Post on 23-Jan-2018
transcript
Know Your Models!(Know your Disease)
Multiple Sclerosis Models &Experience with Clinical Translation
MouseAGE 2015
Prof. David Bakerdavid.baker@qmul.ac.uk
Slides Available on www.ms-res.org
(Slideshare)
MOUSEAGE 2015
• KNOW YOUR DISEASE-DESCRIPTION OF MULTIPLE SCLEROSIS
• KNOW YOUR MODEL- DESCRIPTION OF MS MODELS
• KNOW THE LIMITATIONS: FAILURE TO TRANSLATE
• LIMIT YOUR LIMITATIONS: ARRIVE REPORTING GUIDELINES
• DRUG DEVELOPMENT: BENCH TO BEDSIDE
Slides Available on www.ms-res.org
(Slideshare)
Multiple sclerosis (MS) is a chronic (auto)immune-mediated CNS-confined demyelinating disease affecting 2,500,000 people worldwide
(Restricted Distribution: Northern European Dissent)
Disease onset is usually between 20 and 40 years of age (F:M ratio, 2:1. Polygenic-150 genes identified so far)
(30% concordance in Identical Twins, 100% Infected with EBV)
MS is clinically characterized by a relapsing-remitting course usually followed by a progressive phase
MS is pathologically heterogeneous – inflammation, demyelination and neurodegeneration (axonal loss and neuronal damage)
Spontaneous repair occurs but MS invariably progresses (ambulatory problems in 70-80% patients at 25 years from onset)
KNOW YOUR DISEASE-BIOLOGY OF MS
NERVE LOSS
PROGRESSIVE MSRELAPSING-REMITTING MS
DISABILITY
Frequent inflammation, demyelination,axonal transections, plasticity and remyelination
Inflammation, Persistent Demyelination & Gliosis
Infrequent inflammation, Gliosis, Chronic Neurodegeneration
CLINICAL THRESHOLD
INFLAMMATION
Symptoms
Clinical Effects are Due to Altered Nerve Conduction
CLINICAL COURSE
Immune-MediatedBeta-interferons, Alemtuzumab (CD52),
Cladribine, Bone Marrow Transplantation, Fingolimod (S1P1-modulator) , Rituximab
(CD20) are ACTIVE
NeurodegenerationBeta-interferon, Alemtuzumab,
Cladribine, RituximabBone Marrow Transplantation,
Fingolimod are INACTIVE
KNOW YOUR DISEASE-BIOLOGY OF MS
Eyes,Brain & Spinal Cord
You are told you have Eyes,Brain & Spinal Cord
You are told you have
White Blood Cells
No Repair
• INFLAMMATORY, DEMYELINATING, NEURODEGENERATIVE DISEASE OF CNS
•MULTIFOCAL OLIGODENDROCYTE LOSS
MS plaques
KNOW YOUR DISEASE-BIOLOGY OF MS
Myelin (blue)stain
NORMAL AGINGATROPHY RATE 0.1% p.a.
MULTIPLE SCLEROSISATROPHY RATE 0.4-1.0 %
MS is a Neurodegenerative Disease
KNOW YOUR DISEASE-BIOLOGY OF MS
Brain atrophy occurs across all stages of the disease
De Stefano, et al. Neurology 2010
n= 963 pwMS
KNOW YOUR DISEASE-BIOLOGY OF MS
ATROPHY
KNOW YOUR DISEASE-BIOLOGY OF MS
Onset of Progressive MS is Age-Sensitive andIndependent of Pre-Progression Disease Course
Confavreaux et al. 2006, Tutuncu et al. 2012
0 10 20 30 40 50 60 70 80
100
80
60
40
20
0
PPMS
SPMS
0 10 20 30 40 50 60 70 80
100
80
60
40
20
0
PPMS
SPMS
Mean Age at MS Onset (Years) Mean Age at PMS Onset (Years)
Per
cen
tag
e
Per
cen
tag
e
KNOW YOUR DISEASE-BIOLOGY OF MS
in vitro
monkeys
rodents
clinic
Mechanisms of Disease Design of Therapies Safety & Ethics
monkeys
clinic
discovery/screening
Validation/safety
safety/ preclinical
clinical trials
KNOW YOUR DISEASE-BIOLOGY OF MS MODEL-WHY USE ANIMALS?
