EHA an overview. Christine Chomienne EHA President.
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1 EHA an overview Christine Chomienne EHA President
2 EHA activities Career development Calls are open now EHA Learning Center Annual congress EHA promotes excellence in research, education and patient care in hematology Scientific meetings Tutorials Haematologica Lobby for 20 for young Membership hematologist hematology
3
4 But what if you cannot attend the EHA congress?
5 EHA s Outreach Program EHA s Online Learning Center
6 EHA s Outreach Program Leading principle: Outreach activities are based on the needs of the hosting country Types of meetings: Tutorials Joint Symposia Best of EHA 2015 Taiwan Turkey Iran Russia Argentina India, Czech Republic
7 EHA Learning Center The official learning portal of EHA Integrates an extensive library of all EHA s peer reviewed educational resources into an easy to use learning platform.
8
9 Leukemic stem cell heterogeneity How to monitor and eradicate HST& EHA joint symposium 2015
10 I have nothing to disclose
11 Tumor Burden Diagnosis Prognosis Patient Management Molecular Relapse Molecular complete remission Treatment and disease monitoring Hokland P, and Ommen H B Blood 2011;117:
12 Tumor Burden Tumor Initiating Cells Niche Niche Niche HSC Treatment and disease monitoring Adapted from Hokland P, and Ommen H B Blood 2011
13 Tumor Burden Tumor Heterogeneity- Evolution Niche Niche Niche HSC Treatment and disease monitoring Adapted from Hokland P, and Ommen H B Blood 2011
14 Initiation of hematopoietic malignancies Plasticity, Evolution and Clonal heterogeneity Niche SC Niche LSC MUTATIONS CONFER STEMNESS CAPACITY Activate stem cell pathways Proliferation Survival Tumor Proliferation Differentiation Programmed cell death X Differentiation
15 Methods to assess and enrich cell populations in Tumor initiating cells Common to Solid Tumors and Leukemia CD44, CXCR4, CD133 Leukemia CD34, CD38, CD123, CD47, CD96,CD49 CD34 + CD38 - CD90 - CD45RA + CD34 + CD38 + CD123 +/lo BAH1CD45RA+ CD32,CD25,CD90,CD117 Spheroid Xenografts Leukemia Progenitor colony assays LT-CIC Xenografts Common to Solid Tumors and Leukemia Common to Solid Tumors and Leukemia Common to Solid Tumors and Leukemia Presence of CSC in negative populations CD38+ or CD34- CD133- SP- Taussig 2008, 2010, Sarry 2011, Eppert 2011, Goardon 2011 Shmelkov 2008, Triel 2009 Adapted International Journal of Cancer 2012
16 Molecular targets and signaling pathways shared by tumor initiating cells ROS NFKB Survival (BCL2) PI3K/mTOR/F OXO Tumor Initiating cell Vasculature 02 Cytokines bfgf,igf,vegf Cell of the Niche
17 Tumor initiating cell gene expression and CR
18 Unravelling mutational profiles Two dimensional analysis of tumor/relapse sample PLOS Computational Biology 2014, C Miller
19 But. Abnormal phenotypic or genotypic markers are not sufficient to predict disease initiation capacity Functional assays are necessary to confirm the presence of disease initiating cells
20 Functional characteristics of disease initiating cells Niche 1) Engraftment 2) Self-renewal Maintenance proliferation clonogenicity initiation of disease Proliferation Survival Tumor X Differentiation
21 Engraftment Primary haml In vivo transfer to an immuno-deficient mouse identical blast morphology and dissemination profiles than initial haml Rate of engraftment Frequency of LSC in a patient s leukemic sample Lapidot T, Nature, 1994 Bonnet D and Dick J, Nature Med, 1997
22 Self renewal Primary AML Serial transplantation
