117 results on '"Luc Douay"'
Search Results
2. Comprehensive Proteomic Analysis of Human Erythropoiesis
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Patrick Mayeux, Narla Mohandas, Anna Raimbault, Michael Dussiot, Virginie Salnot, John Hale, Yael Zermati, Sarah Ducamp, Marie-Catherine Giarratana, Luc Douay, François Guillonneau, Marjorie Leduc, Frédérique Verdier, Emilie-Fleur Gautier, Catherine Lacombe, Institut Cochin (IC UM3 (UMR 8104 / U1016)), Université Paris Descartes - Paris 5 (UPD5)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), Plateforme protéomique 3P5 [Institut Cochin] (3P5), Université Paris Descartes - Paris 5 (UPD5)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-Université Paris Descartes - Paris 5 (UPD5)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), Red Cell Physiology Laboratory [New York, USA], New York Blood Center, 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), Ligue Nationale Contre le Cancer - Paris, Ligue Nationnale Contre le Cancer, ANR-11-IDEX-0005,USPC,Université Sorbonne Paris Cité(2011), Institut Cochin ( UM3 (UMR 8104 / U1016) ), Université Paris Descartes - Paris 5 ( UPD5 ) -Institut National de la Santé et de la Recherche Médicale ( INSERM ) -Centre National de la Recherche Scientifique ( CNRS ), Laboratory of Excellence GR-Ex, Sorbonne Paris Cité-Université Paris Descartes - Paris 5 ( UPD5 ) -Imagine Institute, Plateforme protéomique 3P5 [Institut Cochin] ( 3P5 ), Université Paris Descartes - Paris 5 ( UPD5 ) -Institut National de la Santé et de la Recherche Médicale ( INSERM ) -Centre National de la Recherche Scientifique ( CNRS ) -Université Paris Descartes - Paris 5 ( UPD5 ) -Institut National de la Santé et de la Recherche Médicale ( INSERM ) -Centre National de la Recherche Scientifique ( CNRS ), New York Blood Center - NYBC, Centre de Recherche Saint-Antoine ( CR Saint-Antoine ), Université Pierre et Marie Curie - Paris 6 ( UPMC ) -Institut National de la Santé et de la Recherche Médicale ( INSERM ), ANR-11-IDEX-0005-02/11-LABX-0051,GR-Ex,Biogenèse et pathologies du globule rouge ( 2011 ), ANR-11-IDEX-0005-02/11-IDEX-0005,USPC,USPC ( 2011 ), Bos, Mireille, Université Sorbonne Paris Cité - - USPC2011 - ANR-11-IDEX-0005 - IDEX - VALID, and Ligue Nationale Contre le Cancer (LNCC)
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Proteomics ,0301 basic medicine ,Cell type ,Erythroblasts ,Proteome ,Cellular differentiation ,Biology ,Article ,General Biochemistry, Genetics and Molecular Biology ,03 medical and health sciences ,Reticulocyte ,hemic and lymphatic diseases ,medicine ,[SDV.BBM] Life Sciences [q-bio]/Biochemistry, Molecular Biology ,Humans ,Erythropoiesis ,[SDV.BBM]Life Sciences [q-bio]/Biochemistry, Molecular Biology ,RNA, Messenger ,lcsh:QH301-705.5 ,[ SDV.BBM ] Life Sciences [q-bio]/Biochemistry, Molecular Biology ,Cells, Cultured ,Erythroid Precursor Cells ,Messenger RNA ,RNA ,Cell Differentiation ,Molecular biology ,Cell biology ,030104 developmental biology ,medicine.anatomical_structure ,lcsh:Biology (General) - Abstract
SUMMARY Mass spectrometry-based proteomics now enables the absolute quantification of thousands of proteins in individual cell types. We used this technology to analyze the dynamic proteome changes occurring during human erythropoiesis. We quantified the absolute expression of 6,130 proteins during erythroid differentiation from late burst-forming units-erythroid (BFU-Es) to orthochromatic erythroblasts. A modest correlation between mRNA and protein expression was observed. We identified several proteins with unexpected expression patterns in erythroid cells, highlighting a breakpoint in the erythroid differentiation process at the basophilic stage. We also quantified the distribution of proteins between reticulocytes and pyrenocytes after enucleation. These analyses identified proteins that are actively sorted either with the reticulocyte or the pyrenocyte. Our study provides the absolute quantification of protein expression during a complex cellular differentiation process in humans, and it establishes a framework for future studies of disordered erythropoiesis., In Brief Gautier et al. use quantitative mass spectrometry to determine the absolute proteome composition of human erythroid progenitors throughout the differentiation process and the quantitative distribution of proteins between reticulocytes and pyrenocytes after enucleation.
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- 2016
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3. Clonal history of a cord blood donor cell leukemia with prenatal somatic JAK2 V617F mutation
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Dominique Bories, A.-C. Mamez, Ramdane Belhocine, François Delhommeau, O. Legrand, Ludovic Suner, Fanny Fava, Simona Lapusan, Pierre Hirsch, Ruoping Tang, Christophe Marzac, M. Mohty, and Luc Douay
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Male ,0301 basic medicine ,Cancer Research ,medicine.medical_specialty ,Somatic cell ,Cord Blood Stem Cell Transplantation ,Biology ,medicine.disease_cause ,03 medical and health sciences ,0302 clinical medicine ,hemic and lymphatic diseases ,Internal medicine ,medicine ,Humans ,Mutation ,Hematology ,Neoplasms, Second Primary ,Janus Kinase 2 ,Middle Aged ,Allografts ,medicine.disease ,Tissue Donors ,Leukemia, Myeloid, Acute ,Haematopoiesis ,Leukemia ,030104 developmental biology ,Oncology ,030220 oncology & carcinogenesis ,Cord blood ,Immunology ,Stem cell - Abstract
Clonal history of a cord blood donor cell leukemia with prenatal somatic JAK2 V617F mutation
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- 2016
4. Génération de globules rouges de culture à partir de cellules souches : bref récit du futur
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Luc Douay and Christelle Mazurier
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0301 basic medicine ,Blood transfusion ,business.industry ,medicine.medical_treatment ,Biology ,Phenotype ,Embryonic stem cell ,Regenerative medicine ,General Biochemistry, Genetics and Molecular Biology ,Cell biology ,03 medical and health sciences ,030104 developmental biology ,0302 clinical medicine ,030220 oncology & carcinogenesis ,medicine ,Personalized medicine ,Stem cell ,Induced pluripotent stem cell ,business ,Adult stem cell - Abstract
Human adult pluripotent stem cells, stem cells of embryonic origin and induced pluripotent stem cells (iPS) provide cellular sources for new promising regenerative medicine approaches. Because these cells can be patient-specific, they allow considering a personalized medicine appropriate to the diagnosis of each. The generation of cultured red blood cells (cRBC) derived from stem cells is emblematic of personalized medicine. Indeed, these cells have the advantage of being selected according to a blood phenotype of interest and they may provide treatments to patients in situation of impossible transfusion (alloimmunized patients, rare phenotypes). Essential progresses have established proof of concept for this approach, still a concept some years ago. From adult stem cells, all steps of upstream research were successfully achieved, including the demonstration of the feasibility of injection into human. This leads us to believe that Red Blood Cells generated in vitro from stem cells will be the future players of blood transfusion. However, although theoretically ideal, these stem cells raise many biological challenges to overcome, although some tracks are identified.
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- 2016
5. Why industrial production of red blood cells from stem cells is essential for tomorrow's blood transfusion
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Luc Douay
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Embryology ,Blood transfusion ,Erythrocytes ,business.industry ,medicine.medical_treatment ,Blood Safety ,Stem Cells ,Biomedical Engineering ,Cell Culture Techniques ,Blood Donors ,030204 cardiovascular system & hematology ,Regenerative Medicine ,Andrology ,03 medical and health sciences ,0302 clinical medicine ,Medicine ,Humans ,Blood supply ,Blood Transfusion ,Stem cell ,business ,030215 immunology ,Adult stem cell - Published
- 2018
6. Mesenchymal Stem Cell Administration Attenuates Colon Cancer Progression by Modulating the Immune Component within the Colorectal Tumor Microenvironment
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Sabine, François, Benoit, Usunier, Marie-Elisabeth, Forgue-Lafitte, Bruno, L'Homme, Marc, Benderitter, Luc, Douay, Norbert-Claude, Gorin, Annette K, Larsen, and Alain, Chapel
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Polarization of immune cells ,Radiotherapy ,Macrophages ,Endothelial Cells ,Cell Differentiation ,Mesenchymal Stem Cells ,CD8-Positive T-Lymphocytes ,Mesenchymal Stem Cell Transplantation ,Colorectal cancer ,T-Lymphocytes, Regulatory ,Coculture Techniques ,Cell therapy ,Rats ,Rats, Sprague-Dawley ,Disease Models, Animal ,Pelvic radiation disease ,Tissue Engineering and Regenerative Medicine ,Disease Progression ,Tumor Microenvironment ,Cytokines ,Th17 Cells ,Animals ,Humans ,Colorectal Neoplasms - Abstract
We here determine the influence of mesenchymal stem cell (MSC) therapy on the progression of solid tumors. The influence of MSCs was investigated in human colorectal cancer cells as well as in an immunocompetent rat model of colorectal carcinogenesis representative of the human pathology. Treatment with bone marrow (BM)‐derived MSCs significantly reduced both cancer initiation and cancer progression by increasing the number of tumor‐free animals as well as decreasing the number and the size of the tumors by half, thereby extending their lifespan. The attenuation of cancer progression was mediated by the capacity of the MSCs to modulate the immune component. Specifically, in the adenocarcinomas (ADKs) of MSC‐treated rats, the infiltration of CD68+ monocytes/macrophages was 50% less while the presence of CD3+ lymphocytes increased almost twofold. The MSCs reprogrammed the macrophages to become regulatory cells involved in phagocytosis thereby inhibiting the production of proinflammatory cytokines. Furthermore, the MSCs decreased NK (Natural Killer) and rTh17 cell activities, Treg recruitment, the presence of CD8+ lymphocytes and endothelial cells while restoring Th17 cell activity. The expression of miR‐150 and miR‐7 increased up to fivefold indicating a likely role for these miRNAs in the modulation of tumor growth. Importantly, MSC administration limited the damage of healthy tissues and attenuated tumor growth following radiotherapy. Taken together, we here show that that MSCs have durable action on colon cancer development by modulating the immune component of the tumor microenvironment. In addition, we identify two miRNAs associated with the capacity of MSCs to attenuate cancer growth. stem cells translational medicine 2019;8:285&300
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- 2018
7. Production de globules rouges in vitro
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Luc Douay
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Biochemistry (medical) ,Clinical Biochemistry ,Hematology - Abstract
L’exigence de disponibilite des PSL adaptes aux besoins de tous est un defi de sante publique. Si nous n’avons jamais manque de sang dans nos pays developpes, nous en manquerons dans les prochaines decennies en raison du vieillissement de la population. La transfusion sanguine restant le seul traitement symptomatique pour de nombreuses situations, il est temps d’inventer de nouvelles sources complementaires de PSL. Les cellules souches (CS) sont les outils des nouvelles approches prometteuses de la medecine regeneratrice. La production in vitro de globules rouges de culture (GRc) est maintenant possible a partir de trois types principaux de CS : les CS hematopoietiques, les CS pluripotentes induites (iPS) et les lignees erythroides immortalisees. La generation in vitro de GRc pourrait repondre, au cours des prochaines annees, aux defis actuels et futurs de la transfusion sanguine. Les vertus attendues des GRc sont en effet des atouts potentiels : une duree de vie superieure a celle d’une population native de GR, une compatibilite antigenique quasi universelle, une securite infectieuse accrue, une disponibilite permanente. Le moment est venu de tenter de developper leur production industrielle de masse. Si la comprehension des processus biologiques fondamentaux de l’hematopoiese nous a permis d’injecter chez l’homme quelques millilitres de GRc, passer aux essais cliniques grandeur nature avec l’equivalents de plusieurs CGR est sans conteste un defi considerable. Il nous faut en effet aujourd’hui relever plusieurs obstacles biotechnologiques et economiques majeurs. Le point sera fait sur les exigences de rupture pour la conception d’un demonstrateur industriel.
