expression of on this background strongly promoted disease aggressiveness. atRA inhibited leukemia cell viability and stem cell-related properties, and these effects were counteracted by overexpression of . These data further underscore the complexity of the responsiveness of AML LSCs to atRA and point out the need for additional investigations which may lay a foundation for a precision medicine-based use of retinoids in AML.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.3390/biomedicines8100385DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7600968PMC
September 2020

Promotes Aggressiveness and Stem Cell-Related Properties of Acute Myeloid Leukemia.

Cancer Res 2020 10 1;80(20):4527-4539. Epub 2020 Sep 1.

Division of Oncology, Department of Medicine I, Medical University of Vienna, Vienna, Austria.

Overexpression of , which encodes the alpha chain of the IL2 receptor, is associated with chemotherapy resistance and poor outcome in acute myeloid leukemia (AML). The clinical potential of anti-IL2RA therapy is, therefore, being explored in early-stage clinical trials. Notwithstanding, only very limited information regarding the biological function of in AML is available. Using genetic manipulation of expression as well as antibody-mediated inhibition of IL2RA in human cell lines, mouse models, and primary patient samples, we investigated the effects of on AML cell proliferation and apoptosis, and on pertinent signaling pathways. The impact of on the properties of leukemic stem cells (LSC) and on leukemogenesis were queried. promoted proliferation and cell-cycle activity and inhibited apoptosis in human AML cell lines and primary cells. These phenotypes were accompanied by corresponding alterations in cell-cycle machinery and in pathways associated with cell survival and apoptosis. The biological roles of were confirmed in two genetically distinct AML mouse models, revealing that inhibits differentiation, promotes stem cell-related properties, and is required for leukemogenesis. IL2RA antibodies inhibited leukemic, but not normal, hematopoietic cells and synergized with other antileukemic agents in this regard. Collectively, these data show for the first time that plays key biological roles in AML and underscore its value as a potential therapeutic target in this disease. SIGNIFICANCE: This study identifies as a potential therapeutic target in AML, where it is shown to regulate proliferation, differentiation, apoptosis, stem cell-related properties, and leukemogenesis.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1158/0008-5472.CAN-20-0531DOI Listing
October 2020

Synergistic targeting of FLT3 mutations in AML via combined menin-MLL and FLT3 inhibition.

Blood 2020 11;136(21):2442-2456

Department of Hematology, Medical Oncology, and Pulmonary Medicine, University Medical Center, Johannes Gutenberg-University, Mainz, Germany.

The interaction of menin (MEN1) and MLL (MLL1, KMT2A) is a dependency and provides a potential opportunity for treatment of NPM1-mutant (NPM1mut) and MLL-rearranged (MLL-r) leukemias. Concomitant activating driver mutations in the gene encoding the tyrosine kinase FLT3 occur in both leukemias and are particularly common in the NPM1mut subtype. In this study, transcriptional profiling after pharmacological inhibition of the menin-MLL complex revealed specific changes in gene expression, with downregulation of the MEIS1 transcription factor and its transcriptional target gene FLT3 being the most pronounced. Combining menin-MLL inhibition with specific small-molecule kinase inhibitors of FLT3 phosphorylation resulted in a significantly superior reduction of phosphorylated FLT3 and transcriptional suppression of genes downstream of FLT3 signaling. The drug combination induced synergistic inhibition of proliferation, as well as enhanced apoptosis, compared with single-drug treatment in models of human and murine NPM1mut and MLL-r leukemias harboring an FLT3 mutation. Primary acute myeloid leukemia (AML) cells harvested from patients with NPM1mutFLT3mut AML showed significantly better responses to combined menin and FLT3 inhibition than to single-drug or vehicle control treatment, whereas AML cells with wild-type NPM1, MLL, and FLT3 were not affected by either of the 2 drugs. In vivo treatment of leukemic animals with MLL-r FLT3mut leukemia reduced leukemia burden significantly and prolonged survival compared with results in the single-drug and vehicle control groups. Our data suggest that combined menin-MLL and FLT3 inhibition represents a novel and promising therapeutic strategy for patients with NPM1mut or MLL-r leukemia and concurrent FLT3 mutation.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1182/blood.2020005037DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8215191PMC
November 2020

Home and away: clonal hematopoiesis in sibling transplants.

Blood 2020 04;135(18):1511-1512

University of Cambridge.

View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1182/blood.2020005717DOI Listing
April 2020

Dissecting the early steps of MLL induced leukaemogenic transformation using a mouse model of AML.

Nat Commun 2020 03 16;11(1):1407. Epub 2020 Mar 16.

