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1. Developmental interplay between transcriptional alterations and a targetable cytokine signaling dependency in pediatric ETO2::GLIS2 leukemia

Author

Verónica Alonso-Pérez, Klaudia Galant, Fabien Boudia, Elie Robert, Zakia Aid, Laurent Renou, Vilma Barroca, Saryiami Devanand, Loélia Babin, Virginie Rouiller-Fabre, Delphine Moison, Didier Busso, Guillaume Piton, Christophe Metereau, Nassera Abermil, Paola Ballerini, Pierre Hirsch, Rima Haddad, Jelena Martinovic, Arnaud Petit, Hélène Lapillonne, Erika Brunet, Thomas Mercher, and Françoise Pflumio

Subjects
Neoplasms. Tumors. Oncology. Including cancer and carcinogens, RC254-282
Abstract

Abstract Background Several fusion oncogenes showing a higher incidence in pediatric acute myeloid leukemia (AML) are associated with heterogeneous megakaryoblastic and other myeloid features. Here we addressed how developmental mechanisms influence human leukemogenesis by ETO2::GLIS2, associated with dismal prognosis. Methods We created novel ETO2::GLIS2 models of leukemogenesis through lentiviral transduction and CRISPR-Cas9 gene editing of human fetal and post-natal hematopoietic stem/progenitor cells (HSPCs), performed in-depth characterization of ETO2::GLIS2 transformed cells through multiple omics and compared them to patient samples. This led to a preclinical assay using patient-derived-xenograft models to test a combination of two clinically-relevant molecules. Results We showed that ETO2::GLIS2 expression in primary human fetal CD34+ hematopoietic cells led to more efficient in vivo leukemia development than expression in post-natal cells. Moreover, cord blood-derived leukemogenesis has a major dependency on the presence of human cytokines, including IL3 and SCF. Single cell transcriptomes revealed that this cytokine environment controlled two ETO2::GLIS2-transformed states that were also observed in primary patient cells. Importantly, this cytokine sensitivity may be therapeutically-exploited as combined MEK and BCL2 inhibition showed higher efficiency than individual molecules to reduce leukemia progression in vivo. Conclusions Our study uncovers an interplay between the cytokine milieu and transcriptional programs that extends a developmental window of permissiveness to transformation by the ETO2::GLIS2 AML fusion oncogene, controls the intratumoral cellular heterogeneity, and offers a ground-breaking therapeutical opportunity by a targeted combination strategy.

Published
2024
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2. Stepwise GATA1 and SMC3 mutations alter megakaryocyte differentiation in a Down syndrome leukemia model

Author

Brahim Arkoun, Elie Robert, Fabien Boudia, Stefania Mazzi, Virginie Dufour, Aurélie Siret, Yasmine Mammasse, Zakia Aid, Matthieu Vieira, Aygun Imanci, Marine Aglave, Marie Cambot, Rachel Petermann, Sylvie Souquere, Philippe Rameau, Cyril Catelain, Romain Diot, Gérard Tachdjian, Olivier Hermine, Nathalie Droin, Najet Debili, Isabelle Plo, Sébastien Malinge, Eric Soler, Hana Raslova, Thomas Mercher, and William Vainchenker

Subjects
Hematology, Oncology, Medicine
Abstract

Acute megakaryoblastic leukemia of Down syndrome (DS-AMKL) is a model of clonal evolution from a preleukemic transient myeloproliferative disorder requiring both a trisomy 21 (T21) and a GATA1s mutation to a leukemia driven by additional driver mutations. We modeled the megakaryocyte differentiation defect through stepwise gene editing of GATA1s, SMC3+/–, and MPLW515K, providing 20 different T21 or disomy 21 (D21) induced pluripotent stem cell (iPSC) clones. GATA1s profoundly reshaped iPSC-derived hematopoietic architecture with gradual myeloid-to-megakaryocyte shift and megakaryocyte differentiation alteration upon addition of SMC3 and MPL mutations. Transcriptional, chromatin accessibility, and GATA1-binding data showed alteration of essential megakaryocyte differentiation genes, including NFE2 downregulation that was associated with loss of GATA1s binding and functionally involved in megakaryocyte differentiation blockage. T21 enhanced the proliferative phenotype, reproducing the cellular and molecular abnormalities of DS-AMKL. Our study provides an array of human cell–based models revealing individual contributions of different mutations to DS-AMKL differentiation blockage, a major determinant of leukemic progression.

