Allosteric Small Molecule Inhibitors of the NGF/TrkA Pathway
A New Approach to Treating Inflammatory Pain
Steven W. Andrews, Ph.D.
Associate Director of Drug Discovery
1
Challenges Faced with Discovery of New Drugs in the Pain Space
Translation of pre-clinical results to clinical outcome
Efficacy
Poor translation from animal models to man and vice versa
Safety
Mechanism specific toxicities are not always apparent until reaching the clinic
Bar for safety for new pain drugs is very high
Economic Considerations
While existing SOC have deficiencies and liabilities, these treatments are quite cost effective
and offer many convenient dosage forms
Our approach: small molecule inhibitors for antibody validated pathways
AR-797 P38 inhibitor – targeting NSAID resistant pain (Phase II)
Trk inhibitors – targeting the NGF pathway (pre-clinical)
2
Tanezumab- anti-NGF Antibody has Validated Clinical Efficacy
Lane et al. (2010) NEJM 363:1521
Katz et al. (2011) Pain 152:2248
Similarly impressive efficacy reported in Chronic Low Back Pain2
but concerns of joint findings in OA patients with chronic dosing, particularly with NSAIDs
Key Question- is there an alternative and perhaps safer
approach to inhibit this pathway
3
Neurotrophin / Trk Signaling Mediates Peripheral Pain Response
Local release
Peripheral
Hypersensitization
NGF
TrkA
Proinflammatory cell
Central
recruitment and degranulation
Hypersensitization
Neurotrophin Receptors- TrkA and B are critical
signaling partners in the NGF pain cascade
4
Mechanisms for Inhibiting Neurotrophin / Trk Signaling Cascade
Growth Factor Antibodies
NGF
BDNF
NT-4
NT-3
C1
LRR1-3
C2
Ig1
Trk Receptor Antibodies
Ig2
Trk kinase domain inhibitors
CR1
CR2
CR3
CR4
ATP site
100% homology in ATP site
Trk A
TrkB
TrkC
Our ATP site inhibitors selectively inhibit the pan-Trk axis, but not other kinases
5
P75NTR
Key Scientific Questions- Small Molecule Pan-Trk Kinase vs. anti-NGF
Efficacy for Pain Relief?
•
Added efficacy from the TrkB component?
•
Differences related to mechanism of inhibition?
•
Differences related to duration of inhibition?
– Long term vs intermittent target knockdown?
Safety?
•
Differences related to mechanism of inhibition?
•
Differences related to duration of inhibition?
– Long term vs intermittent target knockdown?
•
Safety concerns for added TrkB and TrkC inhibition?
Our approach: in vivo evaluation
with selective small molecule pan-Trk inhibitors –
6
Approach to Finding Highly Selective Trk Chemical Matter
High Throughput Screen
ARRAY diversity
and kinase focused
Trk
potency
Promising
Kinase
<50 compounds
~many compound hits
in 2 novel chemical series
in > 20 chemical series
Selectivity
and
“drug-likeness”
DMPK
Pharmacology
Toxicology
Medicinal Chemistry
Optimization
>1500 designed
compounds
pan-Trk series (ATP site)
7
Trk X-ray
Crystallography
•
Tuned for high kinase selectivity outside of Trk family
•
equipotent for TrkA, TrkB and TrkC
~80TrkA ATP site
x-ray structures
solved to date
Select Properties of Pan-Trk Leads
Program
8
Pan-Trk
Lead
ARRY-470
AR-772
AR-523
hTrkA cell IC50
9.7 nM
1.6 nM
10 nM
hTrkA free cell
IC50
23 nM
12 nM
42 nM
hTrkB / hTrkC
cell
24 nM
1.6 nM
12 nM
230 member
Kinase Panel
Clean
@ 1 µM
Clean
@ 1 µM
Clean
@ 1 µM
Predicted
hepatic Cl
Human, Rat
10, 18
(med, low)
13, 32
(med, med)
8, 13
(med, low)
Plasma protein
binding
Human, Rat
68%, 82%,
79%, 79%,
75%, 89%,
Solubility
(ng/mL)
pH 1.2 / 6.5 / 7.4
>1000, >1000,
>1000
>1000, 780,
820
>1000, 780,
820
peripheral to
CNS exposure
16 to 1
25 to 1
28 to 1
Very potent for inhibiting
neurotrophin driven
TrkA, TrkB and TrkC
signaling in cell
High kinase selectivity
Peripherally Selective:
only peripheral
inhibition in efficacy
studies
Kinase Selectivity of AR-470 compared to literature pan-Trk Kinase Inhibitors