Wellcome and MRC Cambridge Stem Cell Institute and University of Cambridge Department of Haematology, Jeffrey Cheah Biomedical Centre, Puddicombe Way, Cambridge, CB2 0AW, UK.

Leukaemogenic mutations commonly disrupt cellular differentiation and/or enhance proliferation, thus perturbing the regulatory programs that control self-renewal and differentiation of stem and progenitor cells. Translocations involving the Mll1 (Kmt2a) gene generate powerful oncogenic fusion proteins, predominantly affecting infant and paediatric AML and ALL patients. The early stages of leukaemogenic transformation are typically inaccessible from human patients and conventional mouse models. Here, we take advantage of cells conditionally blocked at the multipotent haematopoietic progenitor stage to develop a MLL-r model capturing early cellular and molecular consequences of MLL-ENL expression based on a clear clonal relationship between parental and leukaemic cells. Through a combination of scRNA-seq, ATAC-seq and genome-scale CRISPR-Cas9 screening, we identify pathways and genes likely to drive the early phases of leukaemogenesis. Finally, we demonstrate the broad utility of using matched parental and transformed cells for small molecule inhibitor studies by validating both previously known and other potential therapeutic targets.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/s41467-020-15220-0DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7075888PMC
March 2020

Therapeutic targeting of preleukemia cells in a mouse model of mutant acute myeloid leukemia.

Science 2020 01;367(6477):586-590

Department of Pediatric Oncology, Dana-Farber Cancer Institute, and Division of Hematology/Oncology, Boston, MA, USA.

The initiating mutations that contribute to cancer development are sometimes present in premalignant cells. Whether therapies targeting these mutations can eradicate premalignant cells is unclear. Acute myeloid leukemia (AML) is an attractive system for investigating the effect of preventative treatment because this disease is often preceded by a premalignant state (clonal hematopoiesis or myelodysplastic syndrome). In mutant knock-in mice, a model of AML development, leukemia is preceded by a period of extended myeloid progenitor cell proliferation and self-renewal. We found that this self-renewal can be reversed by oral administration of a small molecule (VTP-50469) that targets the MLL1-Menin chromatin complex. These preclinical results support the hypothesis that individuals at high risk of developing AML might benefit from targeted epigenetic therapy in a preventative setting.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1126/science.aax5863DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7754791PMC
January 2020

Vitamin D Receptor Controls Cell Stemness in Acute Myeloid Leukemia and in Normal Bone Marrow.

Cell Rep 2020 01;30(3):739-754.e4

INSERM UMR 1163, Laboratory of Cellular and Molecular Mechanisms of Hematological Disorders and Therapeutical Implications, 75015 Paris, France; Paris Descartes - Sorbonne Paris Cité University, Imagine Institute, 75015 Paris, France; CNRS ERL 8254, Laboratory of Cellular and Molecular Mechanisms of Hematological Disorders and Therapeutical Implications, 75015 Paris, France; Laboratory of Excellence GR-Ex, 75015 Paris, France.

Vitamin D (VD) is a known differentiating agent, but the role of VD receptor (VDR) is still incompletely described in acute myeloid leukemia (AML), whose treatment is based mostly on antimitotic chemotherapy. Here, we present an unexpected role of VDR in normal hematopoiesis and in leukemogenesis. Limited VDR expression is associated with impaired myeloid progenitor differentiation and is a new prognostic factor in AML. In mice, the lack of Vdr results in increased numbers of hematopoietic and leukemia stem cells and quiescent hematopoietic stem cells. In addition, malignant transformation of Vdr cells results in myeloid differentiation block and increases self-renewal. Vdr promoter is methylated in AML as in CD34 cells, and demethylating agents induce VDR expression. Association of VDR agonists with hypomethylating agents promotes leukemia stem cell exhaustion and decreases tumor burden in AML mouse models. Thus, Vdr functions as a regulator of stem cell homeostasis and leukemic propagation.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.celrep.2019.12.055DOI Listing
January 2020

Analysis pipelines for cancer genome sequencing in mice.

Nat Protoc 2020 02 6;15(2):266-315. Epub 2020 Jan 6.

Institute of Molecular Oncology and Functional Genomics, School of Medicine, Technische Universität München, Munich, Germany.