Published
2023
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3. The Pediatric Acute Leukemia Fusion Oncogene ETO2-GLIS2 Increases Self-Renewal and Alters Differentiation in a Human Induced Pluripotent Stem Cells-Derived Model

Author

Salvatore Nicola Bertuccio, Fabien Boudia, Marie Cambot, Cécile K. Lopez, Larissa Lordier, Alessandro Donada, Elie Robert, Cécile Thirant, Zakia Aid, Salvatore Serravalle, Annalisa Astolfi, Valentina Indio, Franco Locatelli, Andrea Pession, William Vainchenker, Riccardo Masetti, Hana Raslova, and Thomas Mercher

Subjects
Diseases of the blood and blood-forming organs, RC633-647.5
Published
2020
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4. Supplementary Figures and Legends from Ontogenic Changes in Hematopoietic Hierarchy Determine Pediatric Specificity and Disease Phenotype in Fusion Oncogene–Driven Myeloid Leukemia

Author

Thomas Mercher, Juerg Schwaller, Françoise Pflumio, Arnaud Petit, Berthold Göttgens, Olivier A. Bernard, Isabelle Godin, Camille Lobry, Nathalie Droin, Claus Nerlov, Sébastien Malinge, Franco Locatelli, Riccardo Masetti, Muriel Gaudry, William Vainchenker, Antoine H.F.M. Peters, Eric Delabesse, Jean Luc Villeval, Loélia Babin, Erika Brunet, Yann Lecluse, Bastien Job, M'Boyba Diop, Sarah J. Kinston, Marie-Laure Arcangeli, Alexandre Fagnan, Cécile Thirant, Fabien Boudia, Hélène Lapillonne, Chrystèle Bilhou-Nabera, Paola Ballerini, Zakia Aid, Elie Robert, Vaia Stavropoulou, Esteve Noguera, and Cécile K. Lopez

Abstract

Supplementary Figures and Legends

Published
2023

5. Data from Ontogenic Changes in Hematopoietic Hierarchy Determine Pediatric Specificity and Disease Phenotype in Fusion Oncogene–Driven Myeloid Leukemia

Author

Thomas Mercher, Juerg Schwaller, Françoise Pflumio, Arnaud Petit, Berthold Göttgens, Olivier A. Bernard, Isabelle Godin, Camille Lobry, Nathalie Droin, Claus Nerlov, Sébastien Malinge, Franco Locatelli, Riccardo Masetti, Muriel Gaudry, William Vainchenker, Antoine H.F.M. Peters, Eric Delabesse, Jean Luc Villeval, Loélia Babin, Erika Brunet, Yann Lecluse, Bastien Job, M'Boyba Diop, Sarah J. Kinston, Marie-Laure Arcangeli, Alexandre Fagnan, Cécile Thirant, Fabien Boudia, Hélène Lapillonne, Chrystèle Bilhou-Nabera, Paola Ballerini, Zakia Aid, Elie Robert, Vaia Stavropoulou, Esteve Noguera, and Cécile K. Lopez