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- 2019
8. Design and Validation of an Automated Process for the Expansion of Peripheral Blood-Derived CD34
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Claire, Saucourt, Sandrine, Vogt, Amandine, Merlin, Christophe, Valat, Anthony, Criquet, Laurence, Harmand, Brigitte, Birebent, Hélène, Rouard, Christian, Himmelspach, Éric, Jeandidier, Anne-Gaële, Chartois-Leauté, Sophie, Derenne, Laurence, Koehl, Joe-Elie, Salem, Jean-Sébastien, Hulot, Céline, Tancredi, Anne, Aries, Sébastien, Judé, Eric, Martel, Serge, Richard, Luc, Douay, and Philippe, Hénon
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Adult ,Automation, Laboratory ,Male ,Clinical Trials as Topic ,Peripheral Blood Stem Cell Transplantation ,Peripheral blood stem cells ,Primary Cell Culture ,Cellular therapy ,Myocardial Infarction ,Antigens, CD34 ,Middle Aged ,Flow Cytometry ,Immunophenotyping ,Rats ,Manufacturing for Regenerative Medicine ,Animals ,Humans ,Cell culture ,Cardiac ,Cells, Cultured ,CD34+ ,Hematopoietic stem cells - Abstract
We previously demonstrated that intracardiac delivery of autologous peripheral blood‐derived CD34+ stem cells (SCs), mobilized by granulocyte‐colony stimulating factor (G‐CSF) and collected by leukapheresis after myocardial infarction, structurally and functionally repaired the damaged myocardial area. When used for cardiac indication, CD34+ cells are now considered as Advanced Therapy Medicinal Products (ATMPs). We have industrialized their production by developing an automated device for ex vivo CD34+‐SC expansion, starting from a whole blood (WB) sample. Blood samples were collected from healthy donors after G‐CSF mobilization. Manufacturing procedures included: (a) isolation of total nuclear cells, (b) CD34+ immunoselection, (c) expansion and cell culture recovery in the device, and (d) expanded CD34+ cell immunoselection and formulation. The assessment of CD34+ cell counts, viability, and immunophenotype and sterility tests were performed as quality tests. We established graft acceptance criteria and performed validation processes in three cell therapy centers. 59.4 × 106 ± 36.8 × 106 viable CD34+ cells were reproducibly generated as the final product from 220 ml WB containing 17.1 × 106 ± 8.1 × 106 viable CD34+ cells. CD34+ identity, genetic stability, and telomere length were consistent with those of basal CD34+ cells. Gram staining and mycoplasma and endotoxin analyses were negative in all cases. We confirmed the therapeutic efficacy of both CD34+‐cell categories in experimental acute myocardial infarct (AMI) in immunodeficient rats during preclinical studies. This reproducible, automated, and standardized expansion process produces high numbers of CD34+ cells corresponding to the approved ATMP and paves the way for a phase I/IIb study in AMI, which is currently recruiting patients. stem cells translational medicine 2019;8:822&832
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- 2017
9. Transgene-free hematopoietic stem and progenitor cells from human induced pluripotent stem cells
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Nathalie Chevallier, Loïc Garçon, Laurence Guyonneau-Harmand, Hélène Lapillonne, Luc Douay, Marc Benderitter, Brigitte Birebent, François Delhommeau, Christophe Desterke, Thierry Jaffredo, Bruno Homme, Alain Chapel, Université Pierre et Marie Curie - Paris 6 (UPMC), Laboratoire d'Hématologie et d'Immunologie [CHU Saint-Antoine], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-CHU Saint-Antoine [AP-HP], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Sorbonne Université (SU)-Sorbonne Université (SU), EFS Ile de France, Unité d’Ingénierie et de Thérapie Cellulaire, Créteil, F-94017, France., PRP-HOM/SRBE/LRTE, Institut de Radioprotection et de Sûreté Nucléaire (IRSN), Unité de service de l'Institut André Lwoff (US 33 Inserm), Hôpital Paul Brousse-Institut André Lwoff [Villejuif] (IAL), Biomécanique cellulaire et respiratoire (BCR), Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Paris-Est Créteil Val-de-Marne - Paris 12 (UPEC UP12)-Centre National de la Recherche Scientifique (CNRS), CHU Trousseau [APHP], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Sorbonne Université (SU), 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 Saint-Antoine [AP-HP], Migration et différenciation des cellules souches hématopoiétiques = Migration and differentiation of hematopoietic stem cells (LBD-E06), Laboratoire de Biologie du Développement (LBD), Université Pierre et Marie Curie - Paris 6 (UPMC)-Institut de Biologie Paris Seine (IBPS), Université Pierre et Marie Curie - Paris 6 (UPMC)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-Centre National de la Recherche Scientifique (CNRS)-Université Pierre et Marie Curie - Paris 6 (UPMC)-Institut de Biologie Paris Seine (IBPS), Université Pierre et Marie Curie - Paris 6 (UPMC)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-Centre National de la Recherche Scientifique (CNRS)-Institut National de la Santé et de la Recherche Médicale (INSERM), Service d'hématologie clinique et de thérapie cellulaire [CHU Saint-Antoine], Direction Générale de l’Armement ASTRID/ANR program, Etablissement Français du Sang APR 2013, association 'Combattre La Leucémie', joint grant from Agence Nationale pour la Recherche/California Institute for Regenerative Medicine (ANR/CIRM 0001-02), Laboratoire de Radiopathologie et de Thérapies Expérimentales (IRSN/PRP-HOM/SRBE/LRTE), Unité mixte de service UMS33 [Institut André Lwoff] (UMS33 Inserm/IAL), Hôpital Paul Brousse-Institut National de la Santé et de la Recherche Médicale (INSERM)-Institut André Lwoff [Villejuif] (IAL), and Jaffredo, Thierry
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[SDV.MHEP.HEM] Life Sciences [q-bio]/Human health and pathology/Hematology ,Myeloid ,Population ,[SDV.BC.BC]Life Sciences [q-bio]/Cellular Biology/Subcellular Processes [q-bio.SC] ,Biology ,CXCR4 ,03 medical and health sciences ,0302 clinical medicine ,Directed differentiation ,[SDV.BC.BC] Life Sciences [q-bio]/Cellular Biology/Subcellular Processes [q-bio.SC] ,medicine ,Progenitor cell ,education ,hemogenic endothelium ,030304 developmental biology ,Hemogenic endothelium ,0303 health sciences ,education.field_of_study ,[SDV.BDD.EO] Life Sciences [q-bio]/Development Biology/Embryology and Organogenesis ,[SDV.MHEP.HEM]Life Sciences [q-bio]/Human health and pathology/Hematology ,[SDV.BBM.MN]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Molecular Networks [q-bio.MN] ,Cell biology ,hematopoietic stem cells ,human induced pluripotent stem cells ,Haematopoiesis ,hematopoietic reconstitution ,medicine.anatomical_structure ,[SDV.BDD.EO]Life Sciences [q-bio]/Development Biology/Embryology and Organogenesis ,[SDV.BBM.MN] Life Sciences [q-bio]/Biochemistry, Molecular Biology/Molecular Networks [q-bio.MN] ,030220 oncology & carcinogenesis ,Immunology ,Homing (hematopoietic) - Abstract
Introductory paragraphThe successful production of Hematopoietic Stem and Progenitor Cells (HSPCs) from human pluripotent sources is conditioned by transgene delivery1-5. We describe here a dedicated and tractable one step, GMP-grade, transgene-free and stroma-free protocol to produce HSPCs from human induced pluripotent stem cells (hiPSCs). This procedure, applied to several sources of hiPSCs with equal efficiency, is based on a directed differentiation with morphogens and cytokines to generate a cell population close to nascent HSPCs or their immediate forerunners i.e., hemogenic endothelial cells6-9. Following engraftment into immunocompromised recipients, this cell population was proved capable of a robust myeloid, lymphoid and definitive red blood cell production in sequential recipients for at least 40 weeks. Further identification of the repopulating cells show that they express the G protein–coupled receptor APELIN (APLNR) and the homing receptor CXCR4. This demonstrates that the generation of bona fide HSPCs from hiPSCs without transgenes is possible and passes through an early endo-hematopoietic intermediate. This work opens the way to the generation of clinical grade HSPCs for the treatment of hematological diseases and holds promise for the analysis of HSPC development in the human species.
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- 2017
10. Precision and prognostic value of clone-specific minimal residual disease in acute myeloid leukemia
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Pierre, Hirsch, Ruoping, Tang, Nassera, Abermil, Pascale, Flandrin, Hannah, Moatti, Fabrizia, Favale, Ludovic, Suner, Florence, Lorre, Christophe, Marzac, Fanny, Fava, Anne-Claire, Mamez, Simona, Lapusan, Françoise, Isnard, Mohamad, Mohty, Ollivier, Legrand, Luc, Douay, Chrystele, Bilhou-Nabera, and François, Delhommeau
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Acute Myeloid Leukemia ,Adult ,Male ,Neoplasm, Residual ,Adolescent ,Article ,Fluorescence ,Clonal Evolution ,Young Adult ,Biomarkers, Tumor ,Humans ,Precision Medicine ,In Situ Hybridization, Fluorescence ,Alleles ,In Situ Hybridization ,Aged ,Aged, 80 and over ,Chromosome Aberrations ,Leukemia, Myeloid, Acute/diagnosis/genetics/mortality ,Neoplasm, Residual/diagnosis/genetics ,High-Throughput Nucleotide Sequencing ,Middle Aged ,Prognosis ,Leukemia, Myeloid, Acute ,Clonal Evolution/genetics ,Mutation ,Female - Abstract
The genetic landscape of adult acute myeloid leukemias (AML) has been recently unraveled. However, due to their genetic heterogeneity, only a handful of markers are currently used for the evaluation of minimal residual disease (MRD). Recent studies using multi-target strategies indicate that detection of residual mutations in less than 5% of cells in complete remission is associated with a better survival. Here, in a series of 69 AMLs with known clonal architecture, we design a clone-specific strategy based on fluorescent in situ hybridization and high-sensitivity next generation sequencing to detect chromosomal aberrations and mutations, respectively, in follow-up samples. The combination of these techniques allows tracking chromosomal and genomic lesions down to 0.5–0.4% of the cell population in remission samples. By testing all lesions in follow-up samples from 65 of 69 evaluable patients, we find that initiating events often persist and appear to be, on their own, inappropriate markers to predict short-term relapse. In contrast, the persistence of two or more lesions in more than 0.4% of the cells from remission samples is strongly associated with lower leukemia-free and overall survivals in univariate and multivariate analyses. Although larger prospective studies are needed to extend these results, our data show that a personalized, clone-specific, MRD follow up strategy is feasible in the vast majority of AML cases.
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- 2017
11. JAK3 deregulation by activating mutations confers invasive growth advantage in extranodal nasal-type natural killer cell lymphoma
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Laurianne Scourzic, Thomas Mercher, O. De Wever, William Vainchenker, Abdelghani Bouchekioua, P Cervera, Fawzia Louache, Paul Coppo, Luc Douay, Eric Solary, R Nyga, P. Gaulard, Yanyan Zhang, Christian Gespach, A Aline-Fardin, and D Jeziorowska
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Adult ,Male ,Cancer Research ,Tumor suppressor gene ,Cell Survival ,T cell ,medicine.disease_cause ,Mice ,Piperidines ,Cell Line, Tumor ,medicine ,Animals ,Humans ,Neoplasm Invasiveness ,Pyrroles ,Neoplasm Metastasis ,Phosphorylation ,STAT3 ,Protein Kinase Inhibitors ,Aged ,Cell Proliferation ,Neoplasm Staging ,Aged, 80 and over ,Mutation ,biology ,Cell growth ,Janus Kinase 3 ,Hematology ,Middle Aged ,medicine.disease ,Xenograft Model Antitumor Assays ,Tumor Burden ,Lymphoma ,Gene Expression Regulation, Neoplastic ,Lymphoma, Extranodal NK-T-Cell ,Disease Models, Animal ,Pyrimidines ,medicine.anatomical_structure ,Oncology ,Case-Control Studies ,biology.protein ,STAT protein ,Cancer research ,Female ,Janus kinase - Abstract
Extranodal, nasal-type natural killer (NK)/T-cell lymphoma (NKCL) is an aggressive malignancy with poor prognosis in which, usually, signal transducer and activator of transcription 3 (STAT3) is constitutively activated and oncogenic. Here, we demonstrate that STAT3 activation mostly results from constitutive Janus kinase (JAK)3 phosphorylation on tyrosine 980, as observed in three of the four tested NKCL cell lines and in 20 of the 23 NKCL tumor samples under study. In one of the cell lines and in 4 of 19 (21%) NKCL primary tumor samples, constitutive JAK3 activation was related to an acquired mutation (A573V or V722I) in the JAK3 pseudokinase domain. We then show that constitutive activation of the JAK3/STAT3 pathway has a major role in NKCL cell growth and survival and in the invasive phenotype. Indeed, NKCL cell growth was slowed down in vitro by targeting JAK3 with chemical inhibitors or small-interfering RNAs. In a human NKCL xenograft mouse model, tumor growth was significantly delayed by the JAK3 inhibitor CP-690550. Altogether, the constitutive activation of JAK3, which can result from JAK3-activating mutations, is a frequent feature of NKCL that deserves to be tested as a therapeutic target.
