Transcript Lysosomal Storage Disease

Lysosomal Storage Disease
Module 755
The Brain in Health and Disease
Sean Sweeney
Lysosomal Storage Disease (Amaurotic Idiocy)
c.a. 45 autosomal recessive diseases
Individually rare
Collectively occur c.a. 1/8000 live births
Cause death in early to late childhood (after normal infancy)
Varying involvement of the nervous system
All ‘store’ material in the lysosome due to defects in
substrate degradation or biogenesis of the lysosome
The Lysosome
subcellular electron dense organelle
filled with c.a. 70 hydrolytic enzymes: will break down all
biological macromolecules
low pH (~4.0), membrane bound
Considered the ‘gut’ or garbage disposal unit of cell
Material for degradation trafficked to lysosome via endocytosis
or autophagy
Lysosomal enzymes trafficked to lysosome via M6P receptor
pathway
Delivering material for
degradation to the lysosome:
endocytosis and autophagy
Endosome to lysosome:
decreasing pH, membrane
limited.
Autophagy: controls cell size,
used during caloric restriction,
Phagocytosis:- degrades
‘dead’ cells, pathogens
Autophagy and phagocytosis
meet in the Phagolysosome
Professional Phagocytes:
macrophages, neutrophils
Endocytosis in the nervous system
The polarised and extended
structure of the neuron creates
a trafficking problem for neurons:
‘lysosomes’ (as we know them!)
not present at synapse.
Late endosomal markers present:
fuse with lysosomes in the soma
QuickTime™ and a
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Delivering degradative enzymes and cofactors to the lysosome, the M6P/M6PR
pathway.
Mannose-6-phosphate group added to lysosomal
hydrolases via N-linked oligosaccharides as
hydrolases transit through cis-golgi
M6P recognised by M6P-receptors in trans-golgi:
delivers them to late endosome
Lower pH causes dissociation
M6PR then retrieved in late endosome
and trafficked for re-use in trans-golgi
(recognised via C-terminal tail).
General outline of LSD dysfunction:
Mutations arising in hydrolytic enzyme, co-factor or
factor essential of enzyme delivery to lysosome
Also, factors essential for lysosome function and
biogenesis (membrane proteins, channels and proteins of
unknown function) plus factors for protein traffic to lysosome
Material (substrate) continues to be delivered to lysosome
resulting in ‘stored’ material, usually ‘primary’ and ‘secondary’
leads to swollen lysosomes
Developmental dysfunction and early death: symptoms
v. variable, varying involvement of different tissues
General Cellular Phenotype:
Swollen, multilammellar ‘osmiophilic endosomes/lysosomes
(function? pH?)
Accumulation of lipofuscin/ceroid ‘ageing pigment’
Defects in autophagy (?)
Appearance of meganeurites (variable)
QuickTime™ and a
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Cellular phenotype contd.
Excessive synaptogenesis/dendritogenesis
(MPS and sphingolipidoses)
Shrinkage of the CNS (variable)
Mistrafficking of cholesterol
(cholesterol recycling?)
Why are symptoms and effects in different organs variable?
tissue turnover rates?
presence (or relative abundance) of substrate?
sensitivity of cell type (neurons and polarity)?
What is the ‘pathogenic cascade’?
(volume of substrate not key!!!)
Classification :
Mucopolysaccharidoses (variable nervous system involvement)
Mucolipidoses (originally considered an MPS)
Glycoproteinoses
Glycogen storage
Sphingolipidoses
Lipid storage disorders
Multiple enzyme defects
Transport defects
Batten Disease
(Red = nervous system involvement)
Mucopolysaccharides
• Defective metabolism and accumulation of GAGs
• Most abundant polysaccharides
• Long unbranchedstructure containing
disaccharide units:
• High viscosity + rigidity
• Excellent lubricators and shock absorbers
• Important component of cell membranes
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Mucopolysaccharidoses:
Enzyme Defective
MPS-I: (Hurler, Sheie, Hurler/Sheie)
iduronidase
MPS-II: (Hunter)
iduronate-2-sulfatase
MPS-III: (Sanfilippo)
IIIA
heparan-N-sulfatase
IIIB
N-acetyl-glucosaminidase
IIIC
Acetyl Co-A glucosamine
N-acetyl transferase
IIID
N-acetyl-glucosamine-6-sulfatase
MPS-IV (Morquio)
IVA
N-acetyl-galactosamine 6-sulfatase
IVB
ß-galactosidase
MPS-VI (Maroteaux Lamy)
N-acetyl-galatosamine 4-sulfatase
MPS-VII (Sly)
ß-glucuronidase
MPS-IX
hyaluronidase
Sanfilippo Syndrome (MPS III)
Four types: A,B,C,D, cannot break down Heparan sulfate
Most common MPS, 1/70,000 births
hepatosplenomegaly (may resume normal size with age)
Hyperactivity
Speech delay
Mental retardation
Joint stiffness, bone defects (dystosis multiplex)
Coarse features (dysmorphism
Death in middle teens
Screening: GAGs in urine
Diagnostic: WBC enzyme assay or plasma enzyme assay
Prognosis: No effective treatment to date.