MS was called an Autoimmune, Demyelinating Disease of the White Matter
MYELIN = BROWN
DEMYELINATION
MONONUCLEAR CELLINFILTRATE
BLOOD VESSEL(VENULE)
KNOW YOUR DISEASE-BIOLOGY OF MS/MODEL-BIOLOGY OF MS
MS was called an Autoimmune, Demyelinating Disease of the White Matter
CHEMICAL-INDUCED DEMYELINATION
VIRAL-INDUCED DEMYELINATION
AUTOIMMUNE-INDUCED DEMYELINATION
EXPERIMENTAL AUTOIMMUNEENCEPHALOMYELITIS (EAE)
IN MAMMALS(MYELIN-REACTIVE AUTOIMMUNITY)
SEMLIKI FOREST VIRUSMOUSE HEPATITIS VIRUS(NEUROTROPHIC VIRUS)
CUPRIZONE FEEDINGLYSOLECITHIN INJECTION
(OLIGODENDROCYTE TOXIN)
MONONUCLEAR CELL INFILTRATE
TRANSGENE-INDUCED DEMYELINATION
NO ANIMAL GETS SPONTANEOUS MS-LIKE DISEASE
KNOW YOUR DISEASE-BIOLOGY OF MS/MODEL-BIOLOGY OF MS
Myelin Neurofilament
Demyelination
AxonalTransection
Axonal Transections are Associated with Inflammation
Control White Matter <1transection/mm2
Core of Chronic Lesion 875transection/mm2
Edge of Chronic Active Lesion ~3000transection/mm2
Active Lesion ~11000transection/mm2
MS was called an Autoimmune, Demyelinating Disease of the White Matter
KNOW YOUR DISEASE-BIOLOGY OF MS/MODEL-BIOLOGY OF MS
AUTOIMMUNE-INDUCED DEMYELINATION
EXPERIMENTAL AUTOIMMUNEENCEPHALOMYELITIS (EAE)
IN MAMMALS(MYELIN-REACTIVE AUTOIMMUNITY)
MONONUCLEAR CELL INFILTRATE
Grey Matter Demyelination
Grey Matter Lesions have few T cells & macrophages
Experimental Autoimmune Encephalomyelitis
1. Active EAE - induced with myelin proteins or
peptides in CompleteFreund’s Adjuvant.
Myelin proteins in CFA
2. Adoptive EAE - induced by T cell transfer from mice immunised for active EAE.
T cells to myelin proteins
Myelin proteins in CFA
3. Myelin TCR Transgenic - Develop Spontaneous EAE.
KNOW YOUR MODEL-BIOLOGY OF EAE
No Clinical Disease-No Spinal Cord Infiltration- No Cytokine X, Y or Z Limp tail
Impaired righting
reflex
hindlimb paralysis
Moribund
partial paralysis
Normal
Remission
0
1
2
3
4
5
(1)
Clinical Score
Day 7
Spinal cord homogenate in Freund’s complete adjuvant
Day 0
Spasticity & TremorsDevelop
KNOW YOUR MODEL-BIOLOGY OF EAE
Video is Coming Next
43
No Clinical Disease-No Spinal Cord Infiltration- No Cytokine X, Y or Z Limp tail
Impaired righting
reflex
hindlimb paralysis
Moribund
partial paralysis
Normal
Remission
0
1
2
3
4
5
(1)
Clinical Score
Day 7
Spinal cord homogenate in Freund’s complete adjuvant
Day 0
Spasticity & TremorsDevelop
KNOW YOUR MODEL-BIOLOGY OF EAE
Video is Coming Next
43
MULTIPLE SCLEROSIS MODELS
KNOW YOUR MODEL-BIOLOGY OF EAE
Time Post-Induction (Days)
10 15 20 25 30 35 40
Mea
n N
euro
logi
cal S
core
0.0
0.5
1.0
1.5
2.0
2.5
3.0
3.5
4.0
4.5
Acute (ABH, SJL) EAE
NerveLoss
Chronic (C57BL/6) EAE
TailParalysis
HindLimbParalysis
Video is Coming Next
KNOW YOUR MODEL-BIOLOGY OF EAE
ConductionBlock
Strain Antigen
C57BL/6 MOG35-55
ABH PLP56-70
ABH MOG8-22
ABH MOG35-55
PL/J MBP1-9
SJL MBP89-101
SJL PLP139-151
SJL PLP178-191
MOG Myelin oligodendrocyte glycoprotein, MBP Myelin Basic Protein, PLP Proteolipid Protein
Normal Remission 1
Remission 2 Remission 4
NfH
(µ
g/m
g t
ota
l p
rote
in)