Amino pyrimidine
ATP site
Diaryl urea
Type II DFG out
At 1 µM
Potent on Trks
Plus 83 off targets
at 0.5 µM
Potent on Trks
Plus ~7 off targets
No activity at 10 µM
against other pain targets:
AR-470
ATP site
Pan
TRK
inhibitor
Clean pharmacology
for inhibiting
TrkA, B and C
at1 µM
Potent on Trks
One weak off target
9
Summary of in vivo Efficacy Observed for Pan-Trk Leads
Excellent efficacy is observed in pre-clinical models of inflammatory pain
Acute Pain
 UV burn model (thermal hyperalgesia)
 CFA paw model (thermal hyperalgesia, gait analysis)
 CFA joint model (gait analysis)
 Fracture pain (flinching and guarding)
 Bone cancer pain (flinching, guarding and nerve budding)
 Surgical Incision
Chronic Pain
 CIA Model of rheumatoid arthritis (pain and histological evaluation)
 MIA Model of osteoarthritis (pain)
 CFA paw model (mechanical allodynia)
10
ARRY-470 Broad Efficacy Observed in Multiple Pre-Clinical Models
Mantyh et al. Molecular Pain 6 , art. no. 87
p
p
Mantyh et al. Bone 48 (2) , pp. 389-398
vehicle
ARRY-872
ARRY-470 (3 mg/kg)
ARRY-470 (10 mg/kg)
ARRY-872
ARRY-470 (30 mg/kg)
ARRY-872
ibuprofen (100 mg/kg)
11
ARRY-470 is Superior to NSAIDs in the CFA Joint Model
Results shown 2-3 days
after start of treatment
300
200
100
0
Naive
(
Guarding index
400
Vehicle
3
10
30
Dose (mg/kg p.o. twice daily)
Guarding index
Valdecoxib
Naproxen
Rofecoxib
400
400
400
300
300
300
200
200
200
100
100
100
0
0
Naive
Vehicle
10
30
100
Naive
Vehicle
30
100
300
Dose (µMol/kg p.o. twice daily)
12
0
Naive
Vehicle
7.5
30
90
Safety of Pan-Trk Inhibitors – Neuronal Safety
 No changes in functional observations in mice, rats, or monkeys at
therapeutic doses / exposures
 No histological changes in peripheral neuronal density in brain, spinal cord,
sciatic nerve or skin neurons to 300 mg/kg with 28 days of dosing
 No changes in normal pain response at therapeutic doses
Hot Plate (Thermal: C-fibers)
Day 48 post fracture
Hindlimb withdrawl response time at 55C (sec)
Von Frey (Mechanical: A-delta fibers)
Day 48 post fracture
6
50% Withdrawl latency (g)
5
4
3
2
1
0
Fracture + vehicle
n=6
FRX + 30mg/kg AR470
n=3
20
15
10
Col 3
Col 3
5
0
Fracture + vehicle
FRX + 30mg/kg AR470
n=6
n=3
Mantyh et al. Bone 48 (2) , pp. 389-398
No observed effects on neuronal safety
13
Safety of Pan-Trk Inhibitors – On Target Effects
Hyperphagia / weight gain
•
Increased food consumption –
peripheral effect
•
Increase weight gain- even when
food consumption is controlled
•
Likely BDNF / TrkB effect –
rodent specific?
Unger et al. JNEUROSCI, 2007, 27(52): 14265
Lin JC, et al. PLoS ONE 3(4): e1900 (2008)
Ataxia Scoring System
Dose of PanTrk
inhibitor
Ataxia Score
(max)
Incidence
30 mg/kg
1
1 of 3
100 mg/kg
1
3 of 3
4 - plus falling over when on hind legs, sleeping on back
300 mg/kg
2
3 of 3
Correlates with pan-Trk target coverage in the CNS
1 -"swimming” through litter, flattened or splayed on cage bottom
2 - head bobbing, jittery or hyperactive, head bobbing, nervous
3 - head rearing, disoriented, lethargy, agitated
Potential narrow therapeutic window for broad clinical pain treatment
14
Small Molecule Pan-Trk Leads Efficacy
 Great efficacy across pre-clinical pain models
 Equivalent to historical anti-NGF in the same models
 Intermittent target knockdown is sufficient for efficacy
 Partial pathway knockdown is sufficient for efficacy
•
No apparent added effect for TrkB
•
(In the clinic- concerns of too much pain relief with anti-NGFs)
Safety
 No observed adverse effects on peripheral neuronal health or function
•
Hyperphagia and weight gain – rodent specific?
•
Reversible Ataxia when CNS target coverage is achieved. TI related to plasma to
brain ratio
Would selective TrkA inhibition provide a broader TI?
