Disorders of Cholesterol Homeostasis
Niemann-Pick disease, type C
• Autosomal recessive, progressive,
lethal, neurodegenerative disorder
due to mutation of either NPC1
or NPC2
• Endolysosomal storage of
unesterified cholesterol and
lipids
• Incidence: 1/100,000-120,000
• No FDA approved therapies
Niemann-Pick Disease, type C1
• Variable phenotype and age of onset
– Classical Disease: Late-infantile and juvenile (60-70%)
– Infantile (20%)
– Adult
• Late-infantile and juvenile (classical)
– Transient neonatal jaundice
– Splenomegaly
– Neurological symptoms (insidious onset)
•
•
•
•
•
Vertical supranuclear ophthalmoplegia
Cerebellar ataxia and dysfunction
Cognitive impairment and dementia
Gelastic cataplexy
Seizures
Niemann-Pick Disease, type C1
• Therapeutic trial issues
– Rare Disease
– Heterogeneous phenotype
• Variable age of onset
• Variable symptom complex
– Clinical progression occurs over years
– Goal: Stabilization or delay of neurological disease
– Optimal treatment may be prior to the onset of
neurological signs and symptoms
– Significant diagnostic delay
– Lack of defined and accepted outcome measures
Niemann-Pick disease, type C1
• 78 patients enrolled since August 2006
– Age range: 3 months to 54 years
(median 10 years)
– NPC1 Neurological Severity
(range 0-50, median 14)
– Miglustat therapy (42%)
• Goals:
Severity Score
• Natural History Trial
40
20
0
10
20 30 40
Age (years)
– Identify clinical or biochemical markers that can be used as an outcome
measure in a therapeutic trial
– Identify a biochemical marker that can be used for diagnostic testing or
screening
50
Niemann-Pick Disease, type C1
Mean Diffusivity
Volume
Niemann-Pick Disease, type C
Biomarker Development
(Assembling a Tool Box)
Niemann-Pick disease, type C1
• Biomarkers
– Insight into pathology
– Diagnostic/screening test
– Tools to guide therapeutic trials
• Biomarker identification
–
–
–
–
Candidate proteins/lipids
Multi-analyte profiling
Expression analysis
Proteomics
Oxysterols
Blood-based diagnostic test
Niemann-Pick Disease, type C
• Filipin Staining
– Fluorescent antibiotic that binds
unesterified cholesterol
– Specialized testing
– Requires a skin biopsy-invasive
– Variable staining
Filipin Staining
Control
NPC
Vanier and Millat (2003) Clin Genet. 64: 269-281
• 80-85% “classical”
• Molecular Testing
– Expensive and requires a high
index of suspicion
– ~80% sensitive
• 4-5 year diagnostic delay
Dan Ory (personal communication)
NPC1: Clinical Science
• Oxysterols
– Hypothesis: In NPC1 the unique combination of
increased oxidative stress and intracellular
accumulation of unesterified cholesterol will
result in increased nonenzymatic oxysterols.
Intracellular Cholesterol
Accumulation in NPC1
Decreased Serum Antioxidant
Capacity in NPC1 Subjects
TEAC (mM)
10
8
6
n=40
p<0.0001
4
2
0
n=40
Control
NPC
Fu et al (2010) MGM, 101:214
Niemann-Pick Disease, type C1
Plasma Oxysterols
3β,5α,6β-cholestane-triol
7-ketocholesterol
3β,5α,6β-cholestane-triol
7-ketocholesterol
Porter et al. (2010) STM, 2: 56ra81
Niemann-Pick Disease, type C1
Triol (24.5 ng/ml)
Sensitivity 97%
Specificity 100%
Porter et al. (2010) STM, 2: 56ra81
Jiang et al. (2011) JLR, 52:1435
Niemann-Pick Disease, type C1
CSF Protein Biomarkers
Niemann-Pick Disease, type C1