Mouse models of human cancer have transformed our ability to link genetics, molecular mechanisms and phenotypes. Both reverse and forward genetics in mice are currently gaining momentum through advances in next-generation sequencing (NGS). Methodologies to analyze sequencing data were, however, developed for humans and hence do not account for species-specific differences in genome structures and experimental setups. Here, we describe standardized computational pipelines specifically tailored to the analysis of mouse genomic data. We present novel tools and workflows for the detection of different alteration types, including single-nucleotide variants (SNVs), small insertions and deletions (indels), copy-number variations (CNVs), loss of heterozygosity (LOH) and complex rearrangements, such as in chromothripsis. Workflows have been extensively validated and cross-compared using multiple methodologies. We also give step-by-step guidance on the execution of individual analysis types, provide advice on data interpretation and make the complete code available online. The protocol takes 2-7 d, depending on the desired analyses.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/s41596-019-0234-7DOI Listing
February 2020

The curious incident of TdT-mediated mutations in AML.

Blood 2019 12;134(25):2229-2231

University of Cambridge.

View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1182/blood.2019003619DOI Listing
December 2019

HBO1 is required for the maintenance of leukaemia stem cells.

Nature 2020 01 11;577(7789):266-270. Epub 2019 Dec 11.

Cancer Therapeutics CRC, Melbourne, Victoria, Australia.

Acute myeloid leukaemia (AML) is a heterogeneous disease characterized by transcriptional dysregulation that results in a block in differentiation and increased malignant self-renewal. Various epigenetic therapies aimed at reversing these hallmarks of AML have progressed into clinical trials, but most show only modest efficacy owing to an inability to effectively eradicate leukaemia stem cells (LSCs). Here, to specifically identify novel dependencies in LSCs, we screened a bespoke library of small hairpin RNAs that target chromatin regulators in a unique ex vivo mouse model of LSCs. We identify the MYST acetyltransferase HBO1 (also known as KAT7 or MYST2) and several known members of the HBO1 protein complex as critical regulators of LSC maintenance. Using CRISPR domain screening and quantitative mass spectrometry, we identified the histone acetyltransferase domain of HBO1 as being essential in the acetylation of histone H3 at K14. H3 acetylated at K14 (H3K14ac) facilitates the processivity of RNA polymerase II to maintain the high expression of key genes (including Hoxa9 and Hoxa10) that help to sustain the functional properties of LSCs. To leverage this dependency therapeutically, we developed a highly potent small-molecule inhibitor of HBO1 and demonstrate its mode of activity as a competitive analogue of acetyl-CoA. Inhibition of HBO1 phenocopied our genetic data and showed efficacy in a broad range of human cell lines and primary AML cells from patients. These biological, structural and chemical insights into a therapeutic target in AML will enable the clinical translation of these findings.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/s41586-019-1835-6DOI Listing
January 2020

Concordance for clonal hematopoiesis is limited in elderly twins.

Blood 2020 01;135(4):269-273

Wellcome Sanger Institute, Wellcome Genome Campus, Hinxton, Cambridge, United Kingdom.

Although acquisition of leukemia-associated somatic mutations by 1 or more hematopoietic stem cells is inevitable with advancing age, its consequences are highly variable, ranging from clinically silent clonal hematopoiesis (CH) to leukemic progression. To investigate the influence of heritable factors on CH, we performed deep targeted sequencing of blood DNA from 52 monozygotic (MZ) and 27 dizygotic (DZ) twin pairs (aged 70-99 years). Using this highly sensitive approach, we identified CH (variant allele frequency ≥0.5%) in 62% of individuals. We did not observe higher concordance for CH within MZ twin pairs as compared with that within DZ twin pairs, or to that expected by chance. However, we did identify 2 MZ pairs in which both twins harbored identical rare somatic mutations, suggesting a shared cell of origin. Finally, in 3 MZ twin pairs harboring mutations in the same driver genes, serial blood samples taken 4 to 5 years apart showed substantial twin-to-twin variability in clonal trajectories. Our findings propose that the inherited genome does not exert a dominant influence on the behavior of adult CH and provide evidence that CH mutations may be acquired in utero.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1182/blood.2019001807DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6978156PMC
January 2020

Genetic modification of primary human B cells to model high-grade lymphoma.

Nat Commun 2019 10 4;10(1):4543. Epub 2019 Oct 4.

Wellcome MRC Cambridge Stem Cell Institute, Cambridge, CB2 0AW, UK.

Sequencing studies of diffuse large B cell lymphoma (DLBCL) have identified hundreds of recurrently altered genes. However, it remains largely unknown whether and how these mutations may contribute to lymphomagenesis, either individually or in combination. Existing strategies to address this problem predominantly utilize cell lines, which are limited by their initial characteristics and subsequent adaptions to prolonged in vitro culture. Here, we describe a co-culture system that enables the ex vivo expansion and viral transduction of primary human germinal center B cells. Incorporation of CRISPR/Cas9 technology enables high-throughput functional interrogation of genes recurrently mutated in DLBCL. Using a backbone of BCL2 with either BCL6 or MYC, we identify co-operating genetic alterations that promote growth or even full transformation into synthetically engineered DLBCL models. The resulting tumors can be expanded and sequentially transplanted in vivo, providing a scalable platform to test putative cancer genes and to create mutation-directed, bespoke lymphoma models.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/s41467-019-12494-xDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6778131PMC
October 2019

Triple-mutant AML: too clever by HLF?