Number of AttacksN RM1 RM2 RM3
50
100
150
200
250
300
350
400Axonal Content Assessed By Neurofilament ELISA
www.msbrainhealth.org
No-evidence of Disease Activity (NEDA)
KNOW YOUR MODEL-BIOLOGY OF EAE
Stop Relapses Save Brain
Leg moved to full
flexion forassessment
Spastic Leg
RE
SIS
TA
NC
E F
OR
CE
TO
HIN
DLI
MB
FL
EX
ION
(N
)
0.00
0.05
0.10
0.15
0.20
0.25
0.30
0.35
0.40
0.45
Left leg Left legLeft legRight Leg Right Leg Right Leg
Non-spastic
Remission
Paralysed
Relapse
Spastic
Remission
0.08 ± 0.01# 0.03 ± 0.01* 0.17 ± 0.10 *,#
SYMPTOMATIC MODELS OF MS
Chronic EAE RM4(6 months )
Normal
Post Relapse-Remission
Spastic Limbs
Bladder Problems
KNOW YOUR MODEL-BIOLOGY OF EAE
PERIPHERAL IMMUNITY Slow RELAPSE RATE
NEURODEGENERATION Slow PROGRESSION
SYMPTOM CONTROL Improve QUALITY OF LIFE
REPAIR Reverse Deficits
OUTCOME OF CONTROLDISEASE PROCESS
Increasing Number of Effective Drugs
Effective Drugs with Unpleasant Side-Effects
No Treatments
No Treatments
Adaptive Immune –Dependent InflammationAdaptive Immune–Independent, Innate Dependent
Recently DemyelinatedChronic Demyelinated/Gliotic Lesions
KNOW YOUR MODEL-BIOLOGY OF EAE
• PRE-CLINICAL FAILURE• Model does not reflect human disease biology• Drug does not target biology relevant to human application• Lack of appreciation of human disease• Dogma & overstating effect • Model used in a way that does not reflect human indication• Drug doses are not used in at physiological doses• Drugs are not delivered in a way appropriate to how used in humans• Studies are not transparent & not reproducible (Ineffective Study Design)• CLINICAL FAILURE • Lack of clear understanding of human pathology• Drug is seldom investigated by scientists developing the Idea.• Over-interpretation of significance of pre-clinical studies• Drug is not used at a dose relevant to the pre-clinical studies• Population does not respond as predicted. (Ineffective Trial Design)• Dose-limiting side-effects• Study Underpowered, too short or unrealistic expectations• Measurement Instruments Inadequate Clinical Outcomes and Surrogate Markers• Wrong Group of pwMS studied (IneffectiveTrial Design)• Commercial Interests
Mechanism is all Important.
Relevance of Slight Delay of a Few Days, Slight Diminution
Prophylactic/Therapy
“Toxicity leading to Stress”
“Route & Timing”Reporting Issues
KNOW THE LIMITATIONS-FAILURE TO TRANSLATE
Two thousand drugs tested in EAE, only nine-ten classes of drugs approved
Time Post Induction (Days)
10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25
Mean
Clin
ica
l Sco
re
0.0
0.5
1.0
1.5
2.0
2.5
3.0
3.5
4.0Vehicle n= 10/10Test Drug n = 10/10 Test Drug 2 n=0/10
or n = 7/10
ARRIVE Guidelines (UK)Kilkenny C, Browne WJ, Cuthill IC, Emerson M & Altman DG. Improving bioscience research reporting: the ARRIVE guidelines for reporting animal research. PLoS Biol. 2010; 8(6):e1000412 (2010)
National Research Council (USA). Institute for Laboratory Animal Research. Guidance for the Description of Animal Research in Scientific Publications. National Academies Press, Washington (DC) (2011).