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- 2013
12. Genetic hierarchy and temporal variegation in the clonal history of acute myeloid leukaemia
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Hannah Moatti, François Delhommeau, Rémi Favier, Mohamad Mohty, Chrystele Bilhou-Nabera, Fawzia Louache, Virginie Joulin, Ruoping Tang, Aline Betems, Elodie Pronier, Florence Lorre, Pascale Flandrin, Ollivier Legrand, Fanny Fava, Christophe Marzac, Pierre Hirsch, Frédéric Féger, Hélène Boutroux, Hayat Mokrani, Luc Douay, Dominique Bories, Yanyan Zhang, Service d'hématologie clinique et de thérapie cellulaire [CHU Saint-Antoine], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-CHU Saint-Antoine [AP-HP], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Sorbonne Université (SU)-Sorbonne Université (SU), 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), MYPAC, Université Pierre et Marie Curie - Paris 6 (UPMC), 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), CHU Trousseau [APHP], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Sorbonne Université (SU), Institut Gustave Roussy (IGR), CHU Saint-Antoine [AP-HP], Hématologie moléculaire [CHU Mondor], CHU Henri Mondor, Laboratoire commun de biologie et génétique moléculaires [CHU Saint-Antoine], and CHU Henri Mondor [Créteil]
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0301 basic medicine ,Time Factors ,Myeloid ,Science ,Clone (cell biology) ,General Physics and Astronomy ,Biology ,medicine.disease_cause ,Somatic evolution in cancer ,Article ,General Biochemistry, Genetics and Molecular Biology ,Epigenesis, Genetic ,Clonal Evolution ,Mice ,03 medical and health sciences ,hemic and lymphatic diseases ,medicine ,Animals ,Humans ,neoplasms ,Variegation ,Gene Rearrangement ,[SDV.GEN]Life Sciences [q-bio]/Genetics ,Mutation ,Multidisciplinary ,Base Sequence ,General Chemistry ,Gene rearrangement ,medicine.disease ,Clone Cells ,Hematopoiesis ,Leukemia, Myeloid, Acute ,Leukemia ,Haematopoiesis ,030104 developmental biology ,medicine.anatomical_structure ,Immunology ,Single-Cell Analysis - Abstract
In acute myeloid leukaemia (AML) initiating pre-leukaemic lesions can be identified through three major hallmarks: their early occurrence in the clone, their persistence at relapse and their ability to initiate multilineage haematopoietic repopulation and leukaemia in vivo. Here we analyse the clonal composition of a series of AML through these characteristics. We find that not only DNMT3A mutations, but also TET2, ASXL1 mutations, core-binding factor and MLL translocations, as well as del(20q) mostly fulfil these criteria. When not eradicated by AML treatments, pre-leukaemic cells with these lesions can re-initiate the leukaemic process at various stages until relapse, with a time-dependent increase in clonal variegation. Based on the nature, order and association of lesions, we delineate recurrent genetic hierarchies of AML. Our data indicate that first lesions, variegation and treatment selection pressure govern the expansion and adaptive behaviour of the malignant clone, shaping AML in a time-dependent manner., Pre-leukaemic clones, together with the propensity to cause disease in mice, are characterized by appearing early in myeloid leukaemia and being found at relapse. Here, the authors identify clones in human samples and find that they are characterized by hierarchically organized genetic lesions, which can be used to track evolution of the disease.
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- 2016
13. Bio-engineered and native red blood cells from cord blood exhibit the same metabolomic profile
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Paul-Henri Romeo, Samia Boudah, Luc Douay, Tiffany Marie, Lydie Oliveira, Dhouha Darghouth, Christophe Junot, Nathalie Mario, Séverine Jolly, and Marie-Catherine Giarratana
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0301 basic medicine ,Erythrocytes ,Pharmacology ,Bioinformatics ,03 medical and health sciences ,0302 clinical medicine ,Metabolomics ,hemic and lymphatic diseases ,medicine ,Humans ,Online Only Articles ,Cell Engineering ,business.industry ,Stem Cells ,hemic and immune systems ,Hematology ,Leukapheresis ,Fetal Blood ,In vitro ,Peripheral blood ,030104 developmental biology ,medicine.anatomical_structure ,Cord blood ,Bone marrow ,business ,Metabolic profile ,circulatory and respiratory physiology ,030215 immunology - Abstract
The increasing need for red blood cells (RBCs) together with the lack of donors have made the in vitro production of RBCs a major medical challenge.[1][1] We have recently developed a method to produce mature RBCs in vitro , starting from bone marrow, peripheral blood, leukapheresis or cord blood
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- 2016
14. [In vitro generation of blood red cells from stem cells: a sketch of the future]
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Christelle, Mazurier and Luc, Douay
- Subjects
Adult ,Adult Stem Cells ,Erythrocytes ,Induced Pluripotent Stem Cells ,Cell Culture Techniques ,Humans ,Precision Medicine ,Regenerative Medicine ,Cells, Cultured - Abstract
Human adult pluripotent stem cells, stem cells of embryonic origin and induced pluripotent stem cells (iPS) provide cellular sources for new promising regenerative medicine approaches. Because these cells can be patient-specific, they allow considering a personalized medicine appropriate to the diagnosis of each. The generation of cultured red blood cells (cRBC) derived from stem cells is emblematic of personalized medicine. Indeed, these cells have the advantage of being selected according to a blood phenotype of interest and they may provide treatments to patients in situation of impossible transfusion (alloimmunized patients, rare phenotypes). Essential progresses have established proof of concept for this approach, still a concept some years ago. From adult stem cells, all steps of upstream research were successfully achieved, including the demonstration of the feasibility of injection into human. This leads us to believe that Red Blood Cells generated in vitro from stem cells will be the future players of blood transfusion. However, although theoretically ideal, these stem cells raise many biological challenges to overcome, although some tracks are identified.
- Published
- 2016
15. Des banques de cellules souches adultes pluripotentes comme source illimitée de globules rouges de culture: Un scénario pour demain
- Author
-
Thierry Peyrard and Luc Douay
- Subjects
Medical Laboratory Technology ,Biochemistry (medical) ,Analytical Chemistry - Abstract
Resume Les globules rouges peuvent maintenant etre cultives in vitro a partir de plusieurs sources de cellules souches humaines: hematopoietiques, embryonnaires (ES) ou cellules matures humaines reprogrammees en cellules pluripotentes (human induced pluripotent stem cells, hiPSC). La technologie tres prometteuse des hiPSC represente une source potentiellement illimitee de GR et ouvre la voie au possible developpement d’une nouvelle generation de produits transfusionnels allogeniques. En admettant que la culture a grande echelle in vitro des GR s’opere efficacement dans un proche avenir, nous decrivons ici un scenario futuriste, mais realiste, concernant les applications potentielles pour les patients alloimmunises et ceux ayant un groupe sanguin rare. Nous avons etabli que 3 clones d’hiPSC seulement seraient suffisants pour fournir des cellules compatibles a plus que 99% des patients necessitant des transfusions de GR. L’etude du Registre national francais des personnes ayant un phenotype/genotype sanguin rare revele que 15 clones d’hiPSC couvriraient 100% des besoins des patients d’origine caucasienne. De plus, un seul clone d’hiPSC satisferait 73% des besoins des patients drepanocytaires alloimmunises qui necessitaient jusqu’ici des unites de GR rares cryopreserves. Ainsi, un nombre tres limite de clones de GR non seulement serait en mesure de couvrir les besoins de la plupart des patients alloimmunises et de ceux ayant un groupe sanguin rare, mais permettrait aussi une politique efficace de prevention de l’alloimmunisation chez les patients polytransfuses.
- Published
- 2012
16. Design and validation of a consistent and reproducible manufacture process for the production of clinical-grade CD34+ expanded stem cells
- Author
-
E. Mai, A. Black, Hélène Rouard, S. Derenne, Sandrine Vogt, Anthony Criquet, Philippe Henon, Joe-Elie Salem, Laurence Harmand, Luc Douay, Claire Saucourt, Brigitte Birebent, and A. Chartois-Leauté
- Subjects
Cancer Research ,Transplantation ,business.industry ,Computer science ,Immunology ,CD34 ,Clinical grade ,Cell Biology ,Oncology ,Scientific method ,Immunology and Allergy ,Production (economics) ,Stem cell ,Process engineering ,business ,Genetics (clinical) - Published
- 2017
17. Patents on Red Blood Cell Manufacturing In Vitro
- Author
-
Laurence Guyonneau-Harmand and Luc Douay
- Subjects
Red blood cell ,medicine.anatomical_structure ,Developmental Neuroscience ,Chemistry ,medicine ,Cell Biology ,Molecular biology ,In vitro ,Developmental Biology - Published
- 2011
18. RETRACTED: From Stem Cell to Red Blood Cells In Vitro: 'The 12 Labors of Hercules'
- Author
-
Luc Douay
- Subjects
business.industry ,Biochemistry (medical) ,Clinical Biochemistry ,In vitro ,Blood substitute ,Cell biology ,Clinical Practice ,Blood cell ,Haematopoiesis ,Red blood cell ,medicine.anatomical_structure ,medicine ,Stem cell ,business ,Induced pluripotent stem cell - Abstract
This article describes the research in progress that will permit the large-scale production of human red blood cells from hematopoietic stem cells. It also discusses the current state of this research, suggests the obstacles to be overcome to pass from the laboratory model to clinical practice, and analyzes the possible indications in the medium and long term. The potential interest of pluripotent stem cells as an unlimited source of red blood cells is considered. If it succeeds, this new approach could mark a considerable advance in the field of transfusion.
- Published
- 2010
19. Stem cell therapy for the treatment of severe tissue damage after radiation exposure
- Author
-
Noëlle Mathieu, J. Simon, M. Mothy, Christelle Demarquay, Christine Linard, Jean-Jacques Lataillade, Norbert Claude Gorin, Luc Douay, A. Semont, Alain Chapel, Claire Squiban, and Hélène Rouard
- Subjects
Cancer Research ,Transplantation ,Pathology ,medicine.medical_specialty ,business.industry ,medicine.medical_treatment ,Immunology ,Cell ,Cell Biology ,Stem-cell therapy ,Radiation exposure ,medicine.anatomical_structure ,Oncology ,Tissue damage ,medicine ,Immunology and Allergy ,business ,Genetics (clinical) - Published
- 2018
20. Author Correction: Synergistic effect of human Bone Morphogenic Protein-2 and Mesenchymal Stromal Cells on chronic wounds through hypoxia-inducible factor-1 α induction
- Author
-
Véronique Eder, Norbert-Claude Gorin, Karim Belmokhtar, Luc Douay, Sabine François, Marc Benderitter, Alain Chapel, and Marie-Christine Machet
- Subjects
0301 basic medicine ,Hypoxia-Inducible Factor 1 ,Multidisciplinary ,business.industry ,Mesenchymal stem cell ,lcsh:R ,Human bone ,lcsh:Medicine ,03 medical and health sciences ,030104 developmental biology ,Text mining ,Cancer research ,ComputingMethodologies_DOCUMENTANDTEXTPROCESSING ,Medicine ,lcsh:Q ,Author Correction ,business ,lcsh:Science - Abstract
Chronic skin ulcers and burns require advanced treatments. Mesenchymal Stromal Cells (MSCs) are effective in treating these pathologies. Bone Morphogenic Protein-2 (BMP-2) is known to enhance angiogenesis. We investigated whether recombinant human hBMP-2 potentiates the effect of MSCs on wound healing. Severe ulceration was induced in rats by irradiation and treated by co-infusion of MSCs with hBMP-2 into the ulcerated area which accelerated wound healing. Potentiation of the effect of MSCs by hBMP-2 on endothelial repair improved skin healing. HBMP-2 and MSCs synergistically, in a supra additive or enhanced manner, renewed tissue structures, resulting in normalization of the epidermis, hair follicles, sebaceous glands, collagen fibre density, and blood vessels. Co-localization of MSCs with CD31 + cells suggests recruitment of endothelial cells at the site of injection. HBMP-2 and MSCs enhanced angiogenesis and induced micro-vessel formation in the dermis where hair follicles were regenerated. HBMP-2 acts by causing hypoxia-inducible factor-1 α (HIF-1α) expression which impacts endothelial tube formation and skin repair. This effect is abolished by siRNA. These results propose that new strategies adding cytokines to MSCs should be evaluated for treating radiation-induced dermatitis, burns, and chronic ulcers in humans.
- Published
- 2018
21. Perspectives transfusionnelles des cellules souches: le modèle des globules rouges de culture
- Author
-
Luc Douay, Hélène Lapillonne, and Ali G. Turhan
- Subjects
Hematology - Abstract
Dans un contexte de difficulte chronique d’approvisionnement en produits sanguins, l’interet de disposer de sources complementaires de globules rouges pour la transfusion sanguine est evident. La mise au point de molecules chimiques ou naturelles qui remplaceraient l’hemoglobine se revele difficile. Le sang artificiel est encore hors d’atteinte. Ainsi, au lieu de remplacer ce que fait la nature, pourquoi ne pas la copier ? Pour ces raisons, tenter de generer des globules rouges dans un laboratoire, fait sens. Nous decrivons ici les recherches en cours qui permettront la generation massive d’hematies humaines a partir de cellules souches hematopoietiques et pluripotentes. Nous faisons le point sur l’etat de l’art de ce concept, evoquons les obstacles a surmonter pour passer du modele de laboratoire a la clinique, et analysons les indications possibles a moyen et long termes. Si elle aboutit, cette nouvelle approche pourrait etre un progres considerable en matiere de transfusion.