15
Approach to Finding Highly Selective TrkA Chemical Matter
High Throughput Screen
ARRAY diversity
and kinase focused
Trk
potency
Promising
Kinase
<50 compounds
~many compound hits
in 2 novel chemical series
in > 20 chemical series
Selectivity
and
“drug likeness”
2 TrkA vs TrkB
selective hits
DMPK
Pharmacology
Toxicology
TrkA Selective Series (allosteric site)
•
high kinase selectivity outside of Trk family
•
high selectivity for TrkA over TrkB and C
16
Medicinal Chemistry
Optimization
Induced
Trk X-ray
Crystallography
Fit
>2000 designed
compounds
>120 TrkA allosteric + small molecule
x-ray structures
solved to date
Challenges of Drug Design in an Induced Fit Site
existing pocket
ATP site
ATP site
Potency is easy
Selectivity is hard
induced fit
allosteric site
Allosteric site
Potency is hard
Selectivity is easy
Allosteric site optimization is enabled by X-ray crystallography
17
confidential
TrkA Selective Inhibitors- Identifying the Allosteric Site
>100 TrkA Constructs Cloned
Triaged by Expression, Purification, and Binding
Thousands of Crystal Screens
Several Crystal Forms
>120 TrkA/Inhibitor Structures
>120 Selective TrkA Inhibitor Structures
Median Resolution: 2.8 Å
Range: 2.3 Å – 3.3 Å
18
Mechanism for Inhibiting Neurotrophin / Trk Signaling Cascade
Growth Factor Antibodies
NGF
BDNF
NT-4
NT-3
C1
LRR1-3
C2
Ig1
Trk Receptor Antibodies
Ig2
Trk kinase domain inhibitors
CR1
CR2
CR3
CR4
ATP site
high homology in ATP site
allosteric site
low homology in the allosteric site
Trk A
19
TrkB
TrkC
Allosteric site inhibitors selectively inhibit TrkA, but not TrkB, TrkC, or other kinases
P75NTR
Select Properties of Pan-Trk and TrkA Selective Leads
Program
20
TrkA Selective
Lead
AR-786
AR-256
AR-618
hTrkA cell IC50
0.6 nM
0.9 nM
5.6 nM
hTrkA free cell
IC50
12 nM
18 nM
30 nM
hTrkB / hTrkC
cell
>1000 nM
>1000 nM
>1000 nM
230 member
Kinase Panel
Clean
@ 10 µM
Clean
@ 10 µM
Clean
@ 10 µM
Predicted
hepatic Cl
Human, Rat
10, 38
(med, med)
12, 31
(med, med)
7, 17
(med, low)
Solubility
(ng/mL)
pH 1.2 / 6.5 / 7.4
750
60
1
>1000
5
1
>1000
590
130
peripheral to
CNS exposure
10 to 1
48 to 1
48 to 1
Very potent for inhibiting
NGF driven TrkA signaling
in cell
High selectivity over TrkB / C
High kinase selectivity
Peripherally Selective:
Only peripheral
inhibition in efficacy
studies
Kinase Selectivity of Array pan-Trk and TrkA Selective Inhibitors
Amino pyrimidine
ATP site
Diaryl urea
Type II DFG out
At 1 µM
Potent on Trks
Plus 83 off targets
at 0.5 µM
Potent on Trks
Plus ~7 off targets
AR-470
ATP site
Pan
at1 µM
Potent on Trks
One weak off target
21
AR-786
Allosteric
Selective
TRKA
inhibitor
at 10 µM
Potent on TrkA
weak on TrkB/C
No activity at 10 µM: against other pain targets
Clean TrkA
pharmacology
Key Scientific Questions- Small Molecule pan-Trk vs TrkA Selective
Do TrkA selective (allosteric) inhibitors show similar pain efficacy to
Pan-Trk (ATP site) inhibitors?
22
•
Is blocking TrkA upstream of BDNF / TrkB sufficient to alleviate
various modalities of pain / hypersensitization?
•
Is ATP site and allosteric site inhibition functionally equivalent in vivo?
