Nancytomique Wuhan Juin 2013

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La présentation sur Nancytomique faite lors de la réunion scientifique sur le Laboratoire sans murs organisée avec le consulat de France dans le cadre de la Filière Médicale Francophone Nancy-Wuhan à la Faculté de Médecine de Wuhan et à l'Hôpital Zhongnan.

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Nancytomique Wuhan Juin 2013

  1. 1. NANCYTOMIQUE • A unique platform dedicated to clinical and translational research • Focusing on rare cell events – New cellular biomarkers for personalized medicine • In Oncology • In Cardiovascular and immunological diseases
  2. 2. Equipments, Methods and Skills • Technical and Medical competences – Flow cytometry, cell imaging, immunology, hématology, cell biology – Cancerology, Cardiovascular, Internal medicine • 3 complementary Innovative Devices – Multiparametric Flow cytometer (3 lasers, 10 colours) Navios (Beckman Coulter) – Cell Search VERIDEX – ImageStream X AMNIS • Making available for Clinical Research – Multiparametric immunophenotyping – Immunomagnetic enrichment – Multispectral imaging on slide and in flow
  3. 3. MANPOWER • GC FAURE, (MC Béné), M de CARVALHO • Engineer • PhD Students – CHEN Min – CAI Huili – TU Qian • M2 Research – De Kayser Manon – WU Xanglei • Open to other labs in Nancy (CRAN CNRS UMR7039, Fdération de Recherche CNRS) – DING Haixia – WANG YingYing – MASTRONICOLA R, ZUILY S
  4. 4. Filière médicale francophone 法国医疗领域 depuis la classe 2001/2 在医学院总理事会的摩泽尔总理事会的摩泽尔
  5. 5. TOPICS • Onco-Haematology: – Minimal Residual disease in ALL, AML, CLL – Myeloma phenotyping and thrombosis (MCB + VLC) • Solid Oncology – CTCs (Minimal Residual disease: a one century challenge – CSFTCs and ETCs • Immunology: transplantation, tumour – Discrete cells (Tregs, dendritic cells), – Cytokine signatures of cell subpopulations • Cardiovascular diseases: CECs, pECs, microparticles – Vasculitis and Angiogenesis
  6. 6. ONCO-HEMATOLOGY CHEN M, CAI H • From extensive and optimal immunophenotyping in – ALL/AML – CLL • To Minimal residual disease (MRD) and Peripheral Blood Blast decrease • To Evaluation of potential New Therapies (CHEN M) – Thèse UL BIOSE April 8, 2013: Stimulation of CLL cells through TLR-7 and TLR-9 – Chen M et al Leuk & Lymphoma 2012;53:739-742
  7. 7. Multiparametric flow cytometry MC Béné, on behalf of GEIL, GTLLF, GOELAMS Normal Bone marrow Minimal Residual Disease Rare events: detection and quantification in « bermudes » CD34 CD45 CD13 SSC
  8. 8. CLL: Innate immune receptors on B CLL cells, and beyond… • B-CLL cells express TLR-7 and over- express TLR-9 • Extensive study of 40 CLL B-cells stimulated – ODN M362(CpG-C) ligand of TLR-9 – Imiquimod R837, ligand of TLR-7 • Multiparametric flow cytometry for apoptosis, proliferation, immunophenotypic modulation (CD20, CD52) , cytokines (IL-6, IL-8) production, signaling pathways (AKT, STAT, MAPK) • Imiquimod and ODN could be used as an adjuvant immunotherapy strategy for patients with CLL D 0 D 1 D 2 D 3 0 20 40 60 80 100 * * * * * * Time (n=8) S %ofcellslabelled positivelybyTUNEL D 1 D 2 D 3 D 1 D 2 D 3 D 1 D 2 D 3 D 1 D 2 D 3 D 1 D 2 D 3 D 1 D 2 D 3 D 1 D 2 D 3 D 1 D 2 D 3 0 1 2 3 4 * * * * * * * * * ** * * S Time (n=11) RelativeMFI