- Published
- 2010
22. Extensive mutational status of genes and clinical outcome in pediatric acute myeloid leukemia
- Author
-
Philippe N, Llopis L, Guy Leverger, Paola Ballerini, Mazingue F, Claude Preudhomme, Lai Jl, Mircea Adam, Myriam Labopin, Christine Perot, Luc Douay, Hélène Lapillonne, Zurawski, Aline Renneville, A. Auvrignon, and Judith Landman-Parker
- Subjects
Male ,Oncology ,Cancer Research ,medicine.medical_specialty ,Genes, Wilms Tumor ,Adolescent ,Biology ,hemic and lymphatic diseases ,Internal medicine ,medicine ,Humans ,Mutational status ,Child ,Gene ,Hematology ,Pediatric acute myeloid leukemia ,Infant, Newborn ,Infant ,Cancer ,Histone-Lysine N-Methyltransferase ,medicine.disease ,Leukemia, Myeloid, Acute ,Genes, ras ,fms-Like Tyrosine Kinase 3 ,Child, Preschool ,Multivariate Analysis ,Mutation ,Immunology ,Female ,Myeloid-Lymphoid Leukemia Protein - Abstract
Extensive mutational status of genes and clinical outcome in pediatric acute myeloid leukemia
- Published
- 2009
23. Culture de cellules à visée transfusionnelle : le cas des globules rouges
- Author
-
Luc Douay
- Subjects
Biochemistry (medical) ,Clinical Biochemistry ,Hematology - Abstract
Resume Dans un contexte de difficulte chronique d’approvisionnement en produits sanguins, l’interet de disposer de sources complementaires de globules rouges (GR) pour la transfusion sanguine est evident. La mise au point de molecules chimiques ou naturelles qui remplaceraient l’hemoglobine se revele difficile. Le sang artificiel est encore hors d’atteinte. Ainsi au lieu de remplacer ce que fait la nature pourquoi ne pas la copier ? Pour ces raisons, tenter de generer au laboratoire des GR fait sens. Nous decrivons ici les recherches en cours qui permettront la generation massive de GR humains a partir de cellules souches hematopoietiques. Nous faisons le point sur l’etat de l’art de ce concept, evoquons les obstacles a surmonter pour passer du modele de laboratoire a la clinique et analysons les indications possibles a moyen et long terme. Si elle aboutit, cette nouvelle approche pourrait etre un progres considerable en matiere de transfusion sanguine.
- Published
- 2009
24. Molecular signature of erythroblast enucleation in human embryonic stem cells
- Author
-
Nicolas Hebert, Anne-Marie Faussat, Charles Durand, Marie Cambot, Hélène Lapillonne, Shaghayegh Rouzbeh, Christelle Mazurier, Luc Douay, Ladan Kobari, CHU Saint-Antoine [AP-HP], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Sorbonne Université (SU), Institut National de la Transfusion Sanguine [Paris] (INTS), Cancer, Inflammation, Hormones, Université Pierre et Marie Curie - Paris 6 (UPMC)-Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Institut National de la Santé et de la Recherche Médicale (INSERM), Laboratoire de Biologie du Développement [IBPS] (LBD), Sorbonne Université (SU)-Centre National de la Recherche Scientifique (CNRS)-Institut de Biologie Paris Seine (IBPS), Institut National de la Santé et de la Recherche Médicale (INSERM)-Sorbonne Université (SU)-Centre National de la Recherche Scientifique (CNRS)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), CHU Trousseau [APHP], Labex GR-Ex fellowship, Etablissement Francais du Sang (EFS), Fondation pour la recherche medicale, Combattre La Leucemie, DIM Stempole (Paris, France), ANR-11-IDEX-0005,USPC,Université Sorbonne Paris Cité(2011), Laboratoire de Biologie du Développement [Paris] (LBD), Sorbonne Université (SU)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), HAL-UPMC, Gestionnaire, and Université Sorbonne Paris Cité - - USPC2011 - ANR-11-IDEX-0005 - IDEX - VALID
- Subjects
Cell type ,Embryonic stem cells ,Erythroblasts ,Enucleation ,Human Embryonic Stem Cells ,Embryoid body ,Biology ,MiR-30a ,Erythroblast ,hemic and lymphatic diseases ,[SDV.BBM.GTP]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Genomics [q-bio.GN] ,microRNA ,Humans ,Erythropoiesis ,RNA, Messenger ,Cells, Cultured ,Gene knockdown ,Cell Differentiation ,Cell Biology ,Anatomy ,Embryonic stem cell ,Cell biology ,MicroRNAs ,Gene Expression Regulation ,Molecular Medicine ,[SDV.BBM.GTP] Life Sciences [q-bio]/Biochemistry, Molecular Biology/Genomics [q-bio.GN] ,Developmental Biology - Abstract
While enucleation is a critical step in the terminal differentiation of human red blood cells, the molecular mechanisms underlying this unique process remain unclear. To investigate erythroblast enucleation, we studied the erythroid differentiation of human embryonic stem cells (hESCs), which provide a unique model for deeper understanding of the development and differentiation of multiple cell types. First, using a two-step protocol, we demonstrated that terminal erythroid differentiation from hESCs is directly dependent on the age of the embryoid bodies. Second, by choosing hESCs in two extreme conditions of erythroid culture, we obtained an original differentiation model which allows one to study the mechanisms underlying the enucleation of erythroid cells by analyzing the gene and miRNA (miR) expression profiles of cells from these two culture conditions. Third, using an integrated analysis of mRNA and miR expression profiles, we identified five miRs potentially involved in erythroblast enucleation. Finally, by selective knockdown of these five miRs we found miR-30a to be a regulator of erythroblast enucleation in hESCs. Stem Cells 2015;33:2431–2441
- Published
- 2015
25. Autologous Bone Marrow Transplantation with Marrow Decontaminated by Immunotoxin T 101 in the Treatment of Leukemia and Lymphoma
- Author
-
Raphaël David, Stachowiak J, B. Rio, G. A. Voisin, Norbert-Claude Gorin, J. Jansen, G. Le Blanc, P. Poncelet, Luc Douay, J. P. Laporte, M. Lopez, Albert Najman, Salmon C, Duhamel G, R. Zittoun, M. C. Liance, Deloux J, and P. Cazellas
- Subjects
Leukemia ,Immunotoxin ,Marrow transplantation ,business.industry ,Cancer research ,medicine ,medicine.disease ,Autologous bone ,business ,Lymphoma - Published
- 2015
26. A novel micronucleus in vitro assay utilizing human hematopoietic stem cells
- Author
-
Natalia Kotova, Dag Jenssen, Daniel Vare, Luc Douay, Eva-Lena Härnwall, Christelle Mazurier, Nicolas Hebert, Jan Grawé, Administateur, HAL Sorbonne Université, Stockholm University, 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), Etablissement français du sang [Bourgogne-Franche-Comté] (EFS [Bourgogne-Franche-Comté]), Service d'immunologie et hématologies biologiques [CHU Saint-Antoine], Université Pierre et Marie Curie - Paris 6 (UPMC)-Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-CHU Saint-Antoine [AP-HP], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Sorbonne Université (SU)-Sorbonne Université (SU), Uppsala University, Service d'immunologie et hématologies biologiques [Saint-Antoine], Etablissement français du sang [Bourgogne-Franche-Comté] (EFS BFC), CHU Saint-Antoine [AP-HP], and Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Sorbonne Université (SU)
- Subjects
Erythrocytes ,Pharmacology and Toxicology ,Stem cells ,Biology ,Toxicology ,medicine.disease_cause ,Flow cytometry ,03 medical and health sciences ,0302 clinical medicine ,Micronucleus test ,medicine ,Humans ,Cells, Cultured ,030304 developmental biology ,0303 health sciences ,Micronucleus Tests ,medicine.diagnostic_test ,Genotoxicity in vitro ,General Medicine ,Farmakologi och toxikologi ,Hematopoietic Stem Cells ,Molecular biology ,Coculture Techniques ,3. Good health ,[SDV.TOX] Life Sciences [q-bio]/Toxicology ,Haematopoiesis ,medicine.anatomical_structure ,030220 oncology & carcinogenesis ,[SDV.TOX]Life Sciences [q-bio]/Toxicology ,Bone marrow ,Stem cell ,Micronucleus ,Genotoxicity ,Ex vivo ,Mutagens - Abstract
The induction of micronucleated reticulocytes in the bone marrow is a sensitive indicator of chromosomal damage. Therefore, the micronucleus assay in rodents is widely used in genotoxicity and carcinogenicity testing. A test system based on cultured human primary cells could potentially provide better prediction compared to animal tests, increasing patient safety while also implementing the 3Rs principle, i.e. replace, reduce and refine. Hereby, we describe the development of an in vitro micronucleus assay based on animal-free ex vivo culture of human red blood cells from hematopoietic stem cells. To validate the method, five clastogens with direct action, three clastogens requiring metabolic activation, four aneugenic and three non-genotoxic compounds have been tested. Also, different metabolic systems have been applied. Flow cytometry was used for detection and enumeration of micronuclei. Altogether, the results were in agreement with the published data and indicated that a sensitive and cost effective in vitro assay to assess genotoxicity with a potential to high-throughput screening has been developed. (C) 2015 The Authors. Published by Elsevier Ltd.
- Published
- 2015
27. Ex Vivo Expansion Does Not Alter the Capacity of Umbilical Cord Blood CD34+Cells to Generate Functional T Lymphocytes and Dendritic Cells
- Author
-
Jean C. Gluckman, Luc Douay, Marie C. Giarratana, Michelle Rosenzwajg, and Ladan Kobari
- Subjects
Receptors, Antigen, T-Cell, alpha-beta ,T-Lymphocytes ,CD34 ,Antigens, CD34 ,Mice, SCID ,Dendritic cell differentiation ,Biology ,Interferon-gamma ,Mice ,Organ Culture Techniques ,Mice, Inbred NOD ,Animals ,Humans ,Cytotoxic T cell ,Lymphopoiesis ,Antigen-presenting cell ,Cells, Cultured ,Follicular dendritic cells ,Cell Differentiation ,Dendritic Cells ,Cell Biology ,Fetal Blood ,Cell biology ,Phenotype ,Cord blood ,Immunology ,Molecular Medicine ,Stem cell ,Developmental Biology - Abstract
We examined whether ex vivo expansion of umbilical cord blood progenitor cells affected their capacity to generate immune cells such as T lymphocytes (TLs) and dendritic cells (DCs). The capacity to generate TLs from cord blood CD34(+) cells expanded for 14 days (d14) was compared with that of nonexpanded CD34(+) cells (d0) using fetal thymus organ cultures or transfer into nonobese diabetic/severe combined immunodeficient mice. The cell preparations yielded comparable percentages of immature (CD4(+)CD8(-), CD4(+)CD8(+)) TLs and functional mature (CD3(+)CD4(+), CD3(+)CD8(+)) TLs with an analogous TCR (T-cell receptor)-Vbeta repertoire pattern. As regards DCs, d0 and d14 CD34(+) cells also yielded similar percentages of CD1a(+) DCs with the same expression levels of HLA-DR, costimulatory and adhesion molecules, and chemokine receptors. DCs derived from either d14 or d0 CD34(+) stimulated allogeneic TLs to the same extent, and the cytokine pattern production of these allogeneic TLs was similar with no shift toward a predominant Th1 or Th2 response. Even though the intrinsic capacity of d14 CD34(+) cells to generate DCs was 13-fold lower than that of d0 CD34(+) cells, this reduction was offset by the prior amplification of the CD34(+) cells, resulting in the overall production of 15-fold more DCs. These data indicate that ex vivo expansion of CD34(+) cells does not impair T lymphopoiesis nor DC differentiation capacity.
- Published
- 2006
28. High WT1 Expression After Induction Therapy Predicts High Risk of Relapse and Death in Pediatric Acute Myeloid Leukemia
- Author
-
Françoise Mazingue, Myriam Labopin, Jean-Luc Laï, Claude Preudhomme, Axelle Dehee, Judith Landman-Parker, Christine Perot, Mircea Adam, Cyril Flamant, Annick Blaise, Sylvie Fasola, Hélène Lapillonne, Anne Auvrignon, Aline Renneville, Françoise Bellman, Guy Leverger, Luc Douay, and Paola Ballerini
- Subjects
Adult ,Male ,Oncology ,Acute promyelocytic leukemia ,Cancer Research ,medicine.medical_specialty ,Pathology ,Genes, Wilms Tumor ,Myeloid ,Adolescent ,Oncogene Proteins, Fusion ,Gene Dosage ,Gene dosage ,RUNX1 Translocation Partner 1 Protein ,Recurrence ,Internal medicine ,medicine ,Humans ,RNA, Messenger ,Child ,Reverse Transcriptase Polymerase Chain Reaction ,business.industry ,Infant, Newborn ,Cytogenetics ,Infant ,Wilms' tumor ,medicine.disease ,Minimal residual disease ,Neoplasm Proteins ,Leukemia, Myeloid, Acute ,Leukemia ,medicine.anatomical_structure ,El Niño ,Child, Preschool ,Core Binding Factor Alpha 2 Subunit ,Multivariate Analysis ,Female ,business - Abstract
Purpose To determine whether minimal residual disease (MRD) measured by Wilms' tumor gene 1 (WT1) expression is a prognostic marker in pediatric acute myeloid leukemia (AML), we quantified WT1 transcript by real-time quantitative-polymerase chain reaction in 92 AML at diagnosis and during follow-up. Patients and Methods Patients (median age, 6 years; cytogenetics, favorable 27%, intermediate 59%, poor 13%) were treated between 1995 and 2002 and enrolled in Leucémie aiguë Myéloblastique Enfant (LAME) 89/91, LAME 99 pilot study and Acute Promyelocytic Leukemia French collaborative protocols. With a median follow-up of 26 months, event-free survival was 56% with a standard deviation (SD) of 5% and overall survival of 62.5% with an SD of 6%. WT1 copy number was normalized by TATA box binding protein gene transcripts and expressed as WT1/TBP × 1,000 ratio. Median WT1 ratio in normal patient controls was 12 (range, 0 to 57). A level over two SD than normal bone marrow controls (ie, WT1 ratio > 50), was considered as significant overexpression. Results At diagnosis, WT1 overexpression was detected in 78% of patients (72 of 92 patients; median copy ratio, 2231). The WT1 values were significantly higher (P = .01) in favorable cytogenetics and lower (P < .0001) in M5-FAB subtype, 11q23 rearrangements (P < .001), and infants (P = .003) and demonstrate a strong correlation with fusion transcript AML1-ETO, PML-RARα expression. After induction treatment, WT1 ratio was analyzed in 46 of 72 patients and found above 50 in nine of 36 patients and five of 25 patients at D35-50 and 3 to 5 months, respectively. WT1 ratio > 50 after induction is an independent prognostic risk factor of relapse (P = .002) and death (P = .02). Conclusion WT1 quantification is an informative molecular marker for MRD in pediatric AML and is now performed as prospective analysis in ELAM02 protocol.
