Drug Development

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Protein-protein interaction disruptors as
therapeutic targets
(an example of a drug discovery project)
Yvonne Lai, Ph.D.
Department of Psychological and Brain Sciences
Indiana University, Bloomington
Navigator, Indiana CTSI (Bloomington campus)
NMDAR Signaling Cascade
Image from Cayman Chemical
Protein-protein interaction disruptors of
NMDAR-dependent signaling
Tat-NR2B9c (NA-1)
Peptide inhibitor
Pain behavior
NMDAR -----------PSD95----------nNOS-------------NOS1AP--------MAPK------PTSD
(CAPON)
Depression
Stroke
Small molecule
inhibitors:
IC87201/ZL006
Drug Development
Target
Discovery
Lead
Discovery
Assay
development
High-throughput
screen
Lead
Optimization
Candidate
selection
Preclinical
Evaluation
Full
Development
Clinical Trials
Current collaborative Team
Pain efficacy:
Andrea Hohmann (IUB)-Wan-hung Lee
Neurotoxicity:
Andy Hudmon (IUSM)-Aarti Chawla
PTSD efficacy:
Anantha Shekhar (IUSM)-Stephanie Fitz
Chemistry:
Ganesh Thakur (Northeastern University)-Pushkar Kukarni
nNOS-NOS1AP disruption
Michael Courtney (University of Southern Finland)-Lilli Li
Small molecule nNOS-PSD95 protein-protein interaction inhibitors
IC87201:
• Inhibited NMDA-induced nNOS-dependent NO formation
• Efficacious in chronic pain model (Florio, BJP 2009; Lai, Hohmann
unpublished)
• Decrease symptoms of PTSD in a rat preclinical model (Shekhar,
unpublished)
• Efficacious in one depression model (Doucet 2013)
ZL006 (analog of IC87201):
• Blocked focal cerebral ischemic damage in mice and rats (Zhou, Nat Med
2010)
• Efficacious in one depression model (Doucet 2013)
Model
Neuronal cells
Cerebral Ischemia
Neuropathic pain
Fear conditioning
Plasma
Side effect Profile
NOS enzyme
Spatial memory
Motor movement
Effect Measured
Cytotoxicity in vitro
cGMP (NO marker)
Infarct volume
IC87201 (~EC50)
<0.5 mM
3 mM
N.D.
ZL006 (~EC50)
0.08 mM
N.D.
1.5 mg/kg (iv)
Tactile allodynia
Freezing time
Drug level (rat)
2 mg/kg (ip)
4 mg/kg (ip)
6mM-2mg/kg (i.p.)
N.D.
10 mg/kg (ip)
9mM-1.5mg/kg (i.v.)
Catalytic activity
Morris water maze
Rotarod
No effect (10 mM)
N.D.
No effect (4 mg/kg)
No effect (10 mM)
No effect (10 mg/kg)
No effect (10 mg/kg)
Additional profile of IC87201/ZL006
• Efficacious systemic or i.t., found in CSF and brain tissues
• No significant inhibition of 35 target (Receptorgram, MDS Pharma)
• No effect on basal pain sensation
• No motor ataxia
• No effect on novel object recognition
• No effect on Morris water maze
• No effect on other PDZ P-P interactions (e.g. Erb4-PSD95)
HTS (NIH clinical library)
NGP-202-01 (2966)
NOS-NOS1AP
100
50
NOS-PSD95
NOS-NOS1AP
150
100
%Control
%Control
NGP-105-01(3281)
NOS-PSD95
150
50
0
0
0
20
40
60
80
0
20
Well #
NGP-202-02 (3055)
Target
Discovery
Lead
Discovery
NGP-105-02 (3361)
NOS-PSD95
NOS-NOS1AP
100
150
60
80
50
NOS-PSD95
NOS-NOS1AP
100
%Control
%Control
150
40
Well #
50
0
0
0
20
40
60
80
0
20
Well #
NGP-202-03(3144)
150
40
60
80
Well #
NGP-105-03 (3441)
NOS-PSD95
NOS-NOS1AP
200
NOS-PSD95
NOS-NOS1AP
150
%Control
%Control
100
50
Assay
development
50
0
0
0
20
40
60
80
0
20
40
60
80
Well #
NGP-202-04 (3233)
NGP-105-04 (3521)
NOS-PSD95
NOS-NOS1AP
150
100
%Control
%Control
150
High-throughput
screen
100
50
0
NOS-PSD95
NOS-NOS1AP
100
50
0
0
20
40
Well #
60
80
0
20
40
60
Well #
Alpha Screen using NIH clinical library (~800 cpds); nNOS-PSD95 vs nNOS-NOS1AP (Fan,
Courtney, Hohmann, Lai, unpublished)
80
Virtual screen?
Target
Discovery
Lead
Discovery
Virtual Screen
Model of PSD95 PDZ2 and the nNOS PDZ complex (PDZ2 is superimposed on the α-syntrophin
structure. From Tochio 2000)
Drug Development
Preclinical
Evaluation
Side effect
profile
What are the critical No Go side effects for treatment
of pain and PTSD?
Additional:
• Memory models?
• Toxicity profile?
• Oral bioavailability?
Drug Development
CTSI Core Facilities
Target
Discovery
Lead
Discovery
Lead Optimization
Preclinical
Evaluation
HTS: Chemical Genomics
IUB-Light Microscopy Imaging Center (high content screen)
Need for library:
For unusual targets (e.g. P-P interactions)
For cell based or more complicated assays
Computational modeling
Virtual screening
Chemical design and synthesis
Full
Development
Funding
CTSI Core facility grant (collaboration with Purdue’s BAL)
IURCG collaborative grant (Hohmann, IUB and Hudmon, IUSM)
NIH likes our innovative approaches:
• Funded:
o R21/NIDA (pain efficacy)-Hohmann PI; Lai, Co-PI
• Expected to be funded:
o R21/NIMH (PTSD)-Shekhar, PI; Lai, Co-PI
o SBIR phase I (two years)-Lai, PI
What other funding resources? VC, Angel funds?
Collaboration with industry?
Collaboration between start ups with academic universities:
What is the model? How to optimize expertise and minimize
duplications?
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