Abstract

Fusion oncogenes are prevalent in several pediatric cancers, yet little is known about the specific associations between age and phenotype. We observed that fusion oncogenes, such as ETO2–GLIS2, are associated with acute megakaryoblastic or other myeloid leukemia subtypes in an age-dependent manner. Analysis of a novel inducible transgenic mouse model showed that ETO2–GLIS2 expression in fetal hematopoietic stem cells induced rapid megakaryoblastic leukemia whereas expression in adult bone marrow hematopoietic stem cells resulted in a shift toward myeloid transformation with a strikingly delayed in vivo leukemogenic potential. Chromatin accessibility and single-cell transcriptome analyses indicate ontogeny-dependent intrinsic and ETO2–GLIS2-induced differences in the activities of key transcription factors, including ERG, SPI1, GATA1, and CEBPA. Importantly, switching off the fusion oncogene restored terminal differentiation of the leukemic blasts. Together, these data show that aggressiveness and phenotypes in pediatric acute myeloid leukemia result from an ontogeny-related differential susceptibility to transformation by fusion oncogenes.Significance:This work demonstrates that the clinical phenotype of pediatric acute myeloid leukemia is determined by ontogeny-dependent susceptibility for transformation by oncogenic fusion genes. The phenotype is maintained by potentially reversible alteration of key transcription factors, indicating that targeting of the fusions may overcome the differentiation blockage and revert the leukemic state.See related commentary by Cruz Hernandez and Vyas, p. 1653.This article is highlighted in the In This Issue feature, p. 1631

Published
2023

6. Supplementary Tables 1 to 11 from Ontogenic Changes in Hematopoietic Hierarchy Determine Pediatric Specificity and Disease Phenotype in Fusion Oncogene–Driven Myeloid Leukemia

Author

Thomas Mercher, Juerg Schwaller, Françoise Pflumio, Arnaud Petit, Berthold Göttgens, Olivier A. Bernard, Isabelle Godin, Camille Lobry, Nathalie Droin, Claus Nerlov, Sébastien Malinge, Franco Locatelli, Riccardo Masetti, Muriel Gaudry, William Vainchenker, Antoine H.F.M. Peters, Eric Delabesse, Jean Luc Villeval, Loélia Babin, Erika Brunet, Yann Lecluse, Bastien Job, M'Boyba Diop, Sarah J. Kinston, Marie-Laure Arcangeli, Alexandre Fagnan, Cécile Thirant, Fabien Boudia, Hélène Lapillonne, Chrystèle Bilhou-Nabera, Paola Ballerini, Zakia Aid, Elie Robert, Vaia Stavropoulou, Esteve Noguera, and Cécile K. Lopez

Abstract

Supplementary Tables 1 to 11

Published
2023

7. Stepwise GATA1 and SMC3 mutations alter megakaryocyte differentiation in a Down syndrome leukemia model

Author

Brahim Arkoun, Elie Robert, Fabien Boudia, Stefania Mazzi, Virginie Dufour, Aurélie Siret, Yasmine Mammasse, Zakia Aid, Matthieu Vieira, Imanci Aygun, Marine Aglave, Marie Cambot, Rachel Petermann, Sylvie Souquere, Philippe Rameau, Cyril Catelain, Romain Diot, Gérard Tachdjian, Olivier Hermine, Nathalie Droin, Najet Debili, Isabelle Plo, Sébastien Malinge, Eric Soler, Hana Raslova, Thomas Mercher, and William Vainchenker

Subjects
Chondroitin Sulfate Proteoglycans, Chromosomal Proteins, Non-Histone, Leukemia, Megakaryoblastic, Acute, Mutation, Humans, Cell Cycle Proteins, GATA1 Transcription Factor, Trisomy, General Medicine, Down Syndrome, Child, Megakaryocytes, Hematopoiesis
Abstract