- Published
- 2006
29. Génération in vitro de globules rouges humains matures et fonctionnels : un modèle d’étude aux perspectives multidisciplinaires
- Author
-
Marie-Catherine Giarratana and Luc Douay
- Subjects
Blood cell ,Red blood cell ,Haematopoiesis ,medicine.anatomical_structure ,Reticulocyte ,Chemistry ,Cellular differentiation ,medicine ,Erythropoiesis ,Stem cell factor ,General Medicine ,Stem cell ,Cell biology - Abstract
We describe a technical approach permitting massive expansion of CD34+ stem cells (up to 1.95 x 10(6)-fold) and their full ex vivo conversion into mature red blood cells (RBCs). This three-step protocol can be adapted to hematopoietic stem cells (HSC) of various origins. First, cell proliferation and erythroid differentiation are induced in serum-free media supplemented with stem cell factor, interleukin-3 and erythropoietin (Epo) for 8 days. The cells are then co-cultured with either the murine stromal cell line MS-5 or human mesenchymal cells for 3 days in the presence of Epo alone. Finally, all exogenous factors are withdrawn and the cells are incubated on a simple stroma for up to 10 days. The ex vivo microenvironment strongly influences both the terminal maturation of erythroid cells and hemoglobin (Hb) synthesis. Critically, in vitro-generated RBCs have all the characteristics of functional native adult RBCs in terms of their enzyme content, membrane deformability, and capacity to fix and release oxygen. In addition, their behavior in the murine NOD/SCID model mirrors that of native RBCs. This new concept of "cultured RBCs" (cRBC) has major implications for basic research on terminal erythropoiesis and for patient management. Currently, the potential yield of functional red cells is compatible with clinical requirements, as several units of packed RBCs can be produced from a single donation. Importantly, infused cRBC would all have a life-span of about 120 days, whereas the mean half-life of normal donor RBCs is only 28 days. This would help to minimize the transfusion exposure of patients requiring regular treatment, thereby reducing the risk of iron overload and allo-immunization. The use of autologous CD34+ cells isolated from leukapheresis samples could be beneficial for patients who no longer tolerate allogeneic RBCs. This new method should also prove useful for analyzing the mechanisms of terminal erythropoiesis, including hemoglobin synthesis. Finally, it could provide a tool for investigating the lifecycle of blood parasites such as Plasmodium, the agent of malaria.
- Published
- 2005
30. Ex vivo generation of fully mature human red blood cells from hematopoietic stem cells
- Author
-
Laurent Kiger, Michael C. Marden, David Chalmers, Luc Douay, Ladan Kobari, Hélène Lapillonne, Thérèse Cynober, Marie-Catherine Giarratana, and Henri Wajcman
- Subjects
Erythrocytes ,Reticulocytes ,Time Factors ,Stromal cell ,Ultraviolet Rays ,Cellular differentiation ,Cell Culture Techniques ,Biomedical Engineering ,CD34 ,Antigens, CD34 ,Bioengineering ,Cell Separation ,Mice, SCID ,Biology ,Applied Microbiology and Biotechnology ,Hemoglobins ,Mice ,Mice, Inbred NOD ,hemic and lymphatic diseases ,medicine ,Animals ,Humans ,Cells, Cultured ,Erythroid Precursor Cells ,Microscopy, Confocal ,Stem Cells ,Cell Differentiation ,Genetic Therapy ,Flow Cytometry ,Hematopoietic Stem Cells ,Coculture Techniques ,Cell biology ,Oxygen ,Haematopoiesis ,Red blood cell ,medicine.anatomical_structure ,Immunology ,Cytokines ,Molecular Medicine ,Erythropoiesis ,Stem cell ,Ex vivo ,Biotechnology - Abstract
We describe here the large-scale ex vivo production of mature human red blood cells (RBCs) from hematopoietic stem cells of diverse origins. By mimicking the marrow microenvironment through the application of cytokines and coculture on stromal cells, we coupled substantial amplification of CD34(+) stem cells (up to 1.95 x 10(6)-fold) with 100% terminal differentiation into fully mature, functional RBCs. These cells survived in nonobese diabetic/severe combined immunodeficient mice, as do native RBCs. Our system for producing 'cultured RBCs' lends itself to a fundamental analysis of erythropoiesis and provides a simple in vitro model for studying important human viral or parasitic infections that target erythroid cells. Further development of large-scale production of cultured RBCs will have implications for gene therapy, blood transfusion and tropical medicine.
- Published
- 2005
31. A novel real-time RT-PCR assay for quantification of OTT-MAL fusion transcript reliable for diagnosis of t(1;22) and minimal residual disease (MRD) detection
- Author
-
A Blaise, E Gatbois, Beatrice Pellegrino, Roland Berger, Thomas Mercher, Christine Perot, Paola Ballerini, Judith Landman-Parker, J van den Akker, Mircea Adam, Luc Douay, and O. Bernard
- Subjects
Cancer Research ,Oncogene Proteins ,RNA-binding protein ,Chromosomal translocation ,Hematology ,Biology ,medicine.disease ,Molecular biology ,Minimal residual disease ,Leukemia ,Real-time polymerase chain reaction ,Oncology ,Fusion transcript ,Antigen ,hemic and lymphatic diseases ,medicine - Abstract
A novel real-time RT-PCR assay for quantification of OTT-MAL fusion transcript reliable for diagnosis of t(1;22) and minimal residual disease (MRD) detection
- Published
- 2003
32. HOX11L2 expression defines a clinical subtype of pediatric T-ALL associated with poor prognosis
- Author
-
Paola Ballerini, Maryvonne Busson-Le Coniat, Beatrice Pellegrino, Christine Perot, Mircea Adam, Roland Berger, Jessica Zucman-Rossi, Xin Ying Su, Luc Douay, Annick Blaise, Olivier Bernard, Jacqueline Van Den Akker, and Judith Landman-Parker
- Subjects
Male ,medicine.medical_specialty ,Leukemia, T-Cell ,Neoplasm, Residual ,Adolescent ,Immunology ,Locus (genetics) ,Biology ,Biochemistry ,Immunophenotyping ,Proto-Oncogene Proteins ,Acute lymphocytic leukemia ,Metalloproteins ,Basic Helix-Loop-Helix Transcription Factors ,medicine ,Humans ,Gene family ,Child ,Cyclin-Dependent Kinase Inhibitor p16 ,T-Cell Acute Lymphocytic Leukemia Protein 1 ,Adaptor Proteins, Signal Transducing ,Chromosome Aberrations ,Homeodomain Proteins ,Oncogene Proteins ,Gene Expression Profiling ,Cytogenetics ,Infant ,Cell Biology ,Hematology ,Gene rearrangement ,LIM Domain Proteins ,Precursor Cell Lymphoblastic Leukemia-Lymphoma ,Prognosis ,medicine.disease ,Survival Analysis ,Minimal residual disease ,Neoplasm Proteins ,DNA-Binding Proteins ,Childhood T Acute Lymphoblastic Leukemia ,Child, Preschool ,Cytogenetic Analysis ,Cancer research ,Female ,Transcription Factors ,TAL1 - Abstract
The most frequent oncogenic activation events characterized in childhood T acute lymphoblastic leukemia (T-ALL) result in the transcriptional activation of genes coding for transcription factors. The main genes are TAL1/SCL, a member of the basic region helix-loop-helix gene family, and HOX11L2, a member of the homeobox-containing protein family. To gain insight into the pathogenesis of this type of hematologic malignancy, we analyzed 28 T-ALL samples. SIL-TAL1/SCL fusion was detected in 6 patients; expression of HOX11L2 was observed in 6 patients and of HOX11 in 3 patients. With one exception, these activations did not occur simultaneously in the same patients, and they allowed the subclassification of 50% of the patients.SIL-TAL1 fusion was detected in association withHOX11 expression in one patient and with a t(8;14) (q24;q11) in another. High expression of LYL1,LMO2, or TAL1 was observed mainly in samples negative for HOX11L2 expression. HOX11L1 andHOX11 expression were observed in one instance each, in the absence of detectable chromosomal abnormality of their respective loci, on chromosomes 2 and 10, respectively. HOX11L2 expression was associated with a chromosome 5q abnormality, the location of theHOX11L2 locus in each case tested. Finally, our data show that HOX11L2 expression was a suitable marker for minimal residual disease follow-up and was significantly associated with relapse (P = .02).
- Published
- 2002
33. Production massive de précurseurs de globules rougesin vitrovers un nouveau produit sanguin labile ?
- Author
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Luc Douay and Thi My Anh Neildez-Nguyen
- Subjects
Blood cell ,Haematopoiesis ,Red blood cell ,medicine.anatomical_structure ,Chemistry ,medicine ,General Medicine ,Progenitor cell ,Stem cell ,Molecular biology ,General Biochemistry, Genetics and Molecular Biology ,In vitro - Published
- 2002
34. Improved efficiency of remission induction facilitates autologous BMT harvesting and improves overall survival in adults with AML: 108 patients treated at a single institution
- Author
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Laporte Jp, S Lesage, M Aoudjhane, Norbert-Claude Gorin, L. Fouillard, Albert Najman, P Zunic, M Elloumi, Françoise Isnard, Marcelo F. Lopez, J van den Akker, M Guiguet, N. Cheron, Luc Douay, and Deloux J
- Subjects
Adult ,Amsacrine ,Male ,medicine.medical_specialty ,Adolescent ,medicine.medical_treatment ,Urology ,Transplantation, Autologous ,chemistry.chemical_compound ,Mafosfamide ,White blood cell ,Antineoplastic Combined Chemotherapy Protocols ,medicine ,Humans ,Survival rate ,Bone Marrow Transplantation ,Etoposide ,Retrospective Studies ,Transplantation ,Chemotherapy ,business.industry ,Remission Induction ,Cytarabine ,Hematology ,Middle Aged ,medicine.disease ,Surgery ,Survival Rate ,Leukemia ,Treatment Outcome ,medicine.anatomical_structure ,chemistry ,Leukemia, Myeloid ,Acute Disease ,Female ,Bone marrow ,business ,medicine.drug - Abstract
A hundred and eight patients less than 60 years old with de novo acute myeloid leukemia were treated between 1982 and 1994 by protocols including final intensification with a transplant using autologous bone marrow purged by mafosfamide in first remission in the absence of an HLA-matched sibling donor available for allograft. From 1989, we attempted to improve tumor control by using high-dose anthracyclines in induction, by increasing from one to two the number of consolidation courses pre-transplant and by introducing intermediate doses of cytarabine in the first consolidation course. The CR rate was 77% (33/43) before 1989 and 90% (59/65) after 1989 (P = 0.06). Forty-five out of the 59 patients (76%) who achieved CR after 1989 could undergo bone marrow grafting in CR1 vs 16/33 (48%) before 1989 (P = 0.01). In spite of the higher proportion of patients above 50 years after 1989 (32%) toxicity was mild and an adequate graft was obtained more frequently after one collection. The principal factor relating to improvement in graft feasibility was the post-1989 modification of induction and consolidation regimens. This improvement in graft feasibility was associated with a better disease-free survival (DFS) (48 +/- 7% vs 32 +/- 8%, P = 0.04) and overall survival (OS) (53 +/- 6% vs 30 +/- 7%, P = 0.007) at 5 years. By multivariate analysis four factors were associated with overall survival (OS): karyotype, white blood cell count at diagnosis, treatment regimen and bone marrow grafting in CR1. This global approach should be prospectively compared with intensive chemotherapy.