23 Functional assessment of disease initiating cells What can it bring the field? «Engraftment» : a prognostic factor Identification of initiating stem cells in other diseases (lymphoid, MDS, CML) Validation of novel «disease» biomarkers Validation of novel disease drug targeting
24 Overall survival of engrafting and non engrafting AML samples. Non-engrafters N=12 engrafters N=13 Pearce D J et al. Blood 2006;107: Van Rhenen 2005,2007, Ran 2009, Gentiles, 2010, Eppert 2011
25 Functional assessment of disease initiating cells What can it bring the field? «Engraftment» : a prognostic factor Identification of initiating stem cells in other diseases (lymphoid, MDS, CML) Validation of novel «disease» biomarkers Validation of novel disease drug targeting
26 Prediction of the existence of potential relapse initiating clones Flow cytometry Diagnosis Xenograft assays - Disease monitoring in vivo o Relapse Relapse Relapse Xenografts of diagnosis sample
27 Combining Mutational Status and Functional assays Mutation analysis Patient sample analysis Colony assay, LTC-IC in MDS/MPN - Disease definition - Drug assessment in vitro - Disease monitoring in vivo o Flow cytometry Diagnosis Xenograft assays - Disease definition - Drug assessment in vitro - Disease monitoring in vivo Xenografts of diagnosis sample
28 Assessment of MDS-stem cells Impact of combining genetic & functional assays Flow cytometry Stem Cells + Progenitors LTC-IC- Xenografts Exon sequencing on Patient samples Colonies, Single cells Existence of MDS-SC Expansion of progenitors with self renewal Identification of mutation significance Persistance of mutations in CR Identification of «transforming» mutations Serial MDS patients samples at diagnosis and during treatment, in CR or before AML transformation Petter S. Woll ; Cancer Cell, Volume 25, Issue 6, 2014,
29 LSCs in CML express CD26. Identification of a novel biomarker for CML stem cells CD26 (SDF1/CXCR4 pathway) Herrmann H et al. Blood 2014;123:
30 A novel biomarker for CML stem cells (CD26) (2) No disease CML disease Herrmann H et al. Blood 2014;123:
31 A novel biomarker for CML stem cells (CD26) (3) Herrmann H et al. Blood 2014;123:
32 Functional assessment of LSC What can it bring the field? «Engraftment» : a prognostic factor Identification of initiating stem cells in other diseases (lymphoid, MDS, CML) Validation of novel «disease» biomarkers Validation of novel disease drug targeting
33 Means to identify novel approaches to eradicate disease initiating cells 1-Inhibition of homing and/or induction of differentiation - Cytokines: G-CSF, IFN - Antibodies or Antagonists: CXCR4, CD44 Tavor 2004, Esser 2010, Zeng 2009, Jin 2006, 1 2-Inhibition of signaling pathways - Cell surfaces proteins (CD47, CD123, C-LL1) - Chemical inhibitors Majeti 2009, Jin 2009, Van Etten 2010, Takebe 2011 Frankel 2009, Wang
34 Combining Mutational Status and Functional assays Mutation analysis Colony assays, LTC-IC in MDS/MPN - disease definition - drug assessment in vitro - Disease monitoring in vivo o
35 Clonal evolution of JAK2 vs. TET2 variants in MPN patients treated by IFN UPN #1 UPN #2 UPN #3 UPN #4 UPN #5 JAK2 after IFN TET2 before IFN TET2 after IFN JAK2 before IFN
36 UPN #1 UPN #2 UPN #3 UPN #4 UPN #5 JAK2 after IFN TET2 before IFN TET2 after IFN JAK2 before IFN
37 IFNa targets JAK2mut clones but not TET2mut clones UPN #1 UPN #2 UPN #3 UPN #4 UPN #5 JAK2 after IFN TET2 before IFN TET2 after IFN JAK2 before IFN Cassinat & Kiladjian
38 anti-cd44 mab eradicates human AML cells human LAM cells Control and Anti-CD44 3 times/wk for 1 month Xenografts treated Control anti-cd Jin et al. Nat. Med 2006.