Acute megakaryoblastic leukemia of Down syndrome (DS-AMKL) is a model of clonal evolution from a preleukemic transient myeloproliferative disorder requiring both a trisomy 21 (T21) and a GATA1s mutation to a leukemia driven by additional driver mutations. We modeled the megakaryocyte differentiation defect through stepwise gene editing of GATA1s, SMC3+/-, and MPLW515K, providing 20 different T21 or disomy 21 (D21) induced pluripotent stem cell (iPSC) clones. GATA1s profoundly reshaped iPSC-derived hematopoietic architecture with gradual myeloid-to-megakaryocyte shift and megakaryocyte differentiation alteration upon addition of SMC3 and MPL mutations. Transcriptional, chromatin accessibility, and GATA1-binding data showed alteration of essential megakaryocyte differentiation genes, including NFE2 downregulation that was associated with loss of GATA1s binding and functionally involved in megakaryocyte differentiation blockage. T21 enhanced the proliferative phenotype, reproducing the cellular and molecular abnormalities of DS-AMKL. Our study provides an array of human cell-based models revealing individual contributions of different mutations to DS-AMKL differentiation blockage, a major determinant of leukemic progression.

Published
2022

8. High caspase 3 and vulnerability to dual BCL2 family inhibition define ETO2::GLIS2 pediatric leukemia

Author

Zakia Aid, Elie Robert, Cécile K. Lopez, Maxence Bourgoin, Fabien Boudia, Melchior Le Mene, Julie Riviere, Marie Baille, Salima Benbarche, Laurent Renou, Alexandre Fagnan, Cécile Thirant, Laetitia Federici, Laure Touchard, Yann Lecluse, Anton Jetten, Birgit Geoerger, Hélène Lapillonne, Eric Solary, Muriel Gaudry, Soheil Meshinchi, Françoise Pflumio, Patrick Auberger, Camille Lobry, Arnaud Petit, Arnaud Jacquel, and Thomas Mercher

Subjects
Cancer Research, Oncology, Hematology
Abstract

Pediatric acute myeloid leukemia expressing the ETO2::GLIS2 fusion oncogene is associated with dismal prognosis. Previous studies have shown that ETO2::GLIS2 can efficiently induce leukemia development associated with strong transcriptional changes but those amenable to pharmacological targeting remained to be identified. By studying an inducible ETO2::GLIS2 cellular model, we uncovered that de novo ETO2::GLIS2 expression in human cells led to increased CASP3 transcription, CASP3 activation, and cell death. Patient-derived ETO2::GLIS2

Published
2022

9. The pediatric acute leukemia fusion oncogene ETO2-GLIS2 increases self-renewal and alters differentiation in a human induced pluripotent stem cells-derived model

Author

Franco Locatelli, Salvatore Serravalle, Cécile Thirant, Andrea Pession, Riccardo Masetti, Annalisa Astolfi, William Vainchenker, Valentina Indio, Alessandro Donada, Zakia Aid, Thomas Mercher, Salvatore Nicola Bertuccio, Elie Robert, Hana Raslova, Fabien Boudia, Larissa Lordier, Cécile K. Lopez, Marie Cambot, Bertuccio, SN, Boudia, F, Cambot, M, Lopez, CK, Lordier, L, Donada, A, Robert, E, Thirant, C, Aid, Z, Serravalle, S, Astolfi, A, Indio, V, Locatelli, F, Pession, A, Vainchenker, W, Masetti, R, Raslova, H, and Mercher, T

Subjects
Letter, Biology, Self renewal, NO, 03 medical and health sciences, 0302 clinical medicine, Text mining, GLIS2, Human Induced Pluripotent Stem Cells, 030304 developmental biology, 0303 health sciences, Acute leukemia, Oncogene, lcsh:RC633-647.5, business.industry, lcsh:Diseases of the blood and blood-forming organs, Hematology, 3. Good health, Settore MED/38 - PEDIATRIA GENERALE E SPECIALISTICA, 030220 oncology & carcinogenesis, ComputingMethodologies_DOCUMENTANDTEXTPROCESSING, ETO2-GLIS2, Cancer research, na, business
Abstract

Supplemental Digital Content is available in the text.

