De la génomique à la thérapeutique Axel KAHN Directeur de l’Institut COCHIN et de l’IFR Alfred JOST Genopole® : Genopole® La médecine de demainLa médecines’inventede demain aujourd’huis’inventeaujourd’hui THE DISEASES OF THE 21st CENTURY Cancers Neurodegenerative disorders Cardiovascular diseases Nutrition diseases Infectious diseases Monogenic diseases Genopole® La médecinede demains’inventeaujourd’hui AXEL KAHN 3.06.2003 TheThe challengeschallenges forfor progressprogress inin therapeuticstherapeutics inin thethe XXIXXIst centurycentury Variable targets Complex physiopathogenic mechanisms Chronic diseases with irreversible lesions Genopole® LaAXEL médecine KAHNde demains’invente 3.06.2003aujourd’hui GENOMICS = LARGE SCALE STUDY OF THE GENOMES Informational Genomics = maps, sequence Functional Genomics = gene function (reverse genetics) expression maps - transcriptome - proteome Structural Genomics = from genetic information to protein structure accumulation of a huge amount of more or less crude data in extensive databases How can we extract pertinent information from these data? Biology, physiopathology hypotheses bioinformatics ....for raising or studying the hypotheses Genopole® La médecinede demains’inventeaujourd’hui AXEL KAHN 3.06.2003 GENOMICS genetic targets mechanisms r.proteins gene therapy information predictive drugs medicine IMPROVED THERAPY Genopole® La médecinede demains’inventeaujourd’hui AXEL KAHN 3.06.2003 For improving pathophysiological understanding of common diseases and then identifying « validated targets » Weak alleles involved in the common multigenic forms Strong alleles involved in the rare (or exceptional) monogenic forms : Atherosclerosis (LDL-R, ABC-A1, ABC-G5/ABC-G8,ARH) : Alzheimer disease (PS1, PS2, APP) : Obesity (leptin, leptin R, MCH and receptor, MSH and receptors) : Familial hypertrophic cardiomyopathy (AMPK) : Inflammatory anemias : Iron overload Hepcidin Genopole® La médecinede demains’inventeaujourd’hui AXEL KAHN 3.06.2003 Although cancer usually : - results from several associated molecular events - is associated with genetic instability Is it possible to kill or to cure cancerous cells by acting on just one of the involved specific events ? RA in PML Gleevec in CML Kit (+) GIST Genopole® La médecinede demains’inventeaujourd’hui AXEL KAHN 3.06.2003 databases Drug design Bioinformatics Candidate small targets Medicinal chemistry molecules the future? combinatorial chemistry, HTS Physiopathology THE BOTTLENECK Genopole® La médecinede demains’inventeaujourd’hui AXEL KAHN 3.06.2003 WORLWIDE SALES OF SMALL MOLECULES /vs RECOMBINANT PROTEINS (1997) : 3 therapeutic proteins among the top ten selling drugs (EPO, insulin, G-CSF) : 8.4 billions $ vs •• 25 billions total already 1/3 of the sales (among the top ten...) .... while only •• 8.000 / 100.000 genes were exploited Genopole® La médecinede demains’inventeaujourd’hui AXEL KAHN 3.06.2003 SOME PROMISES OF THERAPEUTIC PROTEINS • cancer : antiangiogenic factors : antibodies (Herceptin) • diabetes type 2, lipodystrophies, obesity : leptin-like factors increasing insulin sensitivity ? : adiponectin • cardiovascular diseases : angiogenic factors • regenerative medicine : growth factors for specific cells and tissues - b cells (IDDM) - brain cells - nerves - spinal cord • autoimmune diseases, inflammations : b-interferon and multiple sclerosis : IL 13 antagonists and asthma Genopole® La médecinede demains’inventeaujourd’hui AXEL KAHN 3.06.2003 PharmacogenomicsPharmacogenomics Distinction of different groups of patients with respect to : drug toxicity drug efficacy candidate gene polymorphism or alteration (e.g. P450 isoforms, susceptibility genes,disease genes) tumor identity (gene markers, transcriptome, proteome) full genome screening with DNA microchips Genopole® La médecinede demains’inventeaujourd’hui AXEL KAHN 3.06.2003 WhenWhen humanhuman genomicsgenomics failsfails toto leadlead toto diseasedisease etiologyetiology Gastroduodenal ulcers - anti-histamine - anti-acids - hisR antagonists - proton pump antagonists Helicobacter pylori Genopole® La médecinede demains’inventeaujourd’hui AXEL KAHN 3.06.2003 TREATMENT OF GENETIC DISEASES BY : biological prosthesis / replacement - organ transplantation - administration of the lacking protein - gene transfer : gene correction Genopole® LaAxel médecine KAHNde demain 25.2.03s’inventeaujourd’hui TREATMENT OF GENETIC DISEASES BY SMALL CHEMICALS Tyrosinemia type 1 NTBC Familial ataxia (AVED), Vit E a-TPT deficiency Friedreich ataxia Quinones (Idebenone) Lysosomal diseases Ligands of a-galactosidase Glucosyltransferase inhibitors (NB-DNJ and NB-DGJ) Congenital hypothyroidy T3,T4 Genopole® AxelLa Kahnmédecinede demain6.3.03s’inventeaujourd’hui PROSPECTS IN GENE THERAPY • New mode of recombinant protein delivery; very active protein(EPO, angiogenic factors, clotting factors, etc…) • Corrected