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Praliciguat, a clinical-stage sGCstimulator, improved insulin sensitivity, lipid tolerance and energy utilization in a diet-induced obesity mouse model
Chad D. Schwartzkopf, John R. Hadcock, Julien Roux, Guang Liu, Courtney Shea, Mark G. Currie, G. Todd Milne, Jaime L.
Masferrer and Juli E. Jones
The current prevalence of obesity is alarming
2
WHOBody Mass Index (BMI) = kg/m2
• In 2014, an estimated 900 million worldwide were obese and 1.5 billion were overweight
• Obesity is associated with numerous comorbid conditions such as diabetes, cardiovascular disease and cancer
Clinical experience with the sGC stimulator praliciguat suggests potential metabolic benefit
3
• Praliciguat (IW-1973) is currently in Phase 2 for diabetic nephropathy and HFpEF
• In an exploratory Phase 2 study in 26 patients with type 2 diabetes and hypertension on standard of care therapy, 14 days of praliciguat treatment showed positive trends in reducing
• Fasting plasma glucose
• HOMA-IR
• LDL cholesterol
• Triglycerides
Ø The purpose of the current studies was to explore the mechanism of action behind the potential metabolic benefits of praliciguat utilizing an obese mouse model
Change in HOMA-IR (%)patients not on concomitant insulin therapy
-6 0
-4 0
-2 0
0
2 0
LS
Me
an
(9
5%
CI)
P la c e b o (n = 4 )
P ra lic ig u a t 4 0 m g (n = 1 2 )
-1 3 ( -4 0 , 1 5 ) -3 6 ( -5 3 , -1 9 )
L S M e a n D iffe re n c e (9 5 % C I)-2 3 ( -5 6 , 9 )
NCT03091920
Praliciguat shows extensive distribution to key target tissues
4
Heart Liver Kidney LungSkeletal Muscle
Adipose
Tissue/plasma Cmax ratio 5.0 53.0 9.6 4.1 3.1 12.5
Enhanced NO-sGC-cGMP signaling by praliciguat demonstrates benefits in preclinical models
5Adapted from Tobin, Zimmer et al. J Pharmacol Exp Ther 2018;365:664-675. Copyright © 2018 The Author(s).
• In preclinical models, compared to disease controls, animals treated with praliciguat have:
• Preserved cardiac function• Less cardiac hypertrophy• Lowered biomarkers of
inflammation and fibrosis• Less renal damage
Diet-induced obesity (DIO) mouse model is a common preclinical model of insulin resistance
• Simple model: C57Bl/6 mice are switched to 60% high fat diet (HFD) at 6 weeks of age
• Within 3 weeks mice develop an obese phenotype• Increased adiposity
• Insulin resistance
• Leptin resistance
• HFD is continued throughout study and animals will continue to gain weight
• Widely used in anti-obesity and metabolic syndrome researcho Animals will lose weight on anorectic drugs (GLP-1, phentermine)
o Animals will increase insulin sensitivity/secretion when on anti-diabetic drugs (GLP-1, metformin, rosiglitazone)
Study Design
7
• Study was performed at thermoneutrality (30° C)
• Housing animals below their thermoneutral zone alters energy expenditure and substrate utilization
• Cold stress undermines mouse modeling (Karp, J. Exp. Med. Vol. 209 No. 6 1069-1074 (2012))
Age = 6wk 12wk 14wk 18wk Collect blood, tissues after
overnight fast and 2H post PO
dose
Acclimation, T=30°C
60% HFD Pretreatment (8 weeks)
Maintained on LFD (8 weeks)
Obese Controls
PRL (6 mg/kg) in HFD