Mean Difference in Print Area (%) ±SEM
TrkA Selective and pan-Trk Inhibitors are Equivalent in the Rat CFA Paw Model
80
70
60
50
**
40
***
30
20
10
0
Vehicle
30 mg/kg ARRY-470
(pan-Trk)
30 mg/kg AR786
(TrkA selective)
*p<0.05 by One-Way ANOVA with Bonferroni's correction compared to vehicle
Equivalent and Compelling efficacy for TrkA Selective
and Pan-Trk in a model of inflammatory pain
23
Duration of Action of a TrkA Selective Inhibitor in the Rat CFA Paw Model
30 mg/kg AR786
Mean Difference in Print Area (%)
±SEM
80
Vehicle
30 mg/kg AR-786
70
60
50
40
30
**
20
***
*
10
0
-10
1HR
2HR
4HR
Promising onset and duration of action
24
8HR
TrkA Selective and pan-Trk in a Surgical Incision Model
*
Tim Brennan
Equivalent and compelling efficacy for TrkA Selective
and Pan-Trk in a model of surgical pain
25
Mean HP Print Area (cm2)
SEM
TrkA Selective and pan-Trk in the Rat CIA Model of Polyarthritis
1
**
0.8
0.6
0.4
0.2
0
Vehicle
AR-786
TrkA selective
AR-772
pan-Trk
Equivalent and compelling
efficacy for TrkA Selective
selective
and Pan-Trk in a model of rheumatoid arthritis
26
TrkA Selective Inhibitors are Equivalent to Celecoxib in the Rat MIA Model
Algos
Compelling efficacy for TrkA selective in a model of
Osteoarthritis
27
confidential
Neurotrophin / Trk Signaling Mediates Peripheral Pain Response
Local release
Peripheral
Hypersensitization
NGF
TrkA
Proinflammatory cell
Central
recruitment and degranulation
Hypersensitization
Blocking TrkA at the allosteric site
is sufficient to
block the NGF pain cascade
28
Key Scientific Questions- Small Molecule pan-Trk vs TrkA Selective
Do TrkA selective (allosteric) inhibitors have a similar safety profile
to Pan-Trk (ATP site) inhibitors?
29
•
Hyperphagia and Weight gain?
•
Ataxia with CNS target coverage?
TrkA Selective Inhibitors Do Not Cause Hyperphagia or Ataxia in Rat
Vehicle
10 mg/kg
50 mg/kg
300 mg/kg
Mean Body Weight (gm)
SEM
500
450
400
350
300
250
Hyperphagia / weight gain
•
No increased food consumption
•
No weight gain / hyperphagia
•
Supports pan-Trk effect was
BDNF / TrkB driven
200
150
0
10
20
30
Time (days)
Pan-Trk
Treated
Females
Ataxia
Score
(max)
Incidence
TrkA
Selective
Treated
Females
Ataxia
Score
(max)
Ataxia Scoring System
Incidence
1 -"swimming” through litter, flattened or
splayed on cage bottom
2 - head bobbing, jittery or hyperactive, head
30 mg/kg
AR523
1
1 of 3
10 mg/kg
AR256
0
0 of 3
bobbing, nervous
3 - head rearing, disoriented, lethargy, agitated
30
100
mg/kg
AR523
1
300
mg/kg
AR523
2
3 of 3
3 of 3
50 mg/kg
AR256
0
300 mg/kg
AR256
0
0 of 3
4 - all of above plus falling over when on hind
legs, sleeping on back
0 of 3
Mechanistic Difference- Small Molecule, Oral TrkA kinase
vs anti-NGF m-ABs
Intermittent target knockdown is sufficient for efficacy, required levels seem to vary by model
TrkA
free IC90
Potential Clinically Advantages
31
1.
Ability to adjust inhibition by varying dose and schedule
2.
Ability withdraw drug
3.
Allows flexible titration to address different pain indications
Small Molecule TrkA Selective Inhibitors: What we know so far
Efficacy
 Great efficacy across pre-clinical pain models: similar to pan-Trk
o Looking for Collaborators – new models for mechanistic comparison
 Intermittent target inhibition is sufficient for efficacy
 Similar efficacy to that reported for anti-NGF antibodies
Safety
 No observed adverse effects on peripheral neuronal health
 No hyperphagia or weight gain to 28 days
 No ataxia even when CNS target coverage is achieved for 28 days
TrkA Selective Kinase Inhibition at the Allosteric Site Represents
a Promising New Mechanism for Modulating the NGF / Trk Pain Pathway
32
Acknowledgments
Medicinal Chemistry
Pharmacology / Toxicology
Enzymology
•
Karyn Bouhana
•
Shelley Allen
•
Lisa Pieti Opie
•
Anna Gomez
•
Julia Haas
•
Francis Sullivan
•
Patrice Lee
•
Yutong Jiang
•
Jed Pheneger
•
Tim Kercher
•
Robyn Hamor
•
Vince Murphy
•
Gabrielle Kolakowski
•
Chris Eberhardt
•
Dale Wright
•
Brad Newhouse
•
David Chantry
•
Allen Thomas
Ross Wallace
•
Steve Andrews
•
Brian Baer
•
Larry Burgess
•
Karin Brown
•
Dylan Hartley
•
Gary Hingorani
•
Jennifer Otten
•
Jamie Rogan
•
Susan Rhodes
•
Ron Franklin
DMPK
•
Outsourcing
•
33
Cheryl Napier
•
Tony Morales
•
Walt Voegtli
Computational Chemistry
•
Josh Ballard
Jim Blake
•
Guy Vigers
•
Barb Brandhuber
David Fry
Chris Lindemann
Don Corson
Translational
•
Structural Biology
•
CMC / Formulation
•
•
•
Cell Biology
Steve Miller
Executive Leadership
•
Kevin Koch
•
David Snitman
•
Jim Winkler