+/+
Calbindin ng/mL
Calbindin D
20
15
p<0.001
10
5
0
-/-
Control
NPC1
Calbindin (ng/mL)
20
15
p=0.28
10
5
0
Control Untreated Treated
Niemann-Pick Disease, type C
Calbindin Percent Change
+/- miglustat
Serial Calbindin pre/post miglustat
30
20
Percent Change
15
10
5
10
0
-10
p<0.05
-20
-30
n=16,15,5
A
B
Miglustat
C
D
e-
ed
ea
t
Months
pr
-50
30
tr
20
d
10
ea
te
0
po
st
-40
0
un
tr
Calbindin ng/mL
20
Untreated
Treated
Pre/post
B/A
D/C
C/B
Niemann-Pick disease, type C1
• Fatty Acid Binding Protein 3 (FABP3)
62
62
59
59
Npc1+/+
Npc1-/p<0.0001
CSF
Percent change
FABP3
FABP3 (ng/ml)
100
40
80
p<0.0001
60
40
20
0
Controls
NPC
p=0.05
20
0
-20
untreated
treated
-40
-60
-80
pre/post
miglustat
Niemann-Pick disease, type C1
• Macrophage Inflammatory Protein 1-alpha (Mip1α, CCL3)
– Proinflammatory cytokine
– Activated microglia
30
CSF
Mip1alpha (pg/mL)
Mip1 (pg/mL)
30
p<0.0001
20
10
0
25
20
15
10
5
0
Control
NPC
0
10 20 30 40 50 60
Severity Score Ranking
Control 4.8 ± 1.9 pg/ml
NPC
14.7 ± 4.2
Confidential
CSF Biomarkers
• CSF Biomarkers
– Lipid-based
• 3β,5α,6β-cholestane-triol and cholesterol esters
– Protein-based
• Markers of inflammation
– MIP-1α, IL1a, IL3, IL4, IL12p40, IL13, IL15, and MMP2
• Markers of neuronal injury
– Aβ(42), phosphorylated tau, fatty acid binding protein 3 (FABP3),
and calbindin-D
• Markers of oxidative stress
– glutathione S-transferase alpha (GSTα) and
superoxide dismutase 1 (SOD1)
2-Hydroxypropyl-β-cyclodextrin
NPC1 HPβCD Trial
• 2-Hydroxypropyl-β-cyclodextrin (HPβCD)
– Cyclic oligosaccharide with a hydrophobic core
– Pharmaceutical excipient
• Drug repurposing
NPC1 HPβCD Trial
• Npc1 mouse model
Survival
Ramirez et al. (2010) Ped. Res. 68: 309
Cerebellar pathology: 49-days
Npc1+/+
Npc1-/- Npc1-/- + CD
Liu et al. (2009) PNAS 106: 2377
NPC1 HPβCD Trial
• Npc1 cat model
– 24 week Npc1 mutant cats (HPβCD, miglustat, untreated)
Personal Communication: Charles Vite
NPC1 HPβCD Trial
• HPβCD Therapeutic trial
– Pilot project for TRND/NCATS
• Goal: Improve drug development for rare and neglected
diseases
– HPβCD does not cross the blood-brain barrier
– Dose limiting toxicity: Ototoxicity
• Cat, dog, and mouse
NPC1 HPβCD Trial
Lancet (1963) 282: 983-984
Risks
Bleeding/strokes
Infection
Seizure
NPC1 HPβCD Trial
• Phase 1 trial
– Goal: Establish a safe and biochemically effective
dose
– Cohort dose-escalation
• Safety
• Pharmacokinetics
– Quantitative and validated assay for HPβCD
• Pharmacodynamics (Biochemical efficacy)
– Quantitative and validated assays for 24-hydroxycholesterol
• Pathological efficacy
– CSF protein and lipid biomarkers
NPC1 HPβCD Trial
• Pharmacodynamic response
• Plasma 24(S)-hydroxycholesterol
Neuron
Cholesterol
Synthesis
ApoE
NPC1
HPβCD
CYP46
24(S)HC
CSF
24(S)HC
Blood
NPC1 HPβCD Trial
• Pharmacodynamic response
• Plasma 24(S)-hydroxycholesterol
Neuron
Cholesterol
Synthesis
ApoE
NPC1
HPβCD
CYP46
24(S)HC
CSF
24(S)HC
Blood
NPC1 HPβCD Trial
Saline
CSF and Plasma Sampling
(0 to 72 hours)
HPβCD
CSF and Plasma Sampling
(0 to 72 hours)
• Baseline standard deviation of the saline area under the
curve (AUC) estimated by resampling (bootstrapping)
• Biochemical response defined as:
– In an individual patient the difference between the drug and