- Published
- 2001
35. Clono-Specific Evaluation of Minimal Residual Disease in Acute Myeloid Leukemia
- Author
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Chrystele bilhou Nabera, François Delhommeau, Pierre Hirsch, Ruoping Tang, Ollivier Legrand, Nassera Abermil, Luc Douay, Hannah Moatti, Pascale Flandrin, and Mohamad Mohty
- Subjects
NPM1 ,Chemotherapy ,medicine.medical_specialty ,Pathology ,IDH1 ,business.industry ,medicine.medical_treatment ,Immunology ,Myeloid leukemia ,Cell Biology ,Hematology ,Biochemistry ,Minimal residual disease ,Gastroenterology ,Regimen ,Internal medicine ,CEBPA ,Cytarabine ,Medicine ,business ,medicine.drug - Abstract
Background: The genetic landscape of adult acute myeloid leukemias (AML) has been recently unravelled. This makes achievable the determination of a comprehensive profile of driver lesions for virtually all patients at diagnosis. Recent studies using multi-target minimal residual disease (MRD) strategies with around 1% sensitivity indicate that the clearance of all molecular events after chemotherapy is associated with better survival. To improve the clono-specificity and the sensitivity of this approach, after a precise determination of AML clonal composition, we combined cytogenetic, FISH, and high sensitivity deep sequencing technologies to monitor the MRD in a series of 69 patients. Methods: Forty-five consecutive patients reflecting the genetic diversity of AML were prospectively included and 24 patients were retrospectively studied. All patients received an anthracycline + cytarabine based regimen. The clonal architecture was established at diagnosis based on NGS-targeted resequencing (122 gene panel) and cytogenetic data. Lesions were next investigated in complete remission (CR). Based on the initial clonal composition, targeted resequencing panels were designed to improve the sensitivity by the use of unique molecular barcodes (Haloplex High Sensitivity, or HS-NGS assay). Cytogenetic events were evaluated by FISH. Results: In the 69 patients, a median of 4 genetic or chromosomal events were identified per patient (range 0-10). One patient had no evaluable target allowing MRD evaluation in 68/69 patients. To determine the threshold of detection of the HS NGS assay, we analyzed the frequency of mutant reads in multiple samples expected to be wild type for 31 given SNVs and 2 indels. A consensus threshold of detection was set at a variant allele frequency (VAF) of 0.2% for all lesions. In CR samples, early initiating events frequently persisted after treatment, especially mutations in DNMT3A, TET2, ASXL1, EZH2, IDH1, TP53, SRSF2, and U2AF1. Mutations in FLT3, NRAS, KIT, NPM1, CEBPA, WT1, IDH2 and BCOR were the most frequently cleared events. Seven patients did not reach CR after one course, and two had no available material after one course. In the 59 remaining patients, we tested whether the global response level of all targets was associated with prognosis. We used the median VAF of the first events of all clonal architectures to separate good responder from poor responders (i.e. VAF = 1.66%). At 2 years, there was a trend to lower leukemia free survival (LFS) probability in poor responders (31.7+/-9.9% vs 51.7+/-9.8%, p=0.08) with no translation in overall survival (OS). We next investigated if the persistence of two or more detectable markers was associated with prognosis. The 58 patients with more than one evaluable event were consequently separated in two groups: patients with 0 or 1 marker above the detection threshold after treatment (n=31), and patients with 2 or more detectable lesions (n=27). At 2 years, DFS was 64.9+/-9.3 % in patients with 0 or 1 detectable marker vs 19.8+/-8.7% in patients with 2 or more detectable markers (p=0.001). OS probability was higher in patients with 0 or 1 detectable marker 84+/-6.6% vs 57.1+/-10.5% (p=0.023). When focusing on the 40 patients with intermediate cytogenetics, persistence of 2 or more markers was associated with lower LFS (57+/-11.8% vs 19.4+/-10.5 p=0.0048) and with a trend to lower OS (85+/-8% vs 61+/-11.9% p=0.07). Similar results were observed when restricting the analyses to the 42 prospectively included patients (At 2 years: LFS 73+/-10% vs 24+/-10%, p=0.0026 and OS 90.2+/-6.6% vs 62.8+/-11.5%, p=0.036). In 50 patients with 3 or more identified events, the persistence of 3 or more markers after one course was associated with a very high risk of relapse (DFS 23.5+/-10.3 % vs 75.8 +/-7.5% at one year, p Conclusion Our study shows the high prognostic value of a personalized multi-target clono-specific MRD evaluation that can be used in nearly all AML patients. Detection of two or more events in more than 0.4% cells after one course is associated with lower survival, in particular in intermediate cytogenetic patients. Larger studies are needed to confirm the results and to evaluate if this strategy could be useful to guide treatment decisions. Disclosures No relevant conflicts of interest to declare.
- Published
- 2016
36. New insights for pelvic radiation disease treatment: Multipotent stromal cell is a promise mainstay treatment for the restoration of abdominopelvic severe chronic damages induced by radiotherapy
- Author
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Luc Douay, Jan Voswinkel, Marc Benderitter, Sabine François, and Alain Chapel
- Subjects
Oncology ,medicine.medical_specialty ,Pathology ,Histology ,Stromal cell ,business.industry ,medicine.medical_treatment ,Mesenchymal stem cell ,Cell Biology ,Frontier ,Cell therapy ,Clinical trial ,Radiation therapy ,Gastrointestinal disorder ,Internal medicine ,Genetics ,Medicine ,Stem cell ,business ,Adverse effect ,Molecular Biology ,Genetics (clinical) - Abstract
Radiotherapy may induce irreversible damage on healthy tissues surrounding the tumor. It has been reported that the majority of patients receiving pelvic radiation therapy show early or late tissue reactions of graded severity as radiotherapy affects not only the targeted tumor cells but also the surrounding healthy tissues. The late adverse effects of pelvic radiotherapy concern 5% to 10% of them, which could be life threatening. However, a clear medical consensus concerning the clinical management of such healthy tissue sequelae does not exist. Although no pharmacologic interventions have yet been proven to efficiently mitigate radiotherapy severe side effects, few preclinical researches show the potential of combined and sequential pharmacological treatments to prevent the onset of tissue damage. Our group has demonstrated in preclinical animal models that systemic mesenchymal stromal cell (MSC) injection is a promising approach for the medical management of gastrointestinal disorder after irradiation. We have shown that MSCs migrate to damaged tissues and restore gut functions after irradiation. We carefully studied side effects of stem cell injection for further application in patients. We have shown that clinical status of four patients suffering from severe pelvic side effects resulting from an over-dosage was improved following MSC injection in a compationnal situation.
- Published
- 2013
37. In vitro and in vivo evidence for the long-term multilineage (myeloid, B, NK, and T) reconstitution capacity of ex vivo expanded human CD34+ cord blood cells
- Author
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Xiaxin Li, Monique Titeux, François Leteurtre, Marie-Catherine Giarratana, Luc Douay, Ladan Kobari, Laure Coulombel, Françoise Pflumio, and Brigitte Izac
- Subjects
Cancer Research ,Myeloid ,T-Lymphocytes ,CD34 ,Antigens, CD34 ,Stem cell factor ,Mice, SCID ,Biology ,Mice ,Interleukin 21 ,Mice, Inbred NOD ,Granulocyte Colony-Stimulating Factor ,Genetics ,medicine ,Animals ,Humans ,Lymphocytes ,Lymphopoiesis ,Molecular Biology ,Cells, Cultured ,B-Lymphocytes ,Stem Cell Factor ,Graft Survival ,Hematopoietic Stem Cell Transplantation ,Membrane Proteins ,Cell Differentiation ,Cell Biology ,Hematology ,Fetal Blood ,Hematopoietic Stem Cells ,Hematopoiesis ,Killer Cells, Natural ,Haematopoiesis ,medicine.anatomical_structure ,Thrombopoietin ,Immunology ,Cancer research ,Bone marrow ,Stem cell ,Granulocytes - Abstract
The aim of the present report is to describe clinically relevant culture conditions that support the expansion of primitive hematopoietic progenitors/stem cells, with maintenance of their hematopoietic potential as assessed by in vitro assays and the NOD-SCID in vivo repopulating capacity.CD34(+) cord blood (CB) cells were cultured in serum-free medium containing stem cell factor, Flt3 ligand, megakaryocyte growth and development factor, and granulocyte colony-stimulating factor. After 14 days, the primitive functions of expanded and nonexpanded cells were determined in vitro using clonogenic cell (colony-forming cells, long-term culture initiating cell [LTC-IC], and extended [E]-LTC-IC) and lymphopoiesis assays (NK, B, and T) and in vivo by evaluating long-term engraftment of the bone marrow of NOD-SCID mice. The proliferative potential of these cells also was assessed by determining their telomere length and telomerase activity. Levels of expansion were up to 1,613-fold for total cells, 278-fold for colony-forming unit granulocyte-macrophage, 47-fold for LTC-IC, and 21-fold for E-LTC-IC. Lymphoid B-, NK, and T-progenitors could be detected. When the expanded populations were transplanted into NOD-SCID mice, they were able to generate myeloid progenitors and lymphoid cells for 5 months. These primitive progenitors engrafted the NOD-SCID bone marrow, which contained LTC-IC at the same frequency as that of control transplanted mice, with conservation of their clonogenic capacity. Moreover, human CD34(+)CDl9(-) cells sorted from the engrafted marrow were able to generate CD19(+) B-cells, CD56(+)CD3(-) NK cells, and CD4(+)CD8(+)alphabetaTCR(+) T-cells in specific cultures. Our expansion protocol also maintained the telomere length in CD34(+) cells, due to an 8.8-fold increase in telomerase activity over 2 weeks of culture. These experiments provide strong evidence that expanded CD34(+) CB cells retain their ability to support long-term hematopoiesis, as shown by their engraftment in the NOD-SCID model, and to undergo multilineage differentiation along all myeloid and the B-, NK, and T-lymphoid pathways. The expansion protocol described here appears to maintain the hematopoietic potential of CD34(+) CB cells, which suggests its relevance for clinical applications.
- Published
- 2000
38. Ex vivo expansion of CD34-positive peripheral blood progenitor cells from patients with non-Hodgkin's lymphoma: no evidence of concomitant expansion of contaminating bcl2/JH-positive lymphoma cells
- Author
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Luc Douay, François M. Lemoine, H Firat, G Andreu, Ming Yao, Marc Lopez, Norbert-Claude Gorin, Stéphane Bouchet, and L. Fouillard
- Subjects
Adult ,Male ,Pathology ,medicine.medical_specialty ,Cell Culture Techniques ,CD34 ,Antigens, CD34 ,Stem cell factor ,Cell Separation ,Biology ,Transplantation, Autologous ,Immunophenotyping ,hemic and lymphatic diseases ,Biomarkers, Tumor ,medicine ,Humans ,Progenitor cell ,Gene Rearrangement ,Transplantation ,Lymphoma, Non-Hodgkin ,Membrane Proteins ,Hematopoietic stem cell ,Hematology ,Gene rearrangement ,Middle Aged ,Hematopoietic Stem Cells ,Molecular biology ,Haematopoiesis ,medicine.anatomical_structure ,Proto-Oncogene Proteins c-bcl-2 ,Immunoglobulin Joining Region ,Female ,Stem cell ,Cell Division - Abstract
The aim of the present study was to evaluate the capacity to expand of hematopoietic stem cell (HSC) samples from eight patients with NHL, and to follow in parallel the fate of tumor cells in four of eight samples still containing bcl2/JH+ tumor cells after CD34+ or CD19-/20-/34+ cell selection. The presence of bcl2/JH+ cells was also investigated after expansion in four of eight samples, two of which were bcl2/JH at harvesting and two which were initially bcl2/JH+ but became bcl2/JH (below the level of PCR detection) after cell selection, to assess a possible reappearance of occult tumor cells after expansion culture. We used culture conditions that we previously had established to allow high level expansion of normal precursors, progenitors and LTC-ICs. In this study, particular attention was given to the role of Flt3-ligand, known to favor the growth of B cells. The expansion conditions were: 1.5 x 10(3) cells/ml in serum-free medium containing stem cell factor (SCF), interleukin-3 (IL-3), IL-6, granulocyte-stimulating factor (G-CSF), erythropoietin (Epo) +/- Flt3-ligand (Flt3-L) for 10 days. After culture, total cells, CFU-GMs, BFU-Es and LTC-ICs were expanded to a mean of 833-, 6.6-, 4.6-, and 1.8-fold, respectively with the cocktail of cytokines not including Flt3-L. When Flt3-L was added, the mean expansion values were 1095-, 31-, 15- and three-fold, respectively. Residual bcl2/JH+ cells present in four of eight samples before expansion were not detected after expansion. Similarly, no tumor cells reappeared after expansion of the two samples which had become negative after selection, as well as in the two samples which were bcl2/JH- at harvesting. These results suggest first that ex vivo expansion of hematopoietic stem cells in patients with non-Hodgkin's lymphoma is feasible without incurring the parallel risk of amplifying tumor cells; second, that Flt3-L did not stimulate the growth of tumor cells while it clearly favored the growth of normal progenitors.
