raul krauss*, todd bosanac, thomas engber, rajesh devraj ...€¦ · small molecule inhibitors of...
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Small Molecule Inhibitors of SARM1 Prevent Axonal Degeneration in vitro and in vivoRaul Krauss*, Todd Bosanac, Thomas Engber, Rajesh Devraj, Robert HughesDisarm Therapeutics, Cambridge, Massachusetts 02139, USA
3. A Proprietary Screening Platform Identified Small Molecule SARM1 Inhibitors
7. SARM1 Inhibitors Prevent Neuropathy in vivo in a Paclitaxel CIPN Model
Axonal degeneration is an early and ongoing event that causes disability anddisease progression in many neurodegenerative disorders of the central,peripheral, and ocular nervous systems, including multiple sclerosis, ALS,peripheral neuropathies, and glaucoma. SARM1 is the central driver of anevolutionarily conserved program of axonal degeneration downstream ofinflammatory, mechanical, metabolic, or chemical insults to the axon.SARM1 contains an intrinsic NADase enzymatic activity essential for its pro-degenerative functions, making it a compelling therapeutic target to treatneurodegeneration characterized by axonopathy.Disarm has developed a high-content screening platform and identifiedproprietary drug-like small molecule inhibitors of SARM1. These inhibitorsprotect rodent and human axons in vitro from mechanical,chemotherapeutic and mitochondrial damage. In sciatic nerve transection,SARM1 inhibitors prevent increases in plasma NfL, a clinically accessiblebiomarker of axonal degeneration. In a paclitaxel model of chemotherapy-induced peripheral neuropathy (CIPN) SARM1 inhibitors protect axonalstructure (plasma NfL) and function (SNAP amplitude).This is the first demonstration that pharmacological inhibition of SARM1 canreproduce the axonal protective phenotype observed in SARM1 knockoutmice. The availability of SARM1-dependent biomarkers of axonaldegeneration enables rapid translation of SARM1 inhibitors from discoveryto the clinic. SARM1 inhibitors have the potential to prevent axonaldegeneration in peripheral and central axonopathies and provide atransformational disease-modifying treatment for these disorders.
1. SARM1 Drives Rapid NAD+ Loss Leading to a Bioenergetic Crisis and Axonal Degeneration
Triggers Diseases
Axonal Degeneration Glaucoma
Multiple sclerosis
CIPN
Neurodegenerative (PD, ALS)
Peripheral neuropathies (diabetic, CMT)
CNS, Eye PNS
SARM1
Immuno/inflammatory
Metabolic / Toxins
Trauma
Mitochondrial / Other
Intraocular pressure
2. SARM1 - The Central Driver of Axonal Degeneration in Neurological Diseases
4. SARM1 Inhibitors Protect from Mechanical, Chemical and Mitochondrial InsultsDisarm’s small molecule inhibitors phenocopy SARM1 KO in primary DRG
5. SARM1 Inhibitors Protect Human AxonsHuman IPSC-derived motor neurons
Inhibitors from multiple chemical series inhibit SARM1 NADase
Disarm’s small molecule inhibitors phenocopy SARM1 KOPrevent increase of axonal structure biomarker plasma NfL and functional axonal conduction
Control Cut Cut + DSRM-XX Axonal Fragmentation
Uncut Cut Cut +Series 1
0
20
40
60
80
100
% A
xona
l Deg
ener
atio
n
********
Pharmacological SARM1 InhibitionGenetic Inhibition in SARM1 KO
** p< 0.01; **** p < 0.0001; ANOVA with Bonferroni post-hoc
Conclusions§ Disarm has advanced a proprietary R&D platform to discover
small molecule inhibitors of SARM1, the central driver of axonal degeneration
