New therapeutic approaches for renal inflammation Renato Monteiro, MD, PhD Bichat Hospital – University Paris 7 - Inserm U...
Glomerulonephritis (GN) <ul><li>Inflammation of the glomeruli, or small blood vessels in the kidneys.  </li></ul><ul><li>M...
Geneticfactors Inflammatory reaction and glomerulonephritis Fibrosis and decline in renal function Renal failure Glomerulo...
<ul><li>Young individuals </li></ul><ul><li>responsible for 10% of renal transplant </li></ul><ul><li>altered IgA1 glycosy...
HumanCD89 Humanized model  for IgANephropathy CD11b promotor EGF Stop codon Transgene 83 96 73 Lines Tg CD89 CD11b Hematur...
Mechanism of IgAdeposition in the kidneys Circulating IgA-complexes Mesangial IgA deposits TfR expression IgAN proliferati...
IgE- or IgG-IC FcRs Fab (clone A77) CD89 ITAM dependent renal inflammation 2-Therapeutic antibody: Fab anti-CD89 MAPKs  ac...
Fab Anti-CD89 suppresses  renal inflammation  0 1 2 320 A77 PBS 0 2 4 6 12 10 8 14 Proteinuria (g/dl) * * * Day Proteinuri...
Fab anti-CD89 reduces renal fibrosis  UUO UUO Left Right A77 Fab ctr Fab (320) Left Right Kanamaru  et al. J Immunol  2008...
Other diseases with renal involvement:  diabetes… TNF   (pg/ml) 0 500 1000 1500 2000 2500 LPS * Inflammatory renal diseas...
Fab anti-CD89 : A new anti-inflammatory treatment For severe nephritis targeting macrophages Inhibition Activation
Anti-inflammatory action in other diseases: <ul><li>Prevention of Asthma in a mouse model (Pasquier et al  Immunity  2005)...
INATHER Y S Fusion Immunization Double Tags  Plasmid Protein purification Generating therapeutic monoclonal Antibodies for...
Take home messages: <ul><li>Development of therapeutic antibodies based on physiopathological data and on improved screeni...
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13 rencontres biomédicale LIR Renato Monteiro

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2ND INTERNATIONAL RESEARCH MEETING
4 JUNE 2010- HÔTEL DE MARIGNY PARIS

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  • 4
  • Treatment with A77 Fab inhibits heterologous LPS-induced TNFa production of human blood monocytes and autologous IgA-IC induced b-hexosaminidase release in FcaRI RBL mast cells trnsfectants. A) Human blood monocytes were incubated stimulated with LPS (X mg/ml)and TNFa production (black bars, n= 4) was measured and compared to unstimulated cells (white bars). This response was inhibited (*, p&lt; 0.05) after pretreatment (24 h) of cells with A77 Fab, but not after pretreatment with 320 Fab. B) FcaRI-RBL Fc  RI R209L/  or ITAM-mutated Fc  RI R209L/ g Y268/278F mast cell transfectants were preincubated with either PBS, 320 Fab or A77 Fab before incubation with either PBS or PEG purified IC from 10 patients with IgA nephropathy and b-hexosaminidase release was determined. Data show marked inhibition by A77 Fab in Fc  RI R209L/  . 6 out of the 1O patient IC induced also
  • 13 rencontres biomédicale LIR Renato Monteiro