Published
2020

10. Ontogenic changes in hematopoietic hierarchy determine pediatric specificity and disease phenotype in fusion oncogene-driven myeloid leukemia

Author

Cécile K. Lopez, Marie Laure Arcangeli, Eric Delabesse, Sébastien Malinge, Alexandre Fagnan, Isabelle Godin, Franco Locatelli, Françoise Pflumio, Fabien Boudia, Cécile Thirant, Muriel Gaudry, Vaia Stavropoulou, Arnaud Petit, Claus Nerlov, Nathalie Droin, Riccardo Masetti, Paola Ballerini, Zakia Aid, Berthold Göttgens, Olivier Bernard, Sarah Kinston, Erika Brunet, Hélène Lapillonne, Loelia Babin, Juerg Schwaller, Antoine H.F.M. Peters, Elie Robert, Yann Lécluse, Bastien Job, Chrystele Bilhou-Nabera, Jean-Luc Villeval, Camille Lobry, William Vainchenker, Thomas Mercher, Esteve Noguera, M'Boyba Diop, Service d'hématologie-immunologie-oncologie pédiatrique [CHU Trousseau], Université Pierre et Marie Curie - Paris 6 (UPMC)-Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-CHU Trousseau [APHP], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP), Centre de Recherche Saint-Antoine (UMRS893), Université Pierre et Marie Curie - Paris 6 (UPMC)-Institut National de la Santé et de la Recherche Médicale (INSERM), CHU Trousseau [APHP], Institut de psychiatrie et neurosciences (U894 / UMS 1266), Université Paris Descartes - Paris 5 (UPD5)-Institut National de la Santé et de la Recherche Médicale (INSERM), Stabilité génétique, Cellules Souches et Radiations (SCSR (U_967)), Université Paris-Sud - Paris 11 (UP11)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Paris Cité (UPC), Hématopoïèse normale et pathologique (U1170 Inserm), Université Paris-Sud - Paris 11 (UP11)-Institut Gustave Roussy (IGR)-Institut National de la Santé et de la Recherche Médicale (INSERM), Plateforme de Bioinformatique [Gustave Roussy], Analyse moléculaire, modélisation et imagerie de la maladie cancéreuse (AMMICa), Université Paris-Sud - Paris 11 (UP11)-Institut Gustave Roussy (IGR)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-Université Paris-Sud - Paris 11 (UP11)-Institut Gustave Roussy (IGR)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), Bases fondamentales et stratégies nouvelles en cancérologie (BFSNC - IFR54), Université Paris-Sud - Paris 11 (UP11)-Institut Gustave Roussy (IGR)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), Régulation et dynamique des génomes, Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), Centre de Recherches en Cancérologie de Toulouse (CRCT), Université Toulouse III - Paul Sabatier (UT3), Université Fédérale Toulouse Midi-Pyrénées-Université Fédérale Toulouse Midi-Pyrénées-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), Santa Lucia Foundation, IRCSS, Rome, University of Oxford [Oxford], Images et Modèles, Centre de Recherche en Acquisition et Traitement de l'Image pour la Santé (CREATIS), Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Institut National des Sciences Appliquées de Lyon (INSA Lyon), Université de Lyon-Institut National des Sciences Appliquées (INSA)-Institut National des Sciences Appliquées (INSA)-Hospices Civils de Lyon (HCL)-Université Jean Monnet [Saint-Étienne] (UJM)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Institut National des Sciences Appliquées (INSA)-Institut National des Sciences Appliquées (INSA)-Hospices Civils de Lyon (HCL)-Université Jean Monnet [Saint-Étienne] (UJM)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), University of Cambridge [UK] (CAM), Service Procédés et Innovations Industriels (SPII), eRcane, Institut Cochin (UMR_S567 / UMR 8104), Université Paris Descartes - Paris 5 (UPD5)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), Génétique des tumeurs (U985), Institut Gustave Roussy (IGR)-Université Paris Descartes - Paris 5 (UPD5)-Institut National de la Santé et