Guidelines on EAE StudiesBaker D, Amor S. Publication guidelines for refereeing and reporting on animal use in experimental autoimmune encephalomyelitis. J Neuroimmunol. 2012 242:78-83
LIMIT YOUR LIMITATIONS-EXPERIMENTAL DESIGN
RANDOMISATION
BLINDING
SAMPLE-SIZE
DATA HANDLING
REPLICATION
NINDS GUIDELINE Nature 2012; 490:187
>10%
>20%
>1%
ARRIVE Guidelines (UK)Kilkenny C, Browne WJ, Cuthill IC, Emerson M & Altman DG. Improving bioscience research reporting: the ARRIVE guidelines for reporting animal research. PLoS Biol. 2010; 8(6):e1000412 (2010)
National Research Council (USA). Institute for Laboratory Animal Research. Guidance for the Description of Animal Research in Scientific Publications. National Academies Press, Washington (DC) (2011).
Guidelines on EAE StudiesBaker D, Amor S. Publication guidelines for refereeing and reporting on animal use in experimental autoimmune encephalomyelitis. J Neuroimmunol. 2012 242:78-83
LIMIT YOUR LIMITATIONS-EXPERIMENTAL DESIGN
RANDOMISATION
BLINDING
SAMPLE-SIZE
DATA HANDLING
REPLICATION
NINDS GUIDELINE Nature 2012; 490:187
>10%
>20%
>1%
Parametric(47%)t- test(46%)
NonParametric
39%Not
Reported(14%)
Parametric(65%)
t test (67%)
Non-Parametric (4%)
NotReported
(31%)
All EAE Publications 6 months 1:12:2011-31:5:2012 n=175
All EAE Publications 2 Years1:1:2010-11:9:2012 n=26
All PUBMED PUBLICATIONS
BIG 6 PUBLICATIONSNature, Nat Med, Nat Immunol,
Nat. Neurosci, Science, Cell
T-test CriteriaContinuousNormally DistributedEqual Variances
LIMIT YOUR LIMITATIONS-EXPERIMENTAL DESIGN
Figure legend. Clinical scores of two independent EAE experiments at d23 post disease induction. Individual scores as well as the mean score of two independent experiments are shown. Control: n=10, vehicle: n=13, xxxxx-345: n=11. Control versus vehicle: P=0.620, control versus xxxxx-345: P=0.017, vehicle versus xxxxx-345 P=0.029. * indicate P values <0.05 and ** indicate P values <0.005 based on a non-paired Student’s t test. Error bars are s.e.m
Nature Paper 2015
Drug-treated animals the scores appear to be: 0, 0 ,0, 0.5, 0.5, 2, 2.5, 3, 3.5, 3.5, 3.5 n=11 Vehicle scores appear to be: 0.5, 2.5, 2.5, 2.5, 2,75, 2.75, 3, 3.5, 3.5, 3.5, 3.5, 3.5, 3.5 n=13.Do a t test drug verses vehicle p=0.029
The assumptions of a t test (a) Data is normally distributed. You test this and it passes the test p=0.152,
(b) data groups have equal variances Test for that and it fails P<0.05. (c) Data is Parametric. Fail. It is non-parametric.
t test is not a valid testMann Whitney test. P=0.082.........Ooooops.
LIMIT YOUR LIMITATIONS-EXPERIMENTAL DESIGN
• PRE-CLINICAL FAILURE• Model does not reflect human disease biology• Drug does not target biology relevant to human application• Lack of appreciation of human disease• Dogma & overstating effect • Model used in a way that does not reflect human indication• Drug doses are not used in at physiological doses• Drugs are not delivered in a way appropriate to how used in humans• Studies are not transparent & not reproducible (Ineffective Study Design)• CLINICAL FAILURE • Lack of clear understanding of human pathology• Drug is seldom investigated by scientists developing the Idea.• Over-interpretation of significance of pre-clinical studies• Drug is not used at a dose relevant to the pre-clinical studies• Population does not respond as predicted. (Ineffective Trial Design)• Dose-limiting side-effects• Study Underpowered, too short or unrealistic expectations• Measurement Instruments Inadequate Clinical Outcomes and Surrogate Markers• Wrong Group of pwMS studied (IneffectiveTrial Design)• Commercial Interests
Mechanism is all Important.