cells with a clear selective advantage(bone marrow,liver) • Correction of point mutations in the liver when partial correction is sufficient • …… but safety problems • Cell autonomous effect of transferred or corrected genes • High % of correction required • Disseminated or poorly accessible target cells • High-level/prolonged transgene expression needed Genopole® Axel KAHN 25.2.03 La médecinede demains’inventeaujourd’hui Genopole® AXEL KAHN 3.06.2003 La médecinede demains’inventeaujourd’hui Genopole® AXEL KAHN 3.06.2003 La médecinede demains’inventeaujourd’hui DNA and gene….. as a vaccine Instead of developing antibodies against microorganisms and proteins... Injection of DNA will direct cellular synthesis of antigens safe easy stable cheap cellular immunity May be a breakthrough in preventive or curative vaccinology Genopole® AxelLa médecine KAHNde demain 25.2.03s’inventeaujourd’hui REGENERATIVE MEDICINE A new concept for the future Its goal : to replace aged, diseased cells (or tissues) by young genetically similar cells (or compatible tissues) assuring a restored function Could revolutionize medicine in the new century Genopole® La médecineAxel deKAHN demains’invente 25.2.03aujourd’hui REGENERATIVE MEDICINE Neurodegenerative diseases Eye, ear degenerative diseases Spinal cord damages Bone marrow diseases cells Engineered Diabetes cells Heart failure Joint diseases Skin diseases Tissue engineering (bladder, vessel, skin..) Genopole® AXEL LaKAHN médecine de25.2.2003 demains’inventeaujourd’hui HSC MSC Mesangioblasts Tissue-Specific 3 2 Pluripotent Egg ESC (MAPC) Totipotent Multipotent 1 1 : « therapeutic » cloning 2 : multipotent adult stem cells. 3 : reprogrammed adult cells Genopole® La médecinede demains’inventeaujourd’hui AXEL KAHN 25/02/03 Differentiated ES Cells : therapy in vivo Rats with PD models 1. Undifferentiated cells • Teratomas (5/25) • In situ dif. into dopaminergic neurons 2. Ex vivo differentiation into dopaminergic neurons • transformed with a nurr-1 transgene • normal function in vivo, some improvement Ex vivo differentiation into b cells • 2-5% of insulin content • May be, no insulin synthesis at all • Transient effet Ex vivo differentiation into HSC •<2% of immunocompetent cells Genopole® La médecinede demains’inventeAXELaujourd’hui KAHN 12.06.2002 Cell therapy with somatic (stem) cells already a reality in humans Usually ….. • HSC and hematological diseases • Keratinocytes and skin grafts (burnt patients) Experimentally…… • Fœtal neural cells in neurodegenerative diseases • Islets b-cells in diabetes mellitus • Hepatocytes in liver failure and metabolic diseases • Muscle cells and bone marrow cells in myocardium infarction Genopole® AXELLa médecine KAHNde demain 25.2.2003s’inventeaujourd’hui Genopole® 3.06.2003 La médecinede demains’inventeaujourd’hui Genopole® 3..06.2003 La médecinede demains’inventeaujourd’hui Genopole® 3.06.2003 La médecinede demains’inventeaujourd’hui ADULT CELL REPROGRAMMATION « Therapeutic » cloning Nucleus exchange with ESC In vivo/ex vivo spontaneous reprogrammation ? (e.g,MAPCs) In vivo/ex vivo transfer of differentiation master gene(s) (e.g, Pdx1-Vp16, Neurod, betacellulin) In vivo/ex vivo cell fusion In vitro reprogrammation with cell free extracts (permeabilized cells + nuclear extracts of SC or differentiated cells) Genopole® La médecinede demains’inventeaujourd’hui AXEL KAHN 25/02/03 LIVER OVAL CELL High glucose Insulin- secreting cells Neurod + btc Pdx 1/VP16 « hit and run » system HEPATOCYTES ® AXEL KAHNGenopole La médecinede demains’invente3..06.2003aujourd’hui Multipotent Somatic Stem Cells Bone Marrow MSC Mesoangioblasts Multipotent Adult Stem Cells § bone §Cartilage §Fat Vessels Endothelial progenitors §Vessels Cord §Cardiocytes Cord blood §Myocytes §Hepatocytes Somatic epidermal SC §Glial cells §Neurons §…….etc Skin Skin-derived precursors Genopole® La médecinede demainAXELs’invente aujourd’huiKAHN 12.06.2002 Pluripotent and tissue-specific stem cells • Brain • MSC • Heart • Skeletal muscle • Blood (HSC) • Skin • Pancreas • etc…. Genopole® La médecinede demains’inventeaujourd’hui Axel Kahn 20/1/2004 Major hurdles with stem cell-based regenerative medicine ° To obtain enough starting stem cells (SC) (from adults) ° To control SC in vitro amplification and differentiation ° To eliminate non-differentiated, potentially tumorigenic cells before transplantation (cell sorting, differentiation- specific positive or negative selection markers) Persisting uncertainties ° Even if enough differentiated cells can be obtained, what will be their long-term - function - viability - tumorigenic potential Genopole® LaAXEL médecine KAHNde demains’invente3.06.03aujourd’hui GENOME PROGRAMME Differentiation and Growth factors STEM CELLS expansion (embryonic / DIFFERENTIATED CELLS adult) differentiation Genopole® AXELLa médecine KAHNde demains’invente 3.06.03aujourd’hui.