de la Recherche Médicale (INSERM), Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Sorbonne Université (SU)-Sorbonne Université (SU), Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Sorbonne Université (SU), Université Paris-Sud - Paris 11 (UP11)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université de Paris (UP), Centre National de la Recherche Scientifique (CNRS)-Université Paris-Sud - Paris 11 (UP11)-Institut Gustave Roussy (IGR)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-Université Paris-Sud - Paris 11 (UP11)-Institut Gustave Roussy (IGR)-Institut National de la Santé et de la Recherche Médicale (INSERM), Centre de Recherche et d'Etude en Droit et Science Politique (CREDESPO), Université de Bourgogne (UB), Université Jean Monnet [Saint-Étienne] (UJM)-Hospices Civils de Lyon (HCL)-Institut National des Sciences Appliquées de Lyon (INSA Lyon), Université de Lyon-Institut National des Sciences Appliquées (INSA)-Université de Lyon-Institut National des Sciences Appliquées (INSA)-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Centre National de la Recherche Scientifique (CNRS)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Jean Monnet [Saint-Étienne] (UJM)-Hospices Civils de Lyon (HCL)-Institut National des Sciences Appliquées de Lyon (INSA Lyon), Université de Lyon-Centre National de la Recherche Scientifique (CNRS)-Institut National de la Santé et de la Recherche Médicale (INSERM), Lopez C.K., Noguera E., Stavropoulou V., Robert E., Aid Z., Ballerini P., Bilhou-Nabera C., Lapillonne H., Boudia F., Thirant C., Fagnan A., Arcangeli M.-L., Kinston S.J., Diop M., Job B., Lecluse Y., Brunet E., Babin L., Villeval J.L., Delabesse E., Peters A.H.F.M., Vainchenker W., Gaudry M., Masetti R., Locatelli F., Malinge S., Nerlov C., Droin N., Lobry C., Godin I., Bernard O.A., Gottgens B., Petit A., Pflumio F., Schwaller J., Mercher T., Université Pierre et Marie Curie - Paris 6 (UPMC)-Assistance publique - Hôpitaux de Paris (AP-HP) (APHP)-CHU Trousseau [APHP], Centre de Recherche Saint-Antoine (CR Saint-Antoine), Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Pierre et Marie Curie - Paris 6 (UPMC), Laboratoire d'Hématologie [AP-HP Hôpital Armand Trousseau], Cellules Souches et Radiations (SCSR (U967 / UMR-E_008)), Université Paris-Sud - Paris 11 (UP11)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Paris Diderot - Paris 7 (UPD7)-Institut National de la Santé et de la Recherche Médicale (INSERM), Institut National de la Santé et de la Recherche Médicale (INSERM)-Institut Gustave Roussy (IGR)-Université Paris-Sud - Paris 11 (UP11), Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-Université Paris Descartes - Paris 5 (UPD5), Brunet, Erika [0000-0002-1726-4673], Malinge, Sébastien [0000-0002-9533-7778], Droin, Nathalie [0000-0002-6099-5324], Godin, Isabelle [0000-0001-8577-8388], Göttgens, Berthold [0000-0001-6302-5705], Schwaller, Juerg [0000-0001-8616-0096], Mercher, Thomas [0000-0003-1552-087X], Apollo - University of Cambridge Repository, Centre de Psychiatrie et Neurosciences (U894), Université Paris-Sud - Paris 11 (UP11)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Paris Cité (UPCité), Université de Toulouse (UT)-Université de Toulouse (UT)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), University of Oxford, Université de Lyon-Institut National des Sciences Appliquées (INSA)-Institut National des Sciences Appliquées (INSA)-Université Jean Monnet - Saint-Étienne (UJM)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-Université Claude Bernard Lyon 1 (UCBL), and Université de Lyon-Institut National des Sciences Appliquées (INSA)-Institut National des Sciences Appliquées (INSA)-Université Jean Monnet - Saint-Étienne (UJM)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)