Blood 2019 07;134(3):222-224

University of Cambridge.

View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1182/blood.2019001533DOI Listing
July 2019

SOCS2 is part of a highly prognostic 4-gene signature in AML and promotes disease aggressiveness.

Sci Rep 2019 06 24;9(1):9139. Epub 2019 Jun 24.

Department of Medicine I, Division of Oncology, Medical University of Vienna, Vienna, Austria.

Acute myeloid leukemia (AML) is a heterogeneous disease with respect to its genetic and molecular basis and to patients´ outcome. Clinical, cytogenetic, and mutational data are used to classify patients into risk groups with different survival, however, within-group heterogeneity is still an issue. Here, we used a robust likelihood-based survival modeling approach and publicly available gene expression data to identify a minimal number of genes whose combined expression values were prognostic of overall survival. The resulting gene expression signature (4-GES) consisted of 4 genes (SOCS2, IL2RA, NPDC1, PHGDH), predicted patient survival as an independent prognostic parameter in several cohorts of AML patients (total, 1272 patients), and further refined prognostication based on the European Leukemia Net classification. An oncogenic role of the top scoring gene in this signature, SOCS2, was investigated using MLL-AF9 and Flt3-ITD/NPM1c driven mouse models of AML. SOCS2 promoted leukemogenesis as well as the abundance, quiescence, and activity of AML stem cells. Overall, the 4-GES represents a highly discriminating prognostic parameter in AML, whose clinical applicability is greatly enhanced by its small number of genes. The newly established role of SOCS2 in leukemia aggressiveness and stemness raises the possibility that the signature might even be exploitable therapeutically.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/s41598-019-45579-0DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6591510PMC
June 2019

NPM1c alters FLT3-D835Y localization and signaling in acute myeloid leukemia.

Blood 2019 07 11;134(4):383-388. Epub 2019 Jun 11.

Department of Hematology and Oncology, Freiburg University Medical Center, Albert-Ludwigs-University of Freiburg, Freiburg, Germany.

Activating mutations in FMS-like tyrosine kinase receptor-3 (FLT3) and Nucleophosmin-1 (NPM1) are most frequent alterations in acute myeloid leukemia (AML), and are often coincidental. The mutational status of NPM1 has strong prognostic relevance to patients with point mutations of the FLT3 tyrosine kinase domain (TKD), but the biological mechanism underlying this effect remains unclear. In the present study, we investigated the effect of the coincidence of NPM1c and FLT3-TKD. Although expression of FLT3-TKD is not sufficient to induce a disease in mice, coexpression with NPM1c rapidly leads to an aggressive myeloproliferative disease in mice with a latency of 31.5 days. Mechanistically, we could show that FLT3-TKD is able to activate the downstream effector molecule signal transducer and activator of transcription 5 (STAT5) exclusively in the presence of mutated NPM1c. Moreover, NPM1c alters the cellular localization of FLT3-TKD from the cell surface to the endoplasmic reticulum, which might thereby lead to the aberrant STAT5 activation. Importantly, aberrant STAT5 activation occurs not only in primary murine cells but also in patients with AML with combined FLT3-TKD and NPM1c mutations. Thus, our data indicate a new mechanism, how NPM1c mislocalizes FLT3-TKD and changes its signal transduction ability.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1182/blood.2018883140DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6659255PMC
July 2019

Genome-scale drop-out screens to identify cancer cell vulnerabilities in AML.

Curr Opin Genet Dev 2019 02 4;54:83-87. Epub 2019 May 4.

