Pg. 1
Restoring Biological Harmony for Patients with Debilitating DiseaseRestoring Biological Harmony for Patients with Debilitating Disease
The miR-29b Mimic Remlarsen as an Anti-Fibrotic Therapeutic in the EyeCorrie Gallant-Behm, PhD, MQARS
AOPT 20190310
Pg. 2
Fibrosis is a Significant Cause of Blindness
▪ Corneal fibrosis is second leading cause of blindness (after cataracts) worldwide. Results from corneal damage beyond Bowman’s membrane:▪ Viral keratitis
▪ Bacterial keratitis
▪ Fungal keratitis
▪ Sterile keratitis, dry eye disease
▪ Physical damage to cornea (trauma, surgery, thermal and chemical burns)
▪ Retinal fibrosis is the final common pathophysiological denominator and a major contributor to vision loss from:▪ Age related macular degeneration
▪ Diabetic retinopathy
▪ Retinal detachment (proliferative vitreoretinopathy)
▪ Retinopathy of prematurity
▪ Neovascular glaucoma
Pg. 3
▪ Reduced Expression of miR-29 has been Implicated in the Development and Progression of a Wide Range of Fibrosis Indications
miR-29 is an Anti-Fibrotic miRNA
Ocular fibrosis:Corneal fibrosis
Fuch’s Endothelial Corneal Dystrophy
Glaucoma/trabeculectomy bleb
Retinal fibrosis
Age related macular degeneration
Diabetic retinopathy
Pulmonary fibrosis
Inflammatory bowel disease
Liver fibrosis
Cardiac fibrosis
Tendinopathies
Renal fibrosis
Cutaneous fibrosis
Osteoarthritis
Dupuytren’s contractures
miR-29TGF-b
+ ECMFibrosis
miR-29
agonist
TGF-b
+ ECMFibrosisX X
▪ miR-29 inhibits inflammation, TGF-b activity, EMT, fibroblast-to-myofibroblast transition and ECM synthesis
▪ miR-29 inhibits every step of the collagen fibrillogenesis pathway
miRagen preclinical and/or
clinical data + literature support
Literature support
Pg. 4
Remlarsen Impacts Multiple Fibrotic Processes
Remlarsen Corticosteroids (SOC)
Reduces inflammation ● ●
Accelerates healing ●
Reduces EMT ●
Reduces fibroblast activation ●
Reduces myofibroblasts ●
Inhibits collagen production ●
Pg. 5
The Eye is a Preferred Target Organ for microRNA Therapeutics
▪ Delivery to multiple compartments of the eye, and via multiple routes of administration including injection and topical drops
▪ Stability due to reduced nuclease concentration enables long duration of action and infrequent dosing
▪ Reduced liability for toxicities due to local administration with limited systemic exposure
▪ Remlarsen is a clinical stage miR-29b mimic that has been shown to downregulate the expression of multiple collagens and other ECM molecules in the skin in humans and to inhibit fibroplasia in iatrogenic cutaneous wounds in normal healthy volunteers. It has also been shown to regulate this same expression in in vivo animal models of pulmonary, hepatic, corneal and retinal fibrosis.
Pg. 6
Topically Applied Remlarsen Eye Drops Are Readily Taken Up by Damaged Corneas
▪ Distribution of drug to all layers of the cornea, including corneal endothelium, after drop administration▪ Animal model: Rat alkali burn
▪ Formulated in simple buffer with no additional stabilizers
▪ No penetration enhancement/nanoparticle formulation required
EpitheliumStroma
FITC-remlarsen
DAPI
Pg. 7
▪ Uptake of remlarsen in alkali burned cornea is high in the presence of corneal compromise and remains elevated for up to 28 days
Remlarsen Eye Drops Demonstrate Sustained Uptake in Alkali Burned Cornea
0
5
1 0
1 5
2 0
2 5
6 0
8 0
1 0 0
1 0 0 0
2 0 0 0
3 0 0 0
miR
-29
ex
pre
ss
ion
(fo
ld c
ha
ng
e v
s.