- Published
- 2000
39. Importance of marrow dose on posttransplant outcome in acute leukemia
- Author
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L. Fouillard, Christine Perot, Jean-Pierre Jouet, Francis Bauters, Luc Douay, J. P. Laporte, Jacqueline Van Den Akker, Manuel Lopez, Norbert-Claude Gorin, Albert Najman, Myriam Labopin, Françoise Isnard, S Lesage, and Nassima Bellal
- Subjects
Cancer Research ,medicine.medical_specialty ,medicine.medical_treatment ,Population ,Gastroenterology ,law.invention ,chemistry.chemical_compound ,Randomized controlled trial ,Mafosfamide ,law ,Internal medicine ,Acute lymphocytic leukemia ,Genetics ,medicine ,education ,Molecular Biology ,Acute leukemia ,education.field_of_study ,business.industry ,Cytogenetics ,Cell Biology ,Hematology ,medicine.disease ,Autotransplantation ,Surgery ,chemistry ,Stem cell ,business - Abstract
Several prospective randomized trials in acute myelocytic leukemia (AML) documented a lower relapse rate with autologous bone marrow transplantation (ABMT) than with conventional chemotherapy. However, they also identified some transplant difficulties, such as failure to collect sufficient numbers of stem cells, slow kinetics of engraftment, and a high transplant-related mortality that diminished or negated positive impact on overall survival. Data for ABMT are inconclusive in acute lymphocytic leukemia (ALL) in adults. We retrospectively analyzed patients with acute leukemia autografted with marrow purged with mafosfamide after January 1983 in our institution. The population comprised 229 consecutive patients; 165 with AML [123 in first remission (CR1), 32 in second remission (CR2)]; 61 with ALL (46 in CR1, 4 in CR2); and 3 with undifferentiated acute leukemia. All patients were autografted with marrow purged with mafosfamide. Mafosfamide was given at a constant dose of 50 μg/mL in 103 and adjusted individually to produce a CFU-GM LD 95 (5% residual CFU-GM post purging) in 126. The outcome was analyzed for correlation with patient characteristics, the disease including cytogenetics, and the graft itself. Prognostic factors identified by multivariate analysis were used to derive a prognostic classification. Patients receiving higher doses of marrow submitted to purging (>5.46 × 10 4 CFU-GM/kg) experienced a lower treatment-related mortality (RR = 0.11, p=0.005) and a higher leukemia-free (RR = 0.5, p=0.005) and overall survival (RR = 0.4, p=0.001). Patients receiving 5.46 × 10 4 CFU-GM/kg and doses actually infused post purging of ≤0.02 × 10 4 /kg had a treatment-related mortality of only 2 ± 2%, a leukemia-free survival of 70%, and an overall survival of 77 ± 7% at 10 years. In this study of autotransplantation for acute leukemia using mafosfamide-purged marrow, the stem cell dose used for purging and the intensity of purging were the most important factors predicting outcome.
- Published
- 1999
40. Les développements de la thérapie cellulaire de l'an 2000
- Author
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Luc Douay
- Subjects
Biochemistry (medical) ,Clinical Biochemistry ,Hematology - Abstract
Resume En trente ans, le monde hematologique est passe du concept de greffe de moelle a celui de greffe de cellules souches hematopoietiques (CSH) sanguines ou placentaires, du concept de l'allogreffe a celui de l'autogreffe, du greffon non manipule a l'hyperselection, de la therapie cellulaire hematopoietique a l'immunotherapie. Les indications de ces greffes se sont precisees dans le domaine des maladies malignes et s'etendent maintenant aux pathologies auto-immunes. Une meilleure connaissance de la CSH permet un controle de sa proliferation et de sa differenciation, ouvrant le vaste champ de l'expansion ex vivo des differents compartiments hematopoietiques. Depuis peu, de nouvelles cellules souches sont mises en evidence, montrant ainsi qu'une cellule differenciee garde ses potentialites totipotentes: une cellule nerveuse peut en effet se differencier en CSH qui peut donner ellememe naissance a l'hematopoiese, a des cellules mesenchymateuses ou a des hepatocytes. De nouveaux outils se developpent: cellules embryonnaires humaines, biomateriaux, materiaux fonctionnalises, genie tissulaire, ouvrant la voie a l'ingenierie cellulaire des annees 2000.
- Published
- 1999
41. Cell culture bags allow a large extent of ex vivo expansion of LTC-IC and functional mature cells which can subsequently be frozen: interest for large-scale clinical applications
- Author
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Marcelo F. Lopez, M-C Giarratana, N. C. Gorin, Ladan Kobari, Tma Neildez Nguyen, Stéphane Bouchet, Luc Douay, H Firat, and Dominique Thierry
- Subjects
Cryopreservation ,Transplantation ,Pathology ,medicine.medical_specialty ,Time Factors ,Granulocytic cells ,Cell Culture Techniques ,Hematopoietic Stem Cell Transplantation ,CD34 ,Hematology ,Biology ,Hematopoietic Stem Cells ,Hematopoietic Stem Cell Mobilization ,Andrology ,Haematopoiesis ,Blood Preservation ,Nucleated cell ,Cell culture ,medicine ,Humans ,Ex vivo expansion ,Progenitor cell ,Stem cell - Abstract
The aim of this study was to evaluate the ex vivo expansion of normal CD34 + cells in gas-permeable polypropylene bags suitable for clinical use. Cells were cultured for 14 days in serum-free medium supplemented with SCF, IL3, IL6, FLT3-1, G-CSF ± MGDF or Epo. The bags supported the expansion of hematopoietic cells in a similar manner to small scale well or flask systems, allowing mean expansions of up to 2193-fold for total nucleated cells, 140-fold for CFU-GM and 66-fold for LTC-IC. Increasing the initial cell concentration from 5 x 10 3 to 1 x 10 5 CD34 + cells/ml induced the production of granulocytic cells with terminal differentiation while simultaneously decreasing the overall extent of expansion of the white blood cells produced. We tested the phagocytic activity and oxidative metabolism of the white blood cells produced. The percentage of phagocytic cells was 39 ± 0.5% in expanded cultures derived from fractions initiated at 5 x 10 3 , 104 or 10 5 cells/ml and 45 ± 6% in cultured cells obtained from starting fractions containing 5 x 10 4 cells/ml, as compared to 58 ± 4% in normal controls. A study of the potential for oxygen-dependent microbe killing showed that the expanded cells produced H 2 O 2 , although in lesser quantities than control cells. We subsequently investigated the possibility of freezing expanded cells. Total cell recovery after thawing was 45 ± 4%, while recoveries of progenitors and stem cells ranged from 65 to 90%, without any influence of the initial cell concentration. This new approach could be of major interest for clinical practice, as it would allow evaluation of the quality of a graft prior to its infusion and employs experimental conditions which meet the criteria for potential clinical use.
- Published
- 1998
42. Negative selection and protection of normal progenitor cells for autografting
- Author
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Luc Douay
- Subjects
Male ,Pathology ,medicine.medical_specialty ,medicine.medical_treatment ,Bone Marrow Cells ,Transplantation, Autologous ,Humans ,Medicine ,Progenitor cell ,Cytotoxicity ,Bone Marrow Transplantation ,Transplantation ,Chemotherapy ,business.industry ,Bone Marrow Purging ,Hematology ,Amifostine ,Radiation therapy ,medicine.anatomical_structure ,Toxicity ,Cancer research ,Female ,Bone marrow ,business ,Ex vivo ,medicine.drug - Abstract
Autologous bone marrow transplantation (ABMT) after high-dose chemotherapy is recognized as a curative approach to treating hematologic malignancies and some invasive solid tumors. However, tumor cells present in the bone marrow at the time of harvesting are a potential cause for relapse. Ex vivo marrow purging with very high doses of cytotoxic agents has been introduced in an attempt to remove neoplastic cells contaminating the autograft. The procedure, however, has been limited by its high toxicity to normal bone marrow progenitor cells. In their purging procedures, investigators have used agents such as amifostine, originally developed to protect against the effects of radiation and chemotherapy. In this article, the appropriateness of protecting normal cells with amifostine during various purging procedures will be reviewed.
- Published
- 1998
43. Autologous stem-cell transplantation for non-Hodgkin's lymphomas: the role of graft purging and radiotherapy posttransplantation--results of a retrospective analysis on 120 patients autografted in a single institution
- Author
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M.P. Noel-Walter, P Zunic, Marc Lopez, Yazid Belkacemi, F. Isnard, Pierre Morel, M Aoudjhane, Stachowiak J, S. Lesage, M. Labopin, Luc Douay, N. Cheron, G Andreu, J P Laporte, M P Lemonnier, Albert Najman, J P Jouet, Pierre Fenaux, Francis Bauters, Loic Fouillard, and Norbert-Claude Gorin
- Subjects
Adult ,Male ,Melphalan ,Cancer Research ,medicine.medical_specialty ,medicine.medical_treatment ,Hematopoietic stem cell transplantation ,Disease-Free Survival ,Autologous stem-cell transplantation ,Recurrence ,Antineoplastic Combined Chemotherapy Protocols ,medicine ,Humans ,Cyclophosphamide ,Etoposide ,Retrospective Studies ,Chemotherapy ,Carmustine ,business.industry ,Lymphoma, Non-Hodgkin ,Bone Marrow Purging ,Remission Induction ,Cytarabine ,Hematopoietic Stem Cell Transplantation ,Middle Aged ,Prognosis ,Combined Modality Therapy ,Bone marrow purging ,Surgery ,Oncology ,Female ,business ,medicine.drug - Abstract
PURPOSE To analyze retrospectively survival and prognostic factors of patients with non-Hodgkin's lymphoma (NHL) autografted from 1979 to 1995 in a single institution. PATIENTS AND METHODS A total of 120 patients, 64 with aggressive and 56 with low-grade NHL, were autografted. The carmustine (BCNU), etoposide, cytarabine, and melphalan (BEAM) regimen was used in 104. The autograft was marrow in 101 patients. Marrow was purged in vitro by mafosfamide for 63 patients (adjusted dose [AD] in 32; unique dose [UD] in 31); 27 patients received a CD34+-selected graft. Following intensification, 45 patients received additional radiotherapy on previous sites of involvement. RESULTS Outcome at 5 years for patients transplanted with low-grade NHL in first complete remission (CR1), in first partial remission (PR1), and in second complete remission (CR2) or beyond showed an event-free survival (EFS) of 75% +/- 12%, 46% +/- 18%, and 57% +/- 24%, a relapse incidence (RI) of 21% +/- 12%, 49% +/- 19%, and 43% +/- 25%, and a transplant-related mortality (TRM) of 5% +/- 5%, 10% +/- 7%, and 0%, respectively. For patients with aggressive NHL transplanted in CR1, in PR1, in CR2 or beyond, and in resistant relapse or in primary refractory disease, the EFS was of 73% +/- 9%, 58% +/- 19%, 29% +/- 16%, and 10% +/- 9%, the RI 22% +/- 9%, 14% +/- 9%, 77% +/- 18%, and 66% +/- 20%, and the TRM 6% +/- 6%, 32% +/- 21%, 11% +/- 10%, and 71% +/- 22%, respectively. In patients autografted upfront in first remission, additional radiotherapy was associated with a higher EFS, in univariate (P = .03) and multivariate analysis (P = .02, relative risk [RR] = .021). The role of graft purging with mafosfamide on the outcome reflected by the dose of colony-forming unit-granulocyte-macrophage (CFU-GM) per kilogram infused postpurging was assessed by univariate analysis: patients in first remission who received lower doses of CFU-GM had a lower RI and a higher EFS. CONCLUSION This retrospective analysis suggests that marrow purging and posttransplant radiotherapy improve the outcome of patients with NHL autografted in first remission.
- Published
- 1998
44. Fifteen years of preclinical and clinical experiences about biotherapy treatment of lesions induced by accidental irradiation and radiotherapy
- Author
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Sabine François, Marc Benderitter, Luc Douay, Jan Voswinkel, and Alain Chapel
- Subjects
medicine.medical_specialty ,Severe combined immunodeficiency ,Histology ,business.industry ,medicine.medical_treatment ,Mesenchymal stem cell ,Cell Biology ,Total body irradiation ,medicine.disease ,Frontier ,Surgery ,Radiation therapy ,Therapeutic approach ,Genetics ,medicine ,Radiodermatitis ,Stem cell ,business ,Adverse effect ,Molecular Biology ,Genetics (clinical) - Abstract
High dose radiation exposures involving medical treatments or accidental irradiation may lead to extended damage to the irradiated tissue. Alleviation or even eradication of irradiation induced adverse events is therefore crucial. Because developments in cell therapy have brought some hope for the treatment of tissues damages induced by irradiation, the Institute for Radiation and Nuclear Safety contributed to establish the clinical guidelines for the management of accidentally irradiated victims and to provide the best supportive care to patients all over the world. In the past 15 years, we contributed to develop and test cell therapy for protection against radiation side effects in several animal models, and we proposed mechanisms to explain the benefit brought by this new therapeutic approach. We established the proof of concept that mesenchymal stem cells (MSCs) migrate to damaged tissues in the nonobese diabetic/severe combined immunodeficiency immunotolerant mice model and in non-human primate after radiation exposure. We showed that the intravenous injection of MSCs sustains hematopoiesis after total body irradiation, improves wound healing after radiodermatitis and protects gut function from irradiation damages. Thanks to a tight collaboration with clinicians from several French hospitals, we report successful treatments of therapeutic/accidental radiation damages in several victims with MSC infusions for hematopoiesis correction, radio-induced burns, gastrointestinal disorders and protection homeostatic functions of gut management after radio-therapy.