Wellcome-MRC Cambridge Stem Cell Institute, Department of Haematology, University of Cambridge, Hills Road, Cambridge, CB2 0AW, United Kingdom; Haematological Cancer Genetics, Wellcome Trust Sanger Institute, Hinxton, Cambridge, CB10 1SA, United Kingdom. Electronic address:

Acute myeloid leukemia (AML) is an aggressive cancer that remains lethal to the majority of sufferers. Whilst the mainstay treatments for this condition have remained largely unchanged over the past five decades, progress in deciphering its pathogenesis has accelerated in recent years, propelled in part by advances in cancer genomics and mechanistic studies of leukemogenic mutations. Newer molecular therapies targeting aberrant biological pathways are currently under investigation with a few moving closer to clinical use. However, collectively, these new therapies are not predicted to have a major impact on clinical outcomes and the need for the identification of further therapeutic targets in AML remains critical. Recently the use of CRISPR-Cas9 systems for genome editing and their potential application in genome-wide screening has opened a new frontier for unbiased discovery of therapeutic vulnerabilities in cancer and AML was the first disease in which this technology was systematically applied. In this review we give an overview of recent advances in identifying novel therapeutic vulnerabilities of AML using CRISPR-Cas9 and discuss possible future applications of CRISPR technologies in this field.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.gde.2019.04.004DOI Listing
February 2019

PiggyBac transposon tools for recessive screening identify B-cell lymphoma drivers in mice.

Nat Commun 2019 03 29;10(1):1415. Epub 2019 Mar 29.

The Wellcome Trust Sanger Institute, Genome Campus, Hinxton, Cambridge, CB10 1SA, UK.

B-cell lymphoma (BCL) is the most common hematologic malignancy. While sequencing studies gave insights into BCL genetics, identification of non-mutated cancer genes remains challenging. Here, we describe PiggyBac transposon tools and mouse models for recessive screening and show their application to study clonal B-cell lymphomagenesis. In a genome-wide screen, we discover BCL genes related to diverse molecular processes, including signaling, transcriptional regulation, chromatin regulation, or RNA metabolism. Cross-species analyses show the efficiency of the screen to pinpoint human cancer drivers altered by non-genetic mechanisms, including clinically relevant genes dysregulated epigenetically, transcriptionally, or post-transcriptionally in human BCL. We also describe a CRISPR/Cas9-based in vivo platform for BCL functional genomics, and validate discovered genes, such as Rfx7, a transcription factor, and Phip, a chromatin regulator, which suppress lymphomagenesis in mice. Our study gives comprehensive insights into the molecular landscapes of BCL and underlines the power of genome-scale screening to inform biology.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/s41467-019-09180-3DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6440946PMC
March 2019

Contrasting requirements during disease evolution identify EZH2 as a therapeutic target in AML.

J Exp Med 2019 04 19;216(4):966-981. Epub 2019 Mar 19.

Wellcome Trust-Medical Research Council Cambridge Stem Cell Institute, Cambridge, UK

Epigenetic regulators, such as EZH2, are frequently mutated in cancer, and loss-of-function mutations are common in myeloid malignancies. We have examined the importance of cellular context for Ezh2 loss during the evolution of acute myeloid leukemia (AML), where we observed stage-specific and diametrically opposite functions for Ezh2 at the early and late stages of disease. During disease maintenance, WT Ezh2 exerts an oncogenic function that may be therapeutically targeted. In contrast, Ezh2 acts as a tumor suppressor during AML induction. Transcriptional analysis explains this apparent paradox, demonstrating that loss of derepresses different expression programs during disease induction and maintenance. During disease induction, loss derepresses a subset of bivalent promoters that resolve toward gene activation, inducing a feto-oncogenic program that includes genes such as , whose overexpression phenocopies loss to accelerate AML induction in mouse models. Our data highlight the importance of cellular context and disease phase for the function of Ezh2 and its potential therapeutic implications.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1084/jem.20181276DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6446874PMC
April 2019

TET2 binding to enhancers facilitates transcription factor recruitment in hematopoietic cells.

Genome Res 2019 04 22;29(4):564-575. Epub 2019 Feb 22.

Biotech Research and Innovation Centre (BRIC), Faculty of Health and Medical Sciences, University of Copenhagen, 2200 Copenhagen, Denmark.

The epigenetic regulator is frequently mutated in hematological diseases. Mutations have been shown to arise in hematopoietic stem cells early in disease development and lead to altered DNA methylation landscapes and an increased risk of hematopoietic malignancy. Here, we show by genome-wide mapping of TET2 binding sites in different cell types that TET2 localizes to regions of open chromatin and cell-type-specific enhancers. We find that deletion of in native hematopoiesis as well as fully transformed acute myeloid leukemia (AML) results in changes in transcription factor (TF) activity within these regions, and we provide evidence that loss of TET2 leads to attenuation of chromatin binding of members of the basic helix-loop-helix (bHLH) TF family. Together, these findings demonstrate that TET2 activity shapes the local chromatin environment at enhancers to facilitate TF binding and provides an example of how epigenetic dysregulation can affect gene expression patterns and drive disease development.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1101/gr.239277.118DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6442383PMC
April 2019
-->