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N Cartier Pediatric Neurology Bictre Hospital, Paris INSERM U745 - Facult de Pharmacie, Universit Paris-Descartes
More than 40 different inherited metabolic diseases; 1/7000 births Lysosmal proteins are ubiquitously expressed : Multiple organs Most involve the CNS Subclasses : Transporters Majority : deficit in soluble lysosomal hydrolases Progressive course : implication for therapy
Enzyme is released by normal cells and taken up by deficient cells (mannose 6-phosphate receptor) Low levels of enzyme are presumed to be therapeutic Therapeutic implications:
enzyme replacement strategies engineered tissue sources of enzyme/cell or gene replacement strategies
Giancarlo Parenti
EMBO Mol Med 1, 268279
Standard care for Gaucher type 1 Fabry : reduces cardiomyopathie et cerebraovascular complications, and slows progressiion to renal failure MPS I,II I : positive effect on joint, lung, organomegaly but no effect on the CNS involvment Pompe : partially ameliorates muscle strength and heart function
Limitations : access to organs : CNS +++ Immunological tolerance in CRIM patients Combining therapies ? : Enhanced Enzymatic Delivery synthetic oligos containing M6P chaperones (imino sugars)
Allow constant delivery of the therapeutic protein to target organs (particularly difficult to rich) : bone, brain +++
Secretion of lysosomal enzyme from organs ( enzyme factories ) to blood circulation Liver (MPS1 models) , lung (AAV and Fabry) pb + : brain access Brain ++++ (lipofuscinosis, MPS III, MLD) Ex vivo GT Stem cells , encapsulated fibroblasts HSC transplantation (MPS, MLD, Pompe)
neonatal delivery low AAV particules administration combined with ERT leads to enhanced response
Arylsulfatase A ARSA
GALACTO CEREBROSIDE
Prognosis:
fatal within few years from onset
GA SD
cPPT-CTS PGK
L. enzyme WPRE
SA
6000
5000
4000
3000
2000
1000
0
normal donor pt pt + LV
Prevention and correction of functional and histopathological disease manifestations upon pre-symptomatic and symptomatic treatment Gene corrected HSC are more effective than WT HSC Dose-dependent benefit
Biffi et al., JCI 2004; Biffi et al., JCI 2006
Safety & toxicology addressed in: MLD mice, ARSA tg mice, tumor prone models and human HSC
Capotondo et al, HST 2007; Montini et al., Nature Biotech. 2006 and JCI 2009
9 Rapid delivery of ARSA enzyme into the brain to stop progression of the disease in rapidly progressive forms of MLD
Intracerebral injections of AAV2/5-ARSA in MLD mice allows strong expression and diffusion of recombinant ARSA
*
Untreated
Treated
1000-1500
S3-S4
500-1000
S5-S6
200-500 150-200
Cb
M6
PAS reactivity
Microglial activation
Astrogliosis
MLD
MLD treated
control
Rotarod
Treated
White Matter
18 months
Diffusion Tolerance
Cerebral cortex
* *
1.21 0.05 (0,027) 1.24 0.06 (0,029) 1.14 0.04 (0,042) 1.12 0.03 (0,027) 1.16 0.05 (0,049)
-30%
-63%
9 1.1 10e11 PP / hemisphere = 1x dose for patient 9 3 sites per hemisphere; 2 deposits per site: same device as planned in patients 9No immunosuppression 9Brain imaging (short and long term tolerance) 9Neurological evaluation 9Euthanasia 3 months post injection
Injected hemisphere
Injected hemisphere
73% VG+
62% VG+
Non-injected hemisphere
Non-injected hemisphere
11% VG+
1% VG+
Activity ratio
Noninjected
*
400
C
200
IncreaseinARSAactivityoverthetherapeuticallevel
In comparison with AAV2-5/ARSA injected NHP:
9 Higher levels of ARSA activity in the whole injected hemisphere 9 with ten-fold less vector
Anterior site
Median site
Posterior site
T2FLAIRaxial
T2axial
T2axial
T2TSEaxial
MRI at day 3
CHOPIN
T2TSEsagittal
9 Toxicological study in rat non-human primate with a GLP grade AAVrh10-ARSA vector batch (done) 9 Production of the GMP batch (done) 9 Pre-IND to AFSSAPS (done) 9 Final Submission to AFSSAPS in September 2011
9 5 MLD patients (males or females) with onset of symptoms between 1-5 years (CRIM ARSA +) 9 Total dose of injected AAVrh10-ARSA vector: 4. 1012 vg (3.3 1011 vg / deposit) 9 Targeted regions : anterior medial and posterior part of centrum ovale in each hemisphere 9 3 tracks/hemisphere; 2 deposits/track (12 sites injected simultaneously) 9 No immunosuppression 9 2-years follow-up . Safety . Efficiency (motor and cognitive functions, MRI, electrophysiology)
TIGET San rafaelle A Biffi M Sessa MG Roncarolo L Lorioli T Plati A Capotondo F Fumagalli E Montini L Naldini
C. Sevin N. Cartier F. Piguet M. Zerah T. Roujeau C. Bouquet F Fouquet I Bieche M Vidaud P Aubourg