- Published
- 2013
45. Large-scale production of red blood cells from stem cells: what are the technical challenges ahead?
- Author
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Marie-Catherine Giarratana, Luc Douay, and Guillaume F. Rousseau
- Subjects
Blood transfusion ,Erythrocytes ,business.industry ,Cost effectiveness ,Scale (chemistry) ,medicine.medical_treatment ,Cell Culture Techniques ,General Medicine ,Hematopoietic Stem Cells ,Applied Microbiology and Biotechnology ,Human being ,Biotechnology ,Successful injection ,Bioreactors ,Risk analysis (engineering) ,New product development ,medicine ,Molecular Medicine ,Production (economics) ,Animals ,Humans ,Blood Transfusion ,Stem cell ,business - Abstract
Blood-transfusion centers regularly face the challenge of donor blood shortages, especially for rare blood groups. The possibility of producing universal red blood cells from stem cells industrially has become a possible alternative since the successful injection of blood generated in vitro into a human being in 2011. Although there remains many biological and regulatory issues concerning the efficacy and safety of this new product, the major challenge today for future clinical applications is switching from the current limited 2-dimensional production techniques to large-scale 3-dimensional bioreactors. In addition to requiring technological breakthroughs, the whole process also has to become at least five-fold more cost-efficient to match the current prices of high-quality blood products. The current review sums up the main biological advances of the past decade, outlines the key biotechnological challenges for the large-scale cost-effective production of red blood cells, proposes solutions based on strategies used in the bioindustry and presents the state-of-the-art of large-scale blood production.
- Published
- 2013
46. In vitro generated Rh(null) red cells recapitulate the in vivo deficiency: a model for rare blood group phenotypes and erythroid membrane disorders
- Author
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Jean-Pierre Cartron, Anne Dubart-Kupperschmitt, Marie Cambot, Christelle Mazurier, Véronique Picard, Denis Clay, Julien Picot, Pierre Ripoche, Luc Douay, Nicolas Hebert, and Florence Canoui-Poitrine
- Subjects
Reticulocytes ,Cellular differentiation ,Porphyria, Erythropoietic ,CD47 Antigen ,Anemia, Hemolytic, Congenital ,Cell Line ,Erythroid Cells ,Pregnancy ,Humans ,Progenitor cell ,RNA, Small Interfering ,Cells, Cultured ,Genetics ,Erythroid Precursor Cells ,Fetal Stem Cells ,Membrane Glycoproteins ,Rh-Hr Blood-Group System ,biology ,CD47 ,Genetic Diseases, Inborn ,Cell Differentiation ,Hematology ,Blood Proteins ,Fetal Blood ,Cell biology ,Adult Stem Cells ,Cell culture ,RHAG ,Membrane biogenesis ,biology.protein ,Female ,RNA Interference ,Anemia, Hypoplastic, Congenital ,Cell Adhesion Molecules ,Biogenesis ,Ex vivo - Abstract
Lentiviral modification combined with ex vivo erythroid differentiation was used to stably inhibit RhAG expression, a critical component of the Rh(rhesus) membrane complex defective in the Rh(null) syndrome. The cultured red cells generated recapitulate the major alterations of native Rh(null) cells regarding antigen expression, membrane deformability, and gas transport function, providing the proof of principle for their use as model of Rh(null) syndrome and to investigate Rh complex biogenesis in human primary erythroid cells. Using this model, we were able to reveal for the first time that RhAG extinction alone is sufficient to explain ICAM-4 and CD47 loss observed on native Rh(null) RBCs. Together with the effects of RhAG forced expression in Rh(null) progenitors, this strongly strengthens the hypothesis that RhAG is critical to Rh complex formation. The strategy is also promising for diagnosis purpose in order to overcome the supply from rare blood donors and is applicable to other erythroid defects and rare phenotypes, providing models to dissect membrane biogenesis of multicomplex proteins in erythroid cells, with potential clinical applications in transfusion medicine.
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- 2013
47. Caspase-3 Is Involved in the Signalling in Erythroid Differentiation by Targeting Late Progenitors
- Author
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Daniela Boehm, Christelle Mazurier, Dhouha Darghouth, Marie-Catherine Giarratana, Luc Douay, Laurence Harmand, Anne-Marie Faussat, HAL UPMC, Gestionnaire, Différenciation et prolifération des cellules souches adultes. application à la thérapie cellulaire hématopoiétique, Université Pierre et Marie Curie - Paris 6 (UPMC)-Institut National de la Santé et de la Recherche Médicale (INSERM), Centre de Recherche Saint-Antoine (UMRS893), Etablissement Français du Sang (EFS), EFS, Cytométrie et Imagerie Saint-Antoine (CISA), Unité Mixte de Service d'Imagerie et de Cytométrie (UMS LUMIC), Université Pierre et Marie Curie - Paris 6 (UPMC)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Pierre et Marie Curie - Paris 6 (UPMC)-Institut National de la Santé et de la Recherche Médicale (INSERM), CHU Trousseau [APHP], and Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Sorbonne Université (SU)
- Subjects
[SDV.MHEP.HEM] Life Sciences [q-bio]/Human health and pathology/Hematology ,Myeloid ,Cell division ,Cellular differentiation ,Red Cells ,lcsh:Medicine ,Apoptosis ,Biochemistry ,0302 clinical medicine ,hemic and lymphatic diseases ,Molecular Cell Biology ,lcsh:Science ,Erythroid Precursor Cells ,Cells, Cultured ,0303 health sciences ,Multidisciplinary ,Caspase 3 ,Stem Cells ,Cell Differentiation ,[SDV.MHEP.HEM]Life Sciences [q-bio]/Human health and pathology/Hematology ,Hematology ,Cell cycle ,Caspase Inhibitors ,Cell biology ,Enzymes ,Adult Stem Cells ,medicine.anatomical_structure ,030220 oncology & carcinogenesis ,Medicine ,Cell Division ,Signal Transduction ,Research Article ,G2 Phase ,Biology ,Cell Growth ,03 medical and health sciences ,medicine ,[SDV.BBM] Life Sciences [q-bio]/Biochemistry, Molecular Biology ,Humans ,Cell Lineage ,[SDV.BBM]Life Sciences [q-bio]/Biochemistry, Molecular Biology ,Progenitor cell ,030304 developmental biology ,Cell Proliferation ,Cell growth ,lcsh:R ,Hematopoietic Stem Cells ,Hematopoiesis ,Enzyme Activation ,lcsh:Q ,Developmental Biology - Abstract
International audience; A role for caspase activation in erythroid differentiation has been established, yet its precise mode of action remains elusive. A drawback of all previous investigations on caspase activation in ex vivo erythroid differentiation is the lack of an in vitro model producing full enucleation of erythroid cells. Using a culture system which renders nearly 100% enucleated red cells from human CD34 + cells, we investigated the role of active caspase-3 in erythropoiesis. Profound effects of caspase-3 inhibition were found on erythroid cell growth and differentiation when inhibitors were added to CD34 + cells at the start of the culture and showed dose-response to the concentration of inhibitor employed. Enucleation was only reduced as a function of the reduced maturity of the culture and the increased cell death of mature cells while the majority of cells retained their ability to extrude their nuclei. Cell cycle analysis after caspase-3 inhibition showed caspase-3 to play a critical role in cell proliferation and highlighted a novel function of this protease in erythroid differentiation, i.e. its contribution to cell cycle regulation at the mitotic phase. While the effect of caspase-3 inhibitor treatment on CD34 + derived cells was not specific to the erythroid lineage, showing a similar reduction of cell expansion in myeloid cultures, the mechanism of action in both lineages appeared to be distinct with a strong induction of apoptosis causing the decreased yield of myeloid cells. Using a series of colony-forming assays we were able to pinpoint the stage at which cells were most sensitive to caspase-3 inhibition and found activated caspase-3 to play a signalling role in erythroid differentiation by targeting mature BFU-E and CFU-E but not early BFU-E.
- Published
- 2013
48. Cord-Blood Transplantation from an Unrelated Donor in an Adult with Chronic Myelogenous Leukemia
- Author
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Luc Douay, Marie-France Portnoi, S Lesage, Jean-Philippe Laporte, Albert Najman, Norbert-Claude Gorin, Manuel Lopez, Pablo Rubinstein, and Véronique Barbu
- Subjects
Adult ,Male ,business.industry ,Histocompatibility Testing ,medicine.medical_treatment ,Hematopoietic Stem Cell Transplantation ,Infant, Newborn ,General Medicine ,Hematopoietic stem cell transplantation ,Fetal Blood ,medicine.disease ,Umbilical cord ,Transplantation ,Leukemia ,Myelogenous ,medicine.anatomical_structure ,Leukemia, Myelogenous, Chronic, BCR-ABL Positive ,Cord blood ,Immunology ,medicine ,Humans ,Female ,Stem cell ,business ,Chronic myelogenous leukemia - Abstract
To circumvent the problems inherent in allogeneic bone marrow transplantation, allogeneic cord blood has been studied as an alternative source of hematopoietic stem cells. Preliminary results with cord blood from an HLA-matched sibling are encouraging, but the experience with this procedure is limited.1–5 Although some 200 transplantations of cord blood have been performed in children,6,7 few have been performed in adults. We describe an adult with chronic myelogenous leukemia who underwent the transplantation of cord-blood stem cells successfully. Case Report A 26-year-old woman was found to have chronic myelogenous leukemia in July 1990. All the leukemic cells had . . .
- Published
- 1996
49. A new congenital dysmegakaryopoietic thrombocytopenia (Paris-Trousseau) associated with giant platelet alpha-granules and chromosome 11 deletion at 11q23 [see comments]
- Author
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Rémi Favier, Josette Guichard, Luc Douay, D Cherif, Najet Debili, William Vainchenker, Roland Berger, and J Breton-Gorius
- Subjects
Pathology ,medicine.medical_specialty ,biology ,Platelet disorder ,Immunology ,Paris-Trousseau syndrome ,GATA1 ,Cell Biology ,Hematology ,medicine.disease ,Biochemistry ,Chromosome aberration ,Von Willebrand factor ,biology.protein ,medicine ,Platelet ,Thrombopoiesis ,Jacobsen syndrome - Abstract
This study characterizes a new congenital thrombocytopenia with mild hemorrhagic tendency occurring in a woman and her child with the following features. We found a deletion of the distal part of one chromosome 11 [del(11)q23.3-->qter] that was detected by cytogenetic analysis and confirmed by chromosome painting in the two patients and also an increased number of bone marrow megakaryocytes (MKs), including numerous micromegakaryocytes (mMKs) associated with a normal platelet life span. A normal number of MK colonies in culture was observed with one third of them containing a few large MKs; however, these were always associated with mMKs identified by immunologic staining. A massive cell lysis was observed at the end of the maturation. Fifteen percent of the platelets in the peripheral blood showed giant alpha-granules resulting from the fusion of alpha-granules. These giant granules, which appeared in red on giemsa stain, had a mean diameter of 1.5 microns and showed all markers (detected at electron microscopy by immunogold method) of matrix and alpha-granule membrane, ie, von Willebrand factor, fibrinogen, CD41, CD62P (P-selectin); however, they differed from lysosomes because acid phosphatases were not present. These giant alpha-granules were unable to release their contents after stimulation by thrombin, in contrast to platelets with normal morphology. Abnormalities in bone marrow MK maturation that were detected at the electron microscopic level and that led to lysis of numerous MKs were responsible for thrombocytopenia and were similar in both patients. MK abnormalities are probably the consequence of the chromosome aberration. ETS 1 and FLI, two proto-oncogenes that appear to be essential with GATA1 for the normal expression of MK-specific genes, map to 11q23-q24 and are, thus, deleted in this thrombocytopenia. In conclusion, the association of all these abnormalities constitutes a new familial platelet disorder and may present a valuable model for exploring the role of some genes involved in the regulation of thrombopoiesis.
- Published
- 1995
50. Vers une standardisation de la culture des progéniteurs hématopoïétiques CFU-GM appliquée à la greffe de cellules souches hématopoïétiques en France
- Author
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S. Bouchet, N. C. Gorin, Bardinet D, V. Texier, Manuel Lopez, and Luc Douay
- Subjects
Hematopoietic cell ,Biochemistry (medical) ,Clinical Biochemistry ,Hematology ,Biology ,Molecular biology - Abstract
Resume Le contenu en progeniteurs granulo-macrophagiques (CFU-GM) de 130 echantillons de moelle osseuse et de 105 echantillons de sang a ete etudie en parallele par 2 techniques, l'une en agar avec du milieu placentaire, l'autre en methyl-cellulose supplementee avec des facteurs de croissance recombinants (G-CSF, GM-CSF, IL3 et erythropoietine). Les echantillons de moelle ont ete testes frais (130), apres traitement par mafosfamide (31) et apres congelation-decongelation (33). Les resultats montrent que le nombre de CFU-GM detectees en methyl-cellulose est pour tous les types d'echantillons correle avec celui qui est detecte en agar (p = 0,0001) par une formule de type « log CFU-GM methyl = a log CFU-GM agar + b . Sur la base des correlations trouvees, les doses seuils/kg de CFU-GM detectees en methyl-cellulose necessaires dans notre experience pour constituer un greffon autologue de moelle osseuse et de sang ont ete recalculees a partir des doses seuils que nous avions precedemment determinees en agar (respectivement 104/kg et 5 × 104/kg) : ces doses sont respectivement 2,3 × 104/kg pour la moelle osseuse non traitee, et 2,1 × 105/kg pour les cellules sanguines.
- Published
- 1995
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