39 Engrafment anti-cd44 mab eradicates human leukemia initiating cells Jin et al. Nat. Med human LAM cells Control and Anti-CD44 3 times/wk for 1 month 100 Control + Ac CD44 BM cells nd transplant Weeks after transplantation Similar preclinical models validated anti-cd47 and anti-cd123 mab to eradicate AML and ALL leukemic stem cells
40 % hcd45+ An anti-cd44 mab induces differentiation of haml cells and targets haml LSC in xenografts AML xenografts Control and anti-cd44 treatment 3 times/wk for 1 month First: Eradication of leukemia Second: Eradication of LIC Control + Ac CD44 CD34+CD38- BM cells nd transplant Weeks after transplantation. Coll J DICK Toronto, Jin et al. Nat. Med 2006
41 Cumulative Survival A BCL2 inhitor prolongs survival in an High Risk MDS mouse model and targets Initiating Cells Omidvar Can Res p= MDS + ABT-737 (n=3) p< Untreated (n=27) +ABT-737 (n=23) MDS untreated (n=4) Days from diagnosis Days (post transplant) Beurlet et al Blood 2013
42 Nat Med 2003, Blood 2006, Blood 2010 Immunotherapy targets leukemia initiating cells in transgenic mice models of AML3 AML3 Transplantable model ATRA +PML-RAR FrC n=12 60 ATRA n=12 40 Placebo n= PML-RAR FrC n= Days 120
43 ABT-737 targets leukemia initiating cells in High Risk Myelodysplasia Transgenic Mice Monitoring the disease Blood count Weight Cytology & Histology Progenitor assays hbcl2 expression Arrays ABT times/week 75 mg/kg IP Monitoring the disease Blood count Weight Cytology & Histology Progenitor assays hbcl2 expression Arrays Diagnosis of disease Confirmed hbcl2 Reduced weight Blood count anormal
44 Cumulative Survival ABT-737 prolongs survival in an High Risk MDS preclinical mouse model p= ABT-737 (n=23) Untreated (n=27) Days from diagnosis Beurlet et al Blood 2013
45 ABT-737 prolongs survival by targeting phenotypic LSK+ leukemic stem cells in the BM LSK: Lineage negative/ Sca-1+/c-KIT+ Secondary transplant of lethally irradiated recipients p<0.05 MDS + ABT-737 (n=3) MDS untreated (n=4) Days (post transplant) Beurlet et al Blood 2013
46 Absence of LSC in AML3 long term surviving mice with enhancement of anti-tumor immune response AML3 Transplantable model Treatment Number of mice PML-RAR transcripts PB BM Transferred cells LSC assessment 2 nd transplantation Diseased/ Total Survival no BM (10 4 ) 12/ SPL (10 4 ) 12/ ATRA + DNA 6 neg neg BM (10 7 ) 0/30 >200 SPL (10 7 ) 0/30 >200 Pokorna et al., Molecular and Cellular Probes 2012
47 Role of the microenvironment for initiation or control of the disease
48 Slides courtesy D Bonnet 3D Scaffolds and Biphoton imaging to study the role of the microenvironment 3D Scaffolds to study role of MSC, and modifications Of the microenvironment in vivo Biphoton to study in vivo cells in their environment
49 Summary Urgent need A) to continue and develop more longitudinal, retrospective studies combining genomic and functional studies - Serial Banking and Omics linked to functional studies and patient outcome - validate «danger» or «calm» mutational profiles - novel biomarkers of MRD and novel MRD definition - develop novel therapies (induction, consolidation and maintenance) - companion drugs, microenvironment, enhance antiumor immune responses. - identify parameters that influence tumor evolution in patients in CR - induction/consolidation/maintenance treatments, infections, environment B) to train hematologists, biologists, researchers, bioinformaticians in these new technologies and definitions
50 Institut Universitaire d Hématologie Research Unit UMRS 1131 Group Leaders Christine Chomienne/Fabien Guidez Jean Jacques Kiladjian/Bruno Cassinat Helene Cavé Rose Ann Padua/Pierre Fenaux Christine Dosquet/Dominique Bonnet Institut Universitaire d Hématologie Animal Facility, Imaging plateform FIM, GFM, GFLAP Patients Cancer Stem Cell Networks EHA SWG LSC
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