  9. 9. Solid ONCOLOGY TU Q • CirculatingTumor Cells (CTCs) – STIC DHOS METABREAST (Institut Curie) – Cancéropole Grand Est (Inserm Strasbourg) • CerebroSpinalFluidTCs (Breast, Lung) – DEPOSEIN BMC Clin Pathology 2012 • CSFMelanomaCells – Medical Oncology 2013 • World CTC USA 2012, World CTC Berlin 2013, Lung Cancer 2013 Luxbembourg: Invited speaker, • Cytology 2013 Prize Junior Cytopathologist • EffusionTCs – Tumor Cells quantification in peripheral blood and ascitic fluid from metastatic esophageal cancer submitted %ProbabilityofSurvival 0% 10% 20% 30% 40% 50% 60% 70% 80% 90% 100% Time from Baseline (Months) 0 5 10 15 20 30 35 40 45 5025 Logrank p < 0.0001 Cox Hazards Ratio = 2.4 chi-square = 19.54 (p-value < 0.0001) 21.9 Months 10.9 Months CTC / 7.5mL Median OS in at Baseline N (%) Months (95% C.I.) <5 CTC 89 (50%) 21.9 (20.1 to 28.6) >5 CTC 88 (50%) 10.9 ( 7.0 to 15.2)
  10. 10. LM and « CSFTCs » in Breast Cancer (BMC Clinical Pathology 2012) • Detection • Quantification: 1 to 103-4 • Characterization – Her2-Neu Specificity: no contaminating ependymal cells Sensitivity: detection and quantification in all established LM patients studied compared to cytology High homogeneity (and reproducibility) of images High purity compared to blood samples
  11. 11. Sequential analysis of CSFTCs confirms repetability of numerations with two subgroups (high > 700/mL vs low) BMC Clin Pathol 2012 Sample number 1st 2nd 3rd 4th 5th Tumoralcells/5mLLCR 0,1 1 10 100 1000 10000 100000 DM CTC WA CTC CJ CTC DMB CTC VT CTC PV CTC BE CTC CS CTC HE CMC ST CMC DC CTC
  12. 12. Melanoma CSFMCs Medical Oncology 2013; • CMC kit (J&J, VERIDEX) – CD146 capture, MEL14 , CD45 • Four patients 9 points • Good reproducibility during follow-up • Cell morphology of melanoma cells in CSF far better than in blood
  13. 13. LUNG Cancer CSFTCs + CSFTMs Cytomorphology: cell galleries allow to differentiate SCLC, NSCLC (adenocarcinoma, squamous carcinoma) and detect apoptosis, autophagy... • Numerous CTMs in some NSCLC patients up to 80%, in sequential study
  14. 14. Immunology CHEN M, M de Carvalho, X WU Discrete cells Tregs and Dendritic cell subpopulations Cytokine signatures according to tumoral antigenic load Immunomonitoring for Infectious diseases (HIV, tuberculosis) Transplantation follow-up Chen M et al Clin exp Immunol 2011;163:104-12 Reduced levels of both circulating CD4+CD25+CD127(low/neg) and CD4+CD8(neg) invariant natural killer regulatory T cells in stable heart tranplant recipients Chen M et al 2nd ECI Berlin 2009. B1 cells expressing CXCR3 in cardiac allograft rejection Tumor immunity (Prostate, Colic and ENT Cancer) M2 Project WU X
  15. 15. Phenotypic markers and Cytokine signatures Functional diversity associated with • cell migration to systemic and mucosal lymphoid tissues • proliferation, cytotoxicity •antigenic load Pantaleo et al. Nature Reviews Immunology 6, 417–42
  16. 16. Cardiovascular diseases CAI H, TU Q, DING H • From Endothelial Progenitors to CECs, and microparticles – Number, Morphology, Apoptosis? • Angiogenesis disorders – STB and Rendu-Osler telangiectasia • Inflammation – Vasculitis: SAPL, Wegener – Heart infarction • Myeloma and thrombosis (VLC) • Glioblastoma and exosomes (BF)
  17. 17. CECs: Preliminary results • Number >20/mL • Morphology • Clusters (Wegener) • Multinucleated cells? (Rendu Osler) • Apoptosis (myeloma) • Association with mononuclear cells (Wegener)
  18. 18. Collaborations and projects • France – Nancy (EA4369) CRAN/UMR7039 • Anatomopathologie CAV, Méd Interne, Méd Vasculaire, Urologie, Neuro- oncologie, Pneumologie, MIMOS... – Cancéropole Grand Est (Prostate, Colorectal) – Lille (Centre Oscar Lambret-CSFTCs, CMCs/PHRC) – Paris Institut CURIE (STIC DHOS METABREAST) • International – Estonia (TARTU, GORCE Parrot project) – China (WUHAN...) – USA EPIC: Fluid biopsy and CECs – ScreenCell

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