Relevance of Slight Delay of a Few Days, Slight Diminution
Prophylactic/Therapy
“Building Site Effect”
“Route & Timing”
“Placebo Effect”
Less Circuitry so Less Compensation Capacity
Non-Responders
Immune (T/B cell) or NeurodegenerationProfessional Trialists
Reporting Issues
Two thousand drugs tested only nine-ten classes of drugs approved
KNOW THE LIMITATIONS-FAILURE TO TRANSLATE
KNOW THE LIMITATIONS-FAILURE TO TRANSLATE
Once Drugs Become Available, Clinicians Voice their Opinion that Animal Experiments are not worth while, because of the failures
“It is now possible to conduct a Phase II trial for anti-inflammatory MS drug candidates within a few months…..it is not useful to pre-screen potentially effective drugs using the EAE model (Ransohoff 2006)”_______________________________________________________________________________________
EAE MS____________________________________________________________________________________________________MS DRUGSCopaxone Inhibition InhibitionBeta Interferon Inhibition InhibitionCD49d-specific mAb Inhibition InhibitionGilenya/Fingolimod Inhibition InhibitionAubagio/teriflunomide Inhibition InhibitionTecfidera/Dimethy Fumarate Inhibition InhibitionCD52-specific mAb Inhibition Inhibition(CD20-specific mAb) Inhibition Inhibition
/Context Dependent /Context Dependent /Worsening
/No effect/Worsening
FAILURESCD25-specific mAb Worsening InhibitionGamma Interferon Inhibition WorseningTNF-specific mAb Inhibition WorseningPDE4 inhibitor (rolipram) Inhibition No Effect/WorseningAntigen-Specific Treatments Inhibition No Effect/WorseningTh2 Cytokines (IL-4/IL-10) Inhibition No EffectCD4-specific mAb Inhibition No EffectIL-12/23-specific mAb Inhibition No Effect____________________________________________________________________________________________________
Treg cells important
Th1/Th17 cells important
If 65% T cell Depletion does not Stop EAE, Why would it be expected to stop MS?
The Trial in MS was doomed before it even started?
ACUTE EAE
IL-12/23 KO C57BL/6 EAE non-susceptible (IL12p40 + IL12p35 = IL-12)anti-IL23 (p40) C57BL/6 EAE inhibited (IL-12p40 + IL23p19 = IL-23)anti-IL23 (p40) ABH EAE inhibited
RELAPSING EAEanti-IL-23 (p40) ABH No inhibitory effect (Heremanns et al. 1999)
RELAPSING MSanti-IL23(p40) Human MS No inhibitory effect
Th17
IL-23IL-6/TGFb
Th1
IL-12
Th2
IL-4
B
IL-4, IL10, IL-13 IFNg
IL-4IL-10IL-13
IL-17IL-21IL-22
IFN-g
Ustekinumab(Segal et al 2008)
KNOW THE LIMITATIONS-FAILURE TO TRANSLATE
Time Post-inoculation (Days)
0 7 14 21 28 35 42 49 56 63 70 77Dev
elo
pm
ent
of
Clin
ical
EA
E (
%)
0
10
20
30
40
50
60
70
80
90
100
Untreated
CD4b mAb 3 weeks
CD4d&CD8d mAb D12 + CD4b mAb 3 Wks
“Anti-CD4 antibody Cures EAE”
KNOW THE LIMITATIONS-FAILURE TO TRANSLATE
Time Post-inoculation (Days)
0 7 14 21 28 35 42 49 56 63 70 77Dev
elo
pm
ent
of
Clin
ical
EA
E (
%)
0
10
20
30
40
50
60
70
80
90
100
Untreated
CD4b mAb 3 weeks
CD4d&CD8d mAb D12 + CD4b mAb 3 Wks
Anti-CD4 antibody Causes Transient Immunosuppression in EAE
KNOW THE LIMITATIONS-FAILURE TO TRANSLATE
• CD4 T cell depletion inhibits EAE
• CD4 T cell deletion inhibits virtually every T and B Cell mediated autoimmunity
• CD4 T cell deletion does not inhibit multiple sclerosis (Lindsey et al. 1994 Neurol 44:413 : 810; van Oosten et al. 1998 Mut Scler 1:339)
.