§ We are providing the first demonstration that Disarm’s novel small molecule SARM1 inhibitors reproduce the axonal protection phenotype seen in SARM1 knockout. Disarm’scompounds: • Protect axons in vitro from multiple pathological insults• Protect human iPSC-derived motor axons from traumatic injury• Prevent axonal degeneration and preserve axonal function in vivo in a
rodent model of chemotherapy-induced peripheral neuropathy (CIPN)
Disarm is developing small-molecule SARM1 inhibitors for patients with neurological diseases such as MS, ALS and CIPN
CutUntreated Cut + DSRM-XX Axonal Fragmentation
PaclitaxelUntreated Paclitaxel + DSRM-XX Axonal Fragmentation
Axonal Fragmentation NfL Biomarker
NfL Biomarker
NfL Biomarker
0
2000
4000
6000
8000
10000
12000
NfL
(pg
/ml)
Untreated Rotenone Rotenone +Series 1
**** ****
0
2000
4000
6000
8000
10000
NfL
pg/
ml
**
Uncut Cut Cut + Series 1
***
0
10000
20000
30000
40000
NfL
(pg
/ml)
**
Untreated Paclitaxel Paclitaxel +Series 1
****
0
20
40
60
80
100
% A
xona
l Deg
ener
atio
n
Untreated Paclitaxel Paclitaxel +Series 1
********
0
20
40
60
80
100
% A
xona
l Deg
ener
atio
n
Rotenone Rotenone +Series 1
Untreated
*****
0
20
40
60
80
100
% A
xona
l Deg
ener
atio
n
Uncut Cut Cut +Series 1
********
In vitro Assay in Primary DRG Neurons
** p< 0.01; *** p <0.001; **** p < 0.001; ANOVA with Bonferroni post-hoc
Rotenone Rotenone + DSRM-XXUntreated
Axonal cut
Imaging field Traumatic injury
Non-traumatic injury
Trau
mat
icCh
emot
hera
peut
icM
itoch
ondr
ial
Axon
al S
truc
ture
NfL
Bio
mar
ker
*p<0.05; ** p< 0.01; *** p <0.001; **** p < 0.0001; ANOVA with Bonferroni post-hoc
Pharmacological SARM1 InhibitionGenetic Inhibition in SARM1 KO
Axon
al S
truc
ture
NfL
Bio
mar
ker
Axon
al F
unct
ion
SNAP
Am
plitu
de
0 1 2 3 4 5 6 7 8 90
10
20
30
0
2
4
6
8
Time (h)
NAD
(ng
/ 100
,000
cel
ls) cADPRNAD
cADPR
(ng / 100,000 cells)
ADPR
0 .1 1 1 01 0 0
-2 0-1 0
01 02 03 04 05 06 07 08 09 0
1 0 01 1 01 2 0
T im e d e p e n d e n t in h ib it io n o f '1 2 9 65 µ g /m L N R K 1 -H E K 2 9 3 T + E V 5 4 6 7
5 µ M N A D + , 0 - 2 h p r e in c u b a t io n , 1 2 0 m in a t R T
c (to o l c m p d ) [µ M ]
% S
AR
M1
ac
tiv
ity
1 % D
MS O
w /o ly
sa te
S u b s tra te c o m p e t it io n o f '1 2 9 65 µ g /m L N R K 1 -H E K 2 9 3 T + E V 5 4 6 7
7 5 µ M N A D + , 2 h p r e in c u b a t io n , 1 2 0 m in a t R T
DMSO 0.03 X 0.3 X 3 X 30 X BlankDSRM-XX
Biochemical SARM1 Assay Cell-based DRG Axotomy AssaySARM1 produces cADPR after axotomy SARM1 Inhibitors Prevent cADPR IncreaseDisarm’s Small Molecules Inhibit SARM1
0
5
10
DSRM-Series 1
cAD
PR (n
g / 1
00,0
00 c
ells
)
1 X 10 X0.1 X0
Cut
Uncut
SARM1 Pathway
NAM
NMN
NAD+
NAM+ADPR
cADPR
NAMPT
NMNAT
SARM1
?
NAM + ADPR/cADPR
NAD+ cleaves into ADPR/cADPR
6. SARM1 Inhibitor Protects Axons in vivo After Sciatic Nerve Transection Disarm’s small molecule inhibitors phenocopy SARM1 KO
DSRM-XX
DSRM-XX DSRM-XX
DSRM-XX DSRM-XX
DSRM-XXDSRM-XX
DSRM-XX
0
1000
2000
3000
4000
Plas
ma
NfL
(pg/
ml)
Naive Vehicle DSRM-XXLow
DSRM-XXHigh
**
******
**
0
2000
4000
6000
Plas
ma
NfL
pg/
ml
WT WT SARM1HET
SARM1KO
PaclitaxelVehicle
******
****
0
50
100
150
200
250
SNA
P am
plitu
de (µ
V)
WT WT SARM1HET
SARM1KO
PaclitaxelVehicle
*****
*******
0
2000
4000
6000
Plas
ma
NfL
(pg/
ml)
** **
Vehicle Vehicle DSRM-XXLow
DSRM-XXHigh
Paclitaxel
0
50
100
150
SNA
P A
mpl
itude
(µV) ** **
Vehicle Vehicle DSRM-XXLow
DSRM-XXHigh
Paclitaxel
0
200
400
600
1800
2000
Plas
ma
NfL
pg/
ml
WT SARM1KO
****
Naïve SARM1HET
**
**
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