Subjects
0301 basic medicine, Myeloid, Oncogene Proteins, Fusion, Oncogene Proteins, Fusion gene, Mice, 0302 clinical medicine, AML, CEBPA, GENOMIC ALTERATIONS, Tumor Cells, Cultured, TRANSCRIPTION FACTOR, RNA-SEQ, Child, ComputingMilieux_MISCELLANEOUS, GENE-EXPRESSION, Age Factors, Myeloid leukemia, GATA1, 3. Good health, Leukemia, Myeloid, Acute, Leukemia, Haematopoiesis, medicine.anatomical_structure, DIFFERENTIATION, Oncology, Settore MED/38 - PEDIATRIA GENERALE E SPECIALISTICA, Child, Preschool, 030220 oncology & carcinogenesis, Female, Adolescent, [SDV.CAN]Life Sciences [q-bio]/Cancer, Biology, 03 medical and health sciences, children, ACUTE MEGAKARYOBLASTIC LEUKEMIA, transcription factors, medicine, Animals, Humans, FETAL, pediatric acute myeloid leukemia, LINEAGE COMMITMENT, Infant, medicine.disease, STEM-CELL, SELF-RENEWAL, 030104 developmental biology, Cancer research, Neoplasm Transplantation
Abstract

Fusion oncogenes are prevalent in several pediatric cancers, yet little is known about the specific associations between age and phenotype. We observed that fusion oncogenes, such as ETO2–GLIS2, are associated with acute megakaryoblastic or other myeloid leukemia subtypes in an age-dependent manner. Analysis of a novel inducible transgenic mouse model showed that ETO2–GLIS2 expression in fetal hematopoietic stem cells induced rapid megakaryoblastic leukemia whereas expression in adult bone marrow hematopoietic stem cells resulted in a shift toward myeloid transformation with a strikingly delayed in vivo leukemogenic potential. Chromatin accessibility and single-cell transcriptome analyses indicate ontogeny-dependent intrinsic and ETO2–GLIS2-induced differences in the activities of key transcription factors, including ERG, SPI1, GATA1, and CEBPA. Importantly, switching off the fusion oncogene restored terminal differentiation of the leukemic blasts. Together, these data show that aggressiveness and phenotypes in pediatric acute myeloid leukemia result from an ontogeny-related differential susceptibility to transformation by fusion oncogenes. Significance: This work demonstrates that the clinical phenotype of pediatric acute myeloid leukemia is determined by ontogeny-dependent susceptibility for transformation by oncogenic fusion genes. The phenotype is maintained by potentially reversible alteration of key transcription factors, indicating that targeting of the fusions may overcome the differentiation blockage and revert the leukemic state. See related commentary by Cruz Hernandez and Vyas, p. 1653. This article is highlighted in the In This Issue feature, p. 1631

Published
2019

11. Modeling Acute Megakaryoblastic Leukemia of Down Syndrome Using Induced Pluripotent Stem Cells

Author

Isabelle Plo, Romain Diot, Marie Cambot, G Tachdjian, Eric Soler, Virginie Dufour, Najet Debili, Zakia Aid, Hana Raslova, Cyril Catelain, Rachel Petermann, Stefania Mazzi, Sébastien Malinge, Yasmine Mammasse, Thomas Mercher, Mathieu Vieira, Elie Robert, Sylvie Souquere, Aurelie Siret, Philippe Rameau, William Vainchenker, Fabien Boudia, and Brahim Arkoun

Subjects
Acute megakaryoblastic leukemia, Down syndrome, business.industry, Immunology, Cancer research, Medicine, Cell Biology, Hematology, business, medicine.disease, Induced pluripotent stem cell, Biochemistry
Abstract