• CD4 T cell deletion does not inhibit other human autoimmune diseases
Problem with the StudyPlanned <60% DepletionNaïve CD45RA preferentially affectedPrimed CD45RO T cells relatively unaffected (Llewellyn-Smith et al. 1997 Neurol 48:810; 48)
MS IS NOT CONTROLLED BY T CELL DEPLETION
No effect on Gadolinium enhancing lesions
If 60% T cell Depletion does not Stop EAE, Why would it be expected to stop MS?
The CD4 trial in MS was Probably doomed before it even started
KNOW THE LIMITATIONS-FAILURE TO TRANSLATE
Degree of CD4 depletion was important with regard to treatment efficacy.There was a significant 41% reduction in relapses.
Red = abundant CB1 receptors Black = moderately abundant CB1 receptors
PHARMACEUTICAL CANNABIS
BENCH TO BESIDE – SYMPTOM CONTROL
Time Post-Injection (Min)
0 10 20 30 40 50 60 70 80 90 100 110 120
Cha
nge
in H
indl
imb
Stif
fnes
s (%
) ±
SE
M
-50
-40
-30
-20
-10
0
10
20
Sativex Vehicle
Red = abundant CB1 receptors Black = moderately abundant CB1 receptorsBENCH TO BESIDE – NEUROPROTECTION OR NOT
Placebos did not progress as predicted
Zajicek J et al. 2013
P<0.01
CUPID Cannabinoid Use in Progressive Inflammatory brain Disease.
Animal Studies show that Cannabinoids are Neuroprotective
Compounds in cannabisTHC & CBD causeNeuroprotection
Cannabis Receptor LossCB1 KO exhibitNeurodegeneration
Natural Ligand IncreaseFAAH KO loosesNeuroprotection
FAAH inhibitorsNeuroprotection
Red = abundant CB1 receptors Black = moderately abundant CB1 receptorsBENCH TO BESIDE – NEUROPROTECTION OR NOT
Placebos did not progress as predicted
Zajicek J et al. 2013
P<0.01
Ambulatory PeopleStarting EDSS less than 5.5
CUPID Cannabinoid Use in Progressive Inflammatory brain Disease.
Animal Studies show that Cannabinoids are Neuroprotective
Compounds in cannabisTHC & CBD causeNeuroprotection
Cannabis Receptor LossCB1 KO exhibitNeurodegeneration
Natural Ligand IncreaseFAAH KO loosesNeuroprotection
FAAH inhibitorsNeuroprotection
INFLAMMATORY PENUMBAAnimal Studies indicate that sodium channel blockers can be neuroprotective by: •Inhibiting metabolic overload in nerves •Blocking microglial activity
Lamotrigine trial in secondary progressive MS fails Brain atrophy MRI shows (pseudo) atrophy greater than placebo. Drug was poorly tolerated (Kapoor et al. 2010).