Study Phase
Praliciguat did not alter body weight, food intake or body composition
8
7 14 21 28 35
20
25
30
35
40
45
50
Study Day
Bod
y W
eigh
t, g
7 14 21 28 35
0
10
20
30
40
50
Study Day
Fat M
ass,
% o
f tot
al
7 14 21 28 35
40
50
60
70
80
Study Day
Lean
Mas
s, %
of t
otal
7 14 21 28 35
0
100
200
300
400
500
Study Day
Cum
ulat
ive
Calo
ricIn
take
, kca
l
LeanObesePRL
Body Weight Fat Mass Lean Mass Food Intake
Praliciguat-treated mice had greater insulin sensitivity than control DIO mice
• Compared to obese mice, four weeks of praliciguat resulted in:o Lower fasting insulino Lower C-peptide o Lower HOMA-IR
• In this model, fasting glucose was not altered in obese or PRL-treated mice
9
Insulin Glucose
C-Peptide HOMA-IR
Lean Obese PRL 0
1
2
3
4
Insu
lin, n
g/m
l
****
*
Lean Obese PRL 100
200
300
400
Glu
cose
, mg/
dl
Lean Obese PRL 0
1
2
3
4
C-P
eptid
e, n
g/m
l
****
*
Lean Obese PRL 0
10
20
30
40
50
****
HO
MA-
IR
*
DIO mice treated with praliciguat had lower expression of genes involved in inflammation
10
Liver: Tnf
Lean Obese PRL 0.000
0.005
0.010
0.015
0.020
norm
aliz
ed g
ene
expr
essi
on
****
**
Liver: TNFα SKM: Ccl2
Lean Obese PRL 0.00
0.02
0.04
0.06
0.08
norm
aliz
ed g
ene
expr
essi
on
**
*
Muscle: Ccl2
WAT: Akt1
Lean Obese PRL 0.0
0.5
1.0
1.5
2.0
2.5
norm
aliz
ed g
ene
expr
essi
on
**** *
WAT: Icam1
Lean Obese PRL 0.0
0.1
0.2
0.3
0.4
0.5
norm
aliz
ed g
ene
expr
essi
on
****
Adipose: Akt1 Adipose: Icam1
DIO mice treated with praliciguat had normalized expression of genes involved in lipid handling
11
Liver: Pnpla3
Lean Obese PRL 0.000
0.001
0.002
0.003
norm
aliz
ed g
ene
expr
essi
on
** *
Liver: Lpl
Lean Obese PRL 0.0
0.1
0.2
0.3
norm
aliz
ed g
ene
expr
essi
on
***
Liver: Pnpla3 Liver: Lipoprotein lipaseLiver: Slc27a1
Lean Obese PRL 0.0
0.2
0.4
0.6
0.8
norm
aliz
ed g
ene
expr
essi
on
*****
Liver: Long-chain fatty acid transport protein 1
SKM: Lipe
Lean Obese PRL 0.0
0.5
1.0
1.5
2.0
2.5
norm
aliz
ed g
ene
expr
essi
on
**
Muscle: Hormone-sensitive lipase WAT: Fdft1
Lean Obese PRL 0
1
2
3
4
norm
aliz
ed g
ene
expr
essi
on******
WAT: Ppara
Lean Obese PRL 0.0
0.1
0.2
0.3
norm
aliz
ed g
ene
expr
essi
on ****
**
Adipose: Farnesyl-diphosphate farnesyltransferase 1
Adipose: Pparα
DIO mice treated with praliciguat had lower expression of leptin in liver and muscle suggesting repartitioning of adipose tissue
12
Liver: Lep
Lean Obese PRL 0.000
0.005
0.010
0.015
norm
aliz
ed g
ene
expr
essi
on
***
**
Liver Leptin SKM: Lep
Lean Obese PRL 0
1
2
3
4
norm
aliz
ed g
ene
expr
essi
on
* *
Muscle Leptin
Nor
mal
ized
Gen
e Ex
pres
sion
The assessment of praliciguat on energy expenditure and lipid handling
13
• Energy expenditure (EE) was measured on day 8, 9, 20, 21, 32 and 33
• A lipid tolerance test (LTT) was performed on day 38
Age = 6wk 12wk 14wk 18wk
Acclimation, T=30°C
60% HFD Pretreatment (8 weeks)
Maintained on LFD (8 weeks)
Obese Controls
PRL (6 mg/kg) in HFD
Study Phase
19wk
EE EE EE LTT
Praliciguat-treated mice had a mild increase in energy expenditure compared to obese controls
14
• Increase in energy expenditure was accompanied with an increase in fat oxidation through day 21.