Mucolipidosis (I-Cell disease) and MPS-IV
Mucolipidosis-II
I-Cell (Pseudo-Hurler): first described 1967
I = Inclusion, stored material mucolipid MPS and sphingolipid
Occurrence: 1/640,000 live births
Symptoms: Developmental delay, psychomotor deterioration, dysmorphia, death in
early childhood
Genetic defect: N-acetylglucosaminyl-1-phosphotransferase
Prognosis: v. poor, limited treatment (nutritional), death by 10 years of age.
Mucolipidosis-IV
Storage material: mucolipids, MPS and sphingolipids
Occurrence: carriers in Ashkenazim Jewish population, 1/90 to 1/100
Symptoms: Psychomotor retardation, corneal opacity, retinal degeneration, iron
deficiency, improper stomach pH (achloridia)
Genetic defect: Mucolipin-1 (MCOLN1), a TRP channel (TRPML-1)
Involved in Fe2+ efflux from lysosomes? (Dong et al., (2008) Nature, 455, 992-6)
Prognosis: v. poor. Nutritional supplements, physcial and speech therapy
Sphingolipids: a major component of neural tissue
Ceramide
STRUCTURE
OH
CH2O
H
microdomains (?)
trafficking
NH
O
SIGNALLING
Sphingomyelin
O
OH
CH2O
NH
P
CH3
O
(CH2)2 N+ CH3
CH3
O-
Apoptosis
proliferation
stress
O
Glycosphingolipids
OH
CH2O
NH
O
Glc
n
- Sphingomyelin
- Ceramide
- Sphingosine
- Sphingosine-1-phosphate
- Cerebrosides
- Gangliosides
Sphingolipids are
tightly associated with
cholesterol
The sphingolipidoses: Tay-Sachs (GM2-gangliosidosis)
First described in 1880’s from ‘cherry-red’ spot in fundus (retina) (lipid deposition in bipolar
ganglion cells)
Infantile (death ~ 5yrs), Juvenile (death between 5 and 15yrs) and ‘Late-onset’ forms (v. rare)
All present with increasing neurological and deterioration (ataxia, atrophy, spasticity)
Occurrence: 1/27 to 1/30 Ashkenazim Jews are carriers, also: Acadians, Cajuns
Genetic defect: Hexosaminidase A (HEXA)
storage material: GM2 ganglioside, globoside, glycolipids
cf: Sandhoff Disease: HEXB mutations and GM2 gangliosidosis
(mutations in GM2 activator protein)
Glial Involvement!
Prognosis: early death, ameliorated by treatment
Enzyme Replacement Therapy
Substrate Reduction Therapy
Population Screening
(model of genetic screening for recessive condition)
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Other cellular defects:
Niemann-Pick disease:
occurrence: A, B collectively- 1/1000 Ashkenazim Jews are carriers, type C no ethnic distribution
type A accounts for 85% of cases
Symptoms: enlarged spleen and liver, enlarged lymph nodes, darkening of skin, neurologic
impairment (not in B), cherry red spot
genetic defect: A and B, mutant for sphingomyelinase
Type C mutants: two loci, two proteins, multi-transmembrane protein (related to
hedgehog receptor ‘patched’ and small co-protein(cholesterol binding protein/carrier?).
Homolog NPCL1 involved in cholesterol absorption in gut.
storage material: sphingomyelin, cholesterol and sphingolipids
Diagnosis: ‘filipin’ staining
cell biology (and diagnosis): mislocalised unesterified cholesterol, neurofibrillary tangles
Endosomal trafficking jam? cholesterol and sphingolipids required to organise endosomal
trafficking steps. Cholesterol recycled from lysosome.