50% of people in trial are not drug compliant.Neurofilament Biomarker is reduced in DrugCompliant individuals (Gnanapavan et al. 2013)
Animal studies show that sodium channel blockers are particularly active during the inflammatory penumbra. (Al-Izki et al. 2014)
PLP (brown) myelin Stain)
Red = abundant CB1 receptors Black = moderately abundant CB1 receptorsBENCH TO BESIDE – NEUROPROTECTION
Optic Neuritis isOften First Sign of MS
Most accessible part of Human CNS
Courtesy of Roy Weller
Red = abundant CB1 receptors Black = moderately abundant CB1 receptorsBENCH TO BESIDE – NEUROPROTECTION
EYE
OPTIC NERVE
VISUALCORTEX
The eye is the window to the brain…
OCCIPITALLOBE
The Visual SystemX
Damage in MS
Optic NeuritisCommon First
Sign of MS
Occurs in Over50% of MS
X
Red = abundant CB1 receptors Black = moderately abundant CB1 receptorsBENCH TO BESIDE – NEUROPROTECTION
EYE
OPTIC NERVE
VISUALCORTEX
The eye is the window to the brain…
OCCIPITALLOBE
The Visual SystemX
Damage in MS
Optic NeuritisCommon First
Sign of MS
Occurs in Over50% of MS
X
Red = abundant CB1 receptors Black = moderately abundant CB1 receptorsBENCH TO BESIDE – NEUROPROTECTION
Red = abundant CB1 receptors Black = moderately abundant CB1 receptorsBENCH TO BESIDE – NEUROPROTECTION
C57BL/6-Tg(Tcra2D2,Tcrb2D2)1Kuch /J
Red = abundant CB1 receptors Black = moderately abundant CB1 receptorsBENCH TO BESIDE – NEUROPROTECTION
C57BL/6-Tg(Thy1-CFP)23Jrs /J
Develops Spontaneous/Induced Optic Neuritis.Subclinical Spinal Cord DiseaseLess Severe than Classical EAE
Fluorescent RGC Detection of Nerve Loss in Living Eye
Repeated Monitoring Not Requiring Histology
Disease is Concentrated in the Visual SystemHuman Relevant Outcome Measures
Red = abundant CB1 receptors Black = moderately abundant CB1 receptorsBENCH TO BESIDE – NEUROPROTECTION
C57BL/6-Tg(Tcra2D2,Tcrb2D2)1Kuch.Cg-Tg(Thy1-CFP)23Jrs /J = FLASH GORDON
Low Contrast Eye Chart
Human Test
Visual acuity is a measure of clearness of vision and is used as a measure as to how you can see
Mouse Test
Visual tracking drum
Net
pos
itive
hea
d m
ovem
ents
0
2
4
6
8
10
12
14
Before Optic
Neuritis
AfterOptic
Neuritis
Loss of vision
Head movementsare Reduced after
Optic Neuritis(Nerve Loss)
Red = abundant CB1 receptors Black = moderately abundant CB1 receptorsBENCH TO BESIDE – NEUROPROTECTION
MEASURING SIGHT-VISUAL ACUITY
Reference electrode
Recording electrode
Heating pad
Visual Evoked Potential (VEP)
Z ZZ
Electrophysiology can be used to measure the Visual Evoked Potential (VEP). This
is a measure of the neurotransmission from the
eye to the visual cortex.
Human Test
Mouse Test
Latency
Latency of VEPis Increased after
Optic Neuritis(Demyelination)
BeforeOptic
Neuritis
AfterOptic
Neuritis
Loss ofNerve
Conduction
Amplitude
Amplitude of VEPis Reduced after
Optic Neuritis(Nerve Loss)
Ampli
tude
(µV)
0
10
20
30
40
BeforeOptic
Neuritis
AfterOptic
Neuritis
Loss of vision
Red = abundant CB1 receptors Black = moderately abundant CB1 receptorsBENCH TO BESIDE – NEUROPROTECTION
MEASURING SIGHT-ELECTROPHYSIOLOGY
Optic Nerve HeadGanglion CellLayer (GCL)
OCT is a non-invasive retinal imaging tool to look at the structure of the retina.