sa
lin
e)
S a lin e 1 2 4 7 1 0 1 4 2 1 2 8
D a y s a fte r re m la rs e n tre a tm e n t in it ia tio n
1717x
14x
4x
13x
1 .7x
58x 61x
11x
N=6 per group
Pg. 8
Remlarsen Eye Drops Accelerate Healing of Damaged Cornea
▪ Decreased inflammation and stromal damage
▪ Restored normal corneal architecture after alkali burn
No Burn
Burn
+
Saline
Burn
+
Remlarsen
Images taken Day 4 post-burn
Thin, friable epithelium
Infiltrating immune cells
Stromal damage
Epithelium nearly normal
Minimal inflammatory infiltrate
Stromal damage absent
Pg. 9
Remlarsen Accelerates Corneal Healing and Reduces Scarring
▪ Epithelium thickness restored to normal following remlarsen treatment
▪ Stromal thickness (scarring) reduced with remlarsen treatment
S a lin e R e m la r s e n
0
2 0
4 0
6 0
Ep
ith
eli
um
th
ick
ne
ss
(
M)
p < 0 .0 1
N = 1 2 N = 1 1
S a lin e R e m la r s e n
0
1 0 0
2 0 0
3 0 0
Str
om
al
thic
kn
es
s (
M)
p < 0 .0 5
N = 1 2 N = 1 1
Dotted line indicates normal corneal thickness
Day 14 after corneal burn
Epithelial thickness Stromal thickness
Pg. 10
Remlarsen Reduces Corneal Scarring
▪ Reduced corneal hazing, scarring▪ Increased corneal transparency
▪ Reduced a-SMA expression▪ Reduced EMT
▪ Reduced FMT
S a lin e R e m la r s e n
1
2
3
4
5
Ha
zin
g/s
ca
rrin
g s
co
re
(1
-5)
p = 0 .0 0 3
N = 1 2 N = 1 2
Burn + Saline
Burn + Remlarsen
Images taken Day 10 post-burn, 20x magnification
Double-masked observer determination
according to the following scoring system
(Scoring Day 10 post-burn):
1 = normal
2 = slight haze
3 = moderate haze
4 = severe haze + slight scar
5 = severe haze + overt scar
Pg. 11
▪ miR-29b PD biomarkers are repressed following twice daily remlarsen drop treatments
▪ Consistent in multiple studies
▪ Multiple collagens and pro-fibrotic factors are affected
▪ Remlarsen typifies pathway/network regulation by microRNAs
▪ Regulation of many genes concurrently leads to coordinated impact on pathology
Remlarsen Demonstrates Target Engagement in Cornea in vivo
S a lin e R e m la r s e n
0 .0
0 .5
1 .0
1 .5
T a r g e t E n g a g e m e n tm
RN
A E
xp
re
ss
ion
(Fo
ld C
ha
ng
e)
p < 0 .0 0 0 1
COL1A1
COL1A2
COL5A2
FN1
TGFB2
MMP2
Day 14 after corneal burn
N=6 per group
Pg. 12
Remlarsen is Well Taken Up by All Layers of the Retina After IVT Administration
▪ Distribution of remlarsen to all layers of the retina, including pigmented epithelium via intravitreal injection
▪ Formulated in simple buffer with no nanoparticles or exotic formulation required
Lens
Ganglion cell layer
Inner plexiform layer
Inner nuclear layer
Outer nuclear layer
Photoreceptor inner
segments
Photoreceptor outer
segments
Outer plexiform layer
DAPI
FITC-remlarsen 20x 40x
Photoreceptor outer
segments
RPE layer
Pg. 13
Remlarsen Demonstrates Favorable PK in the Retina
▪ Remlarsen retention in retina and back of the eye for > 1 week after single intravitreal injection; later time points not yet tested
▪ Supports infrequent dosing
S a l in e 2 4 h rs 4 8 h rs 4 d a y s 7 d a y s
0
5 0 0 0
1 0 0 0 0
1 5 0 0 0
2 0 0 0 0
m iR -2 9 b le v e ls in ra t re t in a
miR
NA
ex
pre
ss
ion
(fo
ld c
ha
ng
e)
* *
R e m la r s e n
S a l in e 2 4 h rs 4 8 h rs 4 d a y s 7 d a y s
0
5 0 0 0
1 0 0 0 0
1 5 0 0 0
m iR -2 9 b le v e ls in ra t R P E , c h o ro id & s c le ra
miR
NA
ex
pre
ss
ion
(fo
ld c
ha
ng
e)
*
R e m la r s e n
N=6 per group
Pg. 14
Remlarsen Demonstrates Target Engagement in RPE Cells in vitro
▪ Remlarsen represses collagen expression in the presence and absence of TGF-b1 in multiple human retinal pigment epithelial cell lines
UT
C
MO
CK
0.5
nM
1 n
M
5 n
M
10
nM
25
nM
MO
CK
0.5
nM
1 n
M
5 n
M
10
nM
25
nM
0
1
2
3
4
Fo
ld C
ha
ng
e
+ 5 n g /m L T G F -b 1
CO
L1
A1
CO
L1
A2
CO
L3
A1
** ****
iPS RPE Cells in Culture
N=3 per group
Pg. 15
▪ Rat STZ diabetic retinopathy model
▪ miR-29b PD biomarkers are repressed following 3x weekly IVT remlarsen treatments
▪ Rabbit proliferative vitreoretinopathy model
▪ miR-29b PD biomarkers are repressed following 4x weekly IVT remlarsen treatments
Remlarsen Demonstrates Target Engagement in the Retina in vivoin Multiple Models
S a lin e R e m la rs e n
0 .0
0 .5
1 .0
1 .5
2 .0
mR
NA
Ex
pre
ss
ion
(Fo
ld C
ha
ng
e)
* * *
COL1A1
COL1A2
COL3A1
COL4A3
CTGF
FN1
MMP2
TGFb3
B S S R e m la r s e n
0
1
2
3
mR
NA
ex
pre
ss
ion
(fo
ld c
ha
ng
e)
*
COL1A1
COL1A2
MMP2
TGFB2
N=7 per groupN=6 per group
Pg. 16
▪ miR-29b mimics are well taken up by a damaged/ulcerated cornea following topical drop administration, and by all retinal layers following intravitreal injection
▪ Remlarsen exhibits favorable PK in the cornea and retina
▪ Cornea▪ Remlarsen treatment reduces the severity of alkali burn, accelerates repair
▪ Remlarsen treatment decreases corneal hazing/scarring, corneal stromal thickness, and a-SMA staining post-burn
▪ Remlarsen engages its pharmacodynamic targets and reduces collagen/ECM expression in vivo
▪ Retina▪ Remlarsen engages its pharmacodynamic targets and reduces collagen/ECM expression in vitro
and in vivo in multiple models
▪ Remlarsen represents a potential therapeutic for corneal and retinal scarring regardless of the etiology:▪ Infectious keratitis, surgery, trauma, genetic disease
▪ AMD, DR, retinal detachment
Conclusions