Drosophila models reveal cholesterol is ‘limited’
Batten disease
QuickTime™ and a
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A family of closely related disorders
9 forms: congenital, infantile, late infantile, juvenile
adult
AKA: Neuronal Ceroid Lipofuscinosis (NCL)
Batten (1903)
Incidence: global with hotspots for some loci
Loci: ‘CLN’ genes CLN1, CLN2, CLN3, CLN5, CLN6, CLN8 CTSD
cloned so far, others remain to be mapped.
occurrence: most common childhood neurodegeneration 1/8000 livebirths
Symptoms: visual defects, seizures, stumbling, echolalia, eventual loss of sight speech
and motor skills, early death after blindness, dementia.
storage material: Lipofuscin/ceroid, subunit C of mitochondrial ATP synthase
Phenotype: multilamellar inclusions, selective brain cell death (glia mediated)
infiltration of neuronal tissue with antibodies (defective BBB?)
Prognosis and treatment: anti-convulsives, therapy. Death in childhood
Locus
Disease
Protein deficiency
Function
CLN1
infantile NCL
palmitoyl protein thioesterase
de-palmitoylation
Lysosome
CLN2
late infantile NCL
tripeptidyl peptidase
protease
lysosome
CLN3
juvenile NCL
transmembrane protein
?
lysosome
CLN4
adult (Kuf’s)
Not identified
CLN5
late infantile NCL
(Finnish variant)
transmembrane protein
CLN6
late infantile variant transmembrane protein
CLN7
late infantile variant Not Identified
CLN8
EPMR
transmembrane protein
ER, ER/Golgi
CTSD
Ovine NCL
cathepsin D
protease
lysosome
?
LE/lysosome
ER protein
Endocytosis in the nervous system
Lysosomes (hydrolases!)
not present at synapse
Many of NCL proteins found
at synapse
NPC protein, others?
QuickTime™ and a
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are needed to see this picture.
Identification of proteins involved
in neurodegeneration help
to describe functions in the
neuron
Treatment:
BMT (membrane proteins)
enzyme replacement (BBB?)
gene therapy
substrate reduction- Miglustat (monosaccharide
mimetic-imino sugar)
Neuronal stem cells (membrane proteins?)
Chemical chaperone therapy
Neuroinflammation
Economic cost
ERT is current most effective treatment (non neurodegenerative LSDs):
Disease
Treatment
Annual Cost (per patient in $)
Gaucher
ERT
145,000 - 290,000
Gaucher
SRT
91,000
Fabry
ERT
156,000
Hurler-Scheie (MPS-I)
ERT
340,000
Maroteaux-Lamy (MPS-VI)
ERT
377,000
Reasons:
High regulatory costs
Cost of research
Lack of competition (Orphan Drug Act 1983, US)
Studying the Lysosomal Storage Diseases:
Model Organisms
Sheep (Batten)
sheepdogs (Batten)
mouse (Batten, Tay-Sachs, Sandhoff, NPC)
zebrafish (Batten)
Drosophila (MPS, NPC, Batten, others)
C. elegans (MPS, NPC)
Yeast (cerviseae, pombe) Batten, NPC
Reverse Genetics (qv Tay-Sachs)
Forward Genetics
The Drosophila neuromuscular junction:
A model glutamatergic synapse
http://132.236.112.18/fruitfly/shaker/physiology/
spinster synapses are overgrown
spinster
suppresses
synaptic growth
spinster mutants
have a shortened
lifespan
spinster encodes a twelve transmembrane transporter
4 transcripts = 12 TM domains
1 transcript = 8 TM domains
Spin localises to a low pH late-endosomal compartment
A low pH compartment is expanded in spin mutants
Loss of spinster induces a redistribution of cholesterol
WT
filipin
spin4/spin5
spinster identifies a novel component of the late endosome/lysosome that when mutated gives
rise to all of the hallmarks of lysosomal storage disease
spinster potentially identifies a signalling pathway driving synaptic overgrowth
Summary
Lysosomal storage disease are caused by defects in lysosomal hydrolases and proteins
essential to lysosomal biogenesis/function
LSD lysosomal defects give rise to swollen lysosomes, developmental and degenerative
defects with varying involvement of the nervous system due to ‘storage’ of material
in the lysosome.
Lysosomal storage diseases identify proteins essential to lysosomal function
LSDs cause death in childhood (generally) after normal infancy
LSDs are essentially incurable, but some are treatable to varying degrees.
Model organisms are helping to define the biology of the LSDs, in particular the
‘pathogenic cascade’