    1. 1. New therapeutic approaches for renal inflammation Renato Monteiro, MD, PhD Bichat Hospital – University Paris 7 - Inserm U699
    2. 2. Glomerulonephritis (GN) <ul><li>Inflammation of the glomeruli, or small blood vessels in the kidneys. </li></ul><ul><li>Manifested with hematuria, proteinuria or renal failure. </li></ul><ul><li>IgAnephropathy and systemic lupus erythematous are major causes of </li></ul><ul><li>GN which affects over 2 million people in the US, with 310,000 cases/year. </li></ul><ul><li>No specific treatment is available. </li></ul><ul><li>Market : over 600 M€ and rising at the rate of 10% a year in the US. </li></ul><ul><li>France: Nephrotestcohort (Bichat, Tenon, HEGP hospitals) : </li></ul><ul><li>> 1200 patients with chronic kidney diseases including GN. </li></ul>
    3. 3. Geneticfactors Inflammatory reaction and glomerulonephritis Fibrosis and decline in renal function Renal failure Glomerulonephritis Receptors IFN  TNF GM-CSF Antigen Neutrophil Mastcell Macrophage Dendriticcell T Lymphocyte B Lymphocyte TNF Chemokines TNF Histamines Trauma Antigens… Immune complexes Pathogens LESIONS Cell Signalling
    4. 4. <ul><li>Young individuals </li></ul><ul><li>responsible for 10% of renal transplant </li></ul><ul><li>altered IgA1 glycosylation </li></ul><ul><li>IgA1 complexes in the mesangium </li></ul>IgANephropathy (IgA-N) Human IgA receptor: CD89 = Fc α RI R D1 D2 R ITAM Co IgAN 4 2 Soluble CD89 (Units) GN
    5. 5. HumanCD89 Humanized model for IgANephropathy CD11b promotor EGF Stop codon Transgene 83 96 73 Lines Tg CD89 CD11b Hematuria - + Launay et al J Exp Med 2000 Tg Lt CD89Tg IgA-N SCID S e rum: IgA/sCD89 IC SCID IgA-N Pathogenic role of sCD89
    6. 6. Mechanism of IgAdeposition in the kidneys Circulating IgA-complexes Mesangial IgA deposits TfR expression IgAN proliferation Moura et al J Exp Med 2001 Moura et al Blood 2004 1-Therapeutic antibody: the A24 mAb Mechanism of action Fe-Tf Transferrin receptor APOPTOSIS Patent WO 2005/111082
    7. 7. IgE- or IgG-IC FcRs Fab (clone A77) CD89 ITAM dependent renal inflammation 2-Therapeutic antibody: Fab anti-CD89 MAPKs activation MCP-1 GPCR TNFR TNF  Non- ITAM dependent renal inflammation Kanamaru et al J Immunol 2008 Pasquier et al Immunity 2005 SHP-1 Silva et al Nat Med 2007 Patent WO 2005/089798 Mechanism of action < < < < < < < < < < < < < < < < Ag < < < < < < < < Ag
    8. 8. Fab Anti-CD89 suppresses renal inflammation 0 1 2 320 A77 PBS 0 2 4 6 12 10 8 14 Proteinuria (g/dl) * * * Day Proteinuria Creatinine BUN (mmol/l) 60 40 20 0 d14 d7  mol/l) 40 20 0 * * * Renal parameter Experimental glomerulonephritis Immunohistochemistry Macrophages PBS 320 A77 T cells Injection of rabbit anti-GBM Ab Production of Autologous Ab IgG-IC GN Neutrophil influx Heterologous phase Autologous phase Protocol Tg hFc  RI mice Kanamaru et al. J Immunol 2008
    9. 9. Fab anti-CD89 reduces renal fibrosis UUO UUO Left Right A77 Fab ctr Fab (320) Left Right Kanamaru et al. J Immunol 2008 Tubular dilatation Tubular atrophy Cell infiltration 3 1 0 2 4 * 3 1 0 2 4 * 3 1 0 2 4 * Ligation (left kidney) PBS A77 Fab ctr Fab (320) PBS A77 Fab ctr Fab (320) Macrophages Collagen PBS A77 Fab Ctr Fab (320)
    10. 10. Other diseases with renal involvement: diabetes… TNF  (pg/ml) 0 500 1000 1500 2000 2500 LPS * Inflammatory renal disease: glomerulonephritis TNF  (pg/ml) 0 500 1000 1500 2000 2500 LPS * Kanamaruet al J Immunol 2008 LPS activation of blood cells Fab anti-CD89 reverses activation of patient cells PBS A77 320
    11. 11. Fab anti-CD89 : A new anti-inflammatory treatment For severe nephritis targeting macrophages Inhibition Activation
    12. 12. Anti-inflammatory action in other diseases: <ul><li>Prevention of Asthma in a mouse model (Pasquier et al Immunity 2005) </li></ul><ul><li>Prevention of Arthritis in a mouse model, (unpublished) </li></ul><ul><li>Diabetes? ANR BiotecS to address this issue </li></ul>
    13. 13. INATHER Y S Fusion Immunization Double Tags Plasmid Protein purification Generating therapeutic monoclonal Antibodies for clinical application Human Therapeutic Target In vivo Glomerulonephritis Blood Sample Ex vivo Multiple Screening Strategy Inflammatory Models NUDE Tg Cell proliferation Leads Optimization Sequence analysis Chimeric antibody production (ex A24 and A77) Western Blot Cytometry Hybridoma isolation Cell proliferation Extended period for screening
    14. 14. Take home messages: <ul><li>Development of therapeutic antibodies based on physiopathological data and on improved screening methods. </li></ul><ul><li>Going from production to proof of concept (POC) in animals to allow I.P. development. </li></ul><ul><li>Humanized model of IgA-N ready for POC of new candidate drugs. </li></ul>

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