Human Test Mouse Test
Optic Nerve Head
OCT imageHistology image
Eye from Living AnimalEye from Dead Animal
OuterNuclear Layer
Photo-Receptors
Ganglion Cell Layer
Inner Nuclear Layer
Red = abundant CB1 receptors Black = moderately abundant CB1 receptorsBENCH TO BESIDE – NEUROPROTECTION
OPTICAL COHERENCE TOMOGRAPHY (OCT)
Before Induction
After Induction
Optic Nerve Head
Optic Nerve Head
OCT scan
OCT Can Detect Loss of RGC in Mouse with Optic Neuritis
Red = abundant CB1 receptors Black = moderately abundant CB1 receptorsBENCH TO BESIDE – NEUROPROTECTION
OPTICAL COHERENCE TOMOGRAPHY (OCT)
cSLO is a high resolution retinal imaging tool to look at the structure of the retina
Optic NeuritisPre-Disease
After Disease Onset
Retinal Ganglion Cell Loss
Before Disease Onset
Retinal Ganglion Cell Loss
cSLO Image
BloodVessel
Optic Nerve Head
Retinal GanglionCell
Optic Nerve Head
Pre-Disease 2018 cells/mm2
Optic Neuritis 285 cells/mm2
RGCLoss
P<0.001
Red = abundant CB1 receptors Black = moderately abundant CB1 receptorsBENCH TO BESIDE – NEUROPROTECTION
Confocal SCANNING LASER OPHTHALMOSCOPY (cSLO)
Day 0 2 10 14 21
Demyelinating mAb(250μg Z12 mAb i.p.)
Disease Induction (150ng Pertussis Toxin i.p.)
T cell-Mediated Optic Neuritis
Retinal Flatmount
Microscopy
Treatment Period 5mg/kg i.p.)
Optic Neuritis
A
D
CBA
D
CB
Vehicle
Sodium ChannelBlocker
A
D
CB
Low PowerNormal
Reduced Nerve Damage
Normal mouse
Mea
n re
tina
cell
dens
ity (c
ells
/mm
2)
1000
1100
1200
1300
1400
1500
1600
1700
1800
1900
OPTIC NEURITIS+ Vehicle
OPTIC NEURITIS +drug
Red = abundant CB1 receptors Black = moderately abundant CB1 receptorsBENCH TO BESIDE – NEUROPROTECTION
NEUROPROTECTION WITH SODIUM CHANNEL BLOCKERS
OPTIC NEURITIS
Gabilondo I et al. 2015
Change in Retinal Nerve Fibre Layer Thickness after Optic Neuritis
Double blind, randomised placebo-controlled, parallel group design
Initial dose 15 mg/kg, daily 4 mg/kg (max 300mg) , treatment duration 3 monthsBlinded assessing and treating physicians; 2 imaging sites (Sheffield, London)Outcomes OCT, VEP, MRI, Vision
Red = abundant CB1 receptors Black = moderately abundant CB1 receptorsBENCH TO BESIDE – NEUROPROTECTION
NEUROPROTECTION WITH SODIUM CHANNEL BLOCKERS
OPTIC NEURITIS TRIAL
Red = abundant CB1 receptors Black = moderately abundant CB1 receptorsBENCH TO BESIDE – NEUROPROTECTION
NEUROPROTECTION WITH SODIUM CHANNEL BLOCKERS
300 pwMS
Year 1 Year 2 Year 3
600 pwM
300 pwMS
Active tablet
Placebo tablet
Year -2
Year 4
Month6
Active tablet
Placebo tablet
-6month
month 18
Month12
LP1 LP2 LP3
STANDARD TRIAL DESIGN
NOVEL TRIAL DESIGN
30 pwMS
30 pwMS
60 pwMS
Red = abundant CB1 receptors Black = moderately abundant CB1 receptorsBENCH TO BESIDE – NEUROPROTECTION
NEUROPROTECTION WITH SODIUM CHANNEL BLOCKERS
DMT + Nerve protector
DMT
UCL-INSTITUTE OF NEUROLOGYQueen SquareUCL-INSTITUTE OF NEUROLOGYQueen SquareUCL-INSTITUTE OF NEUROLOGYQueen Square THANK YOU FOR LISTENING
Spasticity StudiesDavid Baker Gareth Pryce Gavin Giovannoni
Neuroprotection StudiesDavid Baker Gareth Pryce Sarah Al-IzkiKatie LidsterSam JacksonGavin Giovannoni
Optic Neuritis TrialRaj Kapoor, Rhian Raftopoulos, Simon Hickman, Basil SharrockKlaus Schmierer, Gavin GiovannoniDavid H Miller & Others
Slides Available on www.ms-res.org(Slideshare)