saline AUC greater than 2.3 x SD is considered a positive
response
– Positive response observed in at least 2 out of 3 patients
NPC1 HPβCD Trial
• 50 mg HPβCD
– No drug related adverse events or reactions
– AUC8-72
• Patient 1 + 3.1 x sd
• Patient 2 + 0.8 x sd
• Patient 3 + 4.1 x sd
• P. acnes
– Infection/Colonization
Plasma 24-OH Cholesterol
(ng/ml)
• 2/3 patients had a biochemical response
Saline
50 mg
35
30
25
0
12
24 36 48
Time (hours)
60
72
Niemann-Pick disease, type C1
• ICV versus lumbar IT
• Dose escalation
response
• Repetitive dosing
response: Attenuation
• Response of other
biomarkers
• Safety-Ototoxicity
Cohort Dose-escalation
Not Tolerated
Cohort 1
50 mg Dose
STOP
Not Tolerated
Tolerated
No Response
Cohort 3
Cohort 1/2
200 mg Dose
Cohort 1/2/3
300 mg
Tolerated
Biochemical Response
Tolerated
Biochemical Response
Tolerated
No Response
Cohort 3/4
400 mg
Cohort 1/2
Maintain
200 mg
Maintain
300 mg
Maintain
400 mg
Niemann-Pick disease, type C1
• No significant response at 50 and 200 mg IT
• Positive response in 2/3 subjects at 300 mg IT
– Response similar to 50 mg ICV
– Grade 1 Ototoxicity in 2 subjects (siblings)
• 400 mg cohort completed this week
Niemann-Pick disease, type C1
• Clinical efficacy trial
• Multicenter: Funding and IRB complexity
• Multinational: Funding, IRB and regulatory complexity
• NeuroNext Network
– Concept proposal accepted
• Clinical outcome measures
– Ataxia (Cerebellar Function)
• Gross motor
• Fine motor
– Swallowing
26 US Sites
Common IRB
Common Trial Agreement
NPC1 HPβCD Trial
• HPβCD selected as a TRND clinical candidate in February 2011
• PreIND meetings with FDA in October and December 2011
• Juvenile dog toxicity study, formulation, and device compatibility
studies performed in 2012
• IRB approval July 2012 and IND filing November 2012
• First ICV infusion February 4, 2013
• First IT infusion September 28. 2013
NPC1 TRND Team
NICHD
Nicole Yanjanin
Aiyi Liu (DESPR)
Roopa Shankar
NHGRI
Washington University School of Medicine
Daniel Ory
University of Pennsylvania
Charles Vite
Bill Pavan
NINDS
Russell Lonser
John Heiss
Albert Einstein Collage of Medicine
Steven Walkley
TRND/NCATS
Johnson and Johnson
Chris Austin
John McKew
Nuria Carrillo
Liz Ottinger
Juan Marugan
Pramod Terse
Xin Xu
Wei Zheng
Steven Silber
Mark Kao
Marcus Brewster
SAIC/Leidos
Jon Stocker
Molly Buehn
Leah Giambarresi
RRD International
Charles Finn
Frank Hurley
Joy Vanderwal
Clinical Center
Carmen Brewer Beth Solomon
Naomi O’Grady Kelly King
Patrick Frenchick
Sandra Morseth
Kimberly Lilly
Acknowledgements
SMD, PDEGEN, NICHD
Nicole Yanjanin
Lee Ann Knight
Roopa Shankar
Chris Wassif
Celine Cluzeau
Stephanie Cologna
Ryan Lee
Cynthia Toth
Jiang Xiao-Sheng
Ian Williams
Antony Cougnoux
Ara Parseghian Medical Research Foundation
National Niemann Pick Disease Foundation
ORD/CC
SOAR-NPC
Bench to Bedside Awards
Dana’s Angels Research Trust
Niemann-Pick disease, type C1
Niemann-Pick disease, type C1
Hydroxypropyl-β-Cyclodextrin
Mouse
Cat
Mouse
Cat
Human
Newborn
0.5 g
30.0
1400.0
375.0
Niemann-Pick Disease, type C1
Vertical supranuclear ophthalmoplegia
Differential
-Niemann-Pick Disease, type C1
-Progressive Supranuclear Palsy
-Midbrain lesions
Niemann-Pick disease, type C1
Attenuation of the 24OHC
response with repetitive dosing