1:00
2:00
3:00
4:00
5:00
6:00
7:00
8:00
9:00
10:0
011
:00
12:0
013
:00
14:0
015
:00
16:0
017
:00
18:0
019
:00
20:0
021
:00
22:0
023
:00
200
250
300
350
400
Day 8
Hour
Ener
gy e
xpen
ditu
re(K
cal/d
ay/k
g le
an)
LeanObesePRL
Obese vs. Lean nsObese vs. PRL ****
1:00
2:00
3:00
4:00
5:00
6:00
7:00
8:00
9:00
10:0
011
:00
12:0
013
:00
14:0
015
:00
16:0
017
:00
18:0
019
:00
20:0
021
:00
22:0
023
:00
200
250
300
350
400
Day 20
HourEn
ergy
exp
endi
ture
(Kca
l/day
/kg
lean
)
LeanObesePRL
Obese vs. Lean nsObese vs. PRL ****
1:00
2:00
3:00
4:00
5:00
6:00
7:00
8:00
9:00
10:0
011
:00
12:0
013
:00
14:0
015
:00
16:0
017
:00
18:0
019
:00
20:0
021
:00
22:0
023
:00
250
300
350
400
450
Day 32
Hour
Ener
gy e
xpen
ditu
re(K
cal/d
ay/k
g le
an)
LeanObesePRL
Obese vs. Lean ***Obese vs. PRL ****
DIO mice treated with praliciguat had lower plasma triglycerides content suggesting improved lipid handling
15
Lean Obese PRL0
50
100
150
Trig
lyce
rides
, mg/
dl *¯13%**¯19%
0
100
200
300
400
ObesePRL
Time, hours
Trig
lyce
rides
, mg/
dl
Lean****
**
**
-0.5 51.5 3
Olive Oil10 µl/g
Oral Lipid Tolerance TestFasting Plasma Triglycerides
PKG VASP NF-kB
Inflammation
Insulin resistance
Ectopic fat accumulationMetabolic syndrome
Type 2 diabetesCardiovascular disease
High Fat Diet
Insulin Receptor
Inflammation and NO insufficiency impair insulin signaling
16
1. The consumption of high-fat diet can lead to obesity resulting in reduced NO signaling
2. Reductions in VASP and high fat diet both increase NF-kB, which increases inflammation and inhibits insulin signaling leading to insulin resistance
3. All of these factors lead to comorbid metabolic conditions
1
2
3
PKG VASP NF-kB
Inflammation
Insulin resistance
Ectopic fat accumulationMetabolic syndrome
Type 2 diabetesCardiovascular disease
High Fat Diet
Insulin Receptor
Praliciguat’s effect on inflammation and insulin sensitivity in DIO mice may be due to inhibition of NF-kB activation
17
1. NO-sGC-cGMP signaling stimulates VASP phosphorylation which inhibits NF-kB
2. Praliciguat-treated mice had lower levels of inflammatory gene expression across target tissues
3. The increased insulin sensitivity in praliciguat-treated mice may be due to its effects on the NF-kB signaling pathway
1
32
Improved lipid handling in praliciguat-treated DIO mice may be in part due to the increase in adipose Pparα
18
1
2
Pparα
High Fat Diet
↓Triglycerides↑FFA Oxidation
1. High fat diet inhibits Pparα; yet praliciguat-treated mice had a restoration in adipose tissue pparα gene expression
2. Pparα decreases triglycerides and increases FFA oxidation
3. Dietary nitrate enhance Pparα and Pparβ/δ activity
4. Pparα agonists such as fenofibrate have been demonstrated to increase NO via eNOS
4
3
Summary
19
§ The positive trends in several metabolic parameters observed in humans following praliciguat treatment is recapitulated in DIO mice including reduced fasting insulin, HOMA-IR, and fasting triglycerides
§ Praliciguat did not affect plasma glucose, body weight, food intake or body composition
§ Praliciguat treated-mice had positive changes in the expression of genes associated with inflammation and lipid metabolism in key metabolic tissues
§ The effects of praliciguat on inflammation and insulin sensitivity may be explained by inhibition of NF-kB signaling
§ The effects of praliciguat on lipid handling may be mediated by activation of Pparα
§ These data demonstrate broad metabolic effects such as improved insulin sensitivity, lipid handling and increased energy utilization in obese mice housed at thermoneutrality