Introduction Development of Acute megakaryoblastic leukemia in Down syndrome children (DS-AMKL) is a multi-step process. Acquired GATA1s mutation during fetal hematopoiesis is responsible of a transient myeloproliferative disorder (TMD) characterized by an accumulation of megakaryoblasts. Although most of TMD regress around birth, some TMD can progress from the initial GATA1s clone to AMKL through the acquisition of additional mutations, including in (i) the cohesin complex (i.e: SMC3), (ii) the JAK/STAT signaling pathway, such as MPL and (iii) the polycomb repressive complex 2 (EZH2). How these mutations cooperate to deregulate megakaryocyte (MK) differentiation and to induce a full-blown AMKL, along with the precise role of trisomy 21 (T21) during this transformation process remain unclear. Because modeling of DS-AMKL is particularly difficult in mice, we performed a step-wise introduction of GATA1s, a gain of function mutation of MPL (MPLW515K) and a heterozygous loss of function mutation in a cohesin (SMC3), separately or in combination, in T21 and isogenic disomic 21 (Dis21) human induced Pluripotent Stem Cells (iPSCs). Methods Trisomy 21 iPSCs were kindly provided by M. Weiss (Memphis, TN). CRISPR/Cas 9 genome editing of GATA1 or SMC3 allowed the generation of GATA1s T21, SMC3+/- T21 and GATA1s SMC3+/- T21 iPSC clones. CRISPR/Cas9-mediated knock-in of MPLW515K was performed in T21 GATA1s iPSCs. The subsequent T21 GATA1sMPLW515K/W515Kclones were selected as well as a revertant Dis21 GATA1sMPLW515K/W515Kclone. Finally, SMC3 insertion/deletion were obtained in isogenic T21 and Dis21 GATA1s MPLW515K/W515K SMC3+/-iPSCs clones. Hematopoietic differentiation was induced in 2D cultures in presence of a matrix and a cocktail of cytokines followed by a MK differentiation with SCF and TPO. MK differentiation was studied by clonogenic assays, flow cytometry, confocal microscopy and ultrastructural studies. Gene expression analyses were performed by RNA-seq on highly purified MK from all genotypes. Results GATA1s alone blocked MK maturation characterized by a persistent CD34 expression, an accumulation of abnormal large granules, a defect in the development of demarcation membranes (DMS), and a marked decrease in proplatelet formation. The typical GATA1s MK were large megakaryoblasts with numerous large granules and rare DMS. However, GATA1s alone had no effect on the clonogenic activity in CFU-MK assays and MK numbers. The introduction of the MPLW515K mutation did not modify this phenotype either in Dis21 or T21 GATA1s MK, but induced a complete TPO independence. SMC3+/- alone enhanced the MK maturation allowing the generation of a higher number of proplatelets-generating MK. Importantly, the combination of GATA1s and SMC3+/- mutations had a marked cooperative effect that worsened the MK maturation defect, led to the generation of abnormal megakaryoblasts with only a pre-DMS and resulted in enhanced proliferation and ploidization both in Dis21 and T21 iPSCs. Interestingly, the proliferation was markedly higher in T21 clones compared to Dis21 counterparts. RNA-seq and GSEA analyses showed that T21 GATA1s SMC3+/- mutant MK exhibited transcriptional signatures consistent with a dramatic decrease in the expression of maturation genes, including GATA1 target genes, while DNA replication gene markers were increased compared with GATA1s alone. T21 GATA1s MPLW515K/W515K SMC3+/- MK were enriched for AMKL signatures as compared to isogenic Dis21 GATA1sMPLW515K/W515K SMC3+/- MK. Ongoing ATAC-seq analyses will define the consequence of the different mutations on chromatin accessibility. Conclusion Using iPSC modeling, we analyzed in a human cell-context the consequences of the different combination of mutations associated with DS-AMKL that would be difficult to model using human primary cells. Our data demonstrate that GATA1s expression cooperates with SMC3+/- to enhance proliferation of megakaryoblasts from T21 iPSCs and isogenic Dis21 iPSCs hence reproducing the abnormalities observed in DS-AMKL. T21 is not directly involved in the MK differentiation defects but rather give a proliferative advantage supporting its role in leukemia development. Disclosures No relevant conflicts of interest to declare.

Published
2020

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