Gene Therapy for Neovascular AMD: Intravitreal Delivery of AAV-7m8 Vectors

David M. Brown, MD

Retina Consultants of Houston

KEY POINTS

▪ Non Human Primate Models Demonstrate:

▪Robust expression of aflibercept sustained 22.5 months after single IVT injection

▪ADVM-022 delivered aflibercept is sustained at levels similar to the aflibercept

recombinant protein-injected eyes 21-31 days post-dose.

▪ Human Phase 1 Trial ADVM-022 – OPTIC

▪ IND Active – August 2018

▪ Granted Fast Track – September 2018

▪ First patient dosed November 2018 – Preliminary Data expected 1Q20.

2

3

wt AAV2

rAAV2 vector

rep cap

Gene of Interest

PROMOTER GENE/ PROTEIN

• Simple virus made safe for gene therapy

• Protein on outside, DNA on inside

• Non-pathogenic, non-replicating, non-integrating

AAV

Adeno-Associated Virus (AAV) as a Gene Therapy Vector

Gene Therapy rAAV Toolbox Options

CapsidImprove transduction efficiency upon intravitreal delivery

cDNA: sFLT vs standard of care proteinssFLTAfliberceptRanibizumab

Expression cassetteCodon optimizationCombination of various regulatory elements for enhanced

protein expression

rAAV Capsid Variants

Capsid

AAV150

Summary of Trials

Novel Capsid: AAV.7m8

Variant of AAV2 with a 10-aa insertion in Loop IV of AAV2 VP3

Discovered by directed evolution (UC Berkeley)

Screened in vivo in murine retina

Exhibits robust tropism for photoreceptor via intravitreal injection

Dalkara et al. Sci. Transl. Med. (2013)

0.

250.

500.

750.

1000.

AAV2-CMV-sFLT 7m8-CMV-sFLT

Explant 1

Explant 3

sVEG

FR1

co

nce

ntr

atio

n (

pg/

ml)

Pig retinal explant

AAV2.CMV-GFP 7m8.CMV-GFP

Rat retina

AAV2

AAV.7m8

HEK293

AAV.7m8 Improves Transduction

Improved Transduction in Nonhuman Primate

➢ GFP expression mediated by 7m8 comes on faster

➢ 7m8 vector mediates highest level of GFP expression across the retina and inside the fovea

AAV2.CMV-GFP

7m8.CMV-GFP

4 wks 8 wks 12 wks

Fluorescence Fundus Images Heidelberg Spectralis Images

12 wks

• 5E+11 vg/eye

cDNA: sFLT vs Standard of Care Proteins

Codon optimization and assessment of regulatory elements

0. 1.25 2.5 3.75 5.

123456789

101112131415161718

Expression relative to CO1

Ca

sse

tte

Va

ria

nts

Evaluation of expression cassettes in pig retinal explants

Explant 1

Explant 2

Explant 3

22,139

3,159

16

353

77

* 3-day media

4E4 MOI, 2-wks timepoint.

12

ADVM-022: AAV.7m8-aflibercept

ADVM-022

Aflibercept expression cassette

AAV.7m8 capsid

C11=> strong ubiquitous expression cassette

AAV.7m8: Novel Capsid:

➢ AAV.7m8 Variant of AAV2

➢ Discovered by directed evolution(UC Berkeley)

➢ Exhibits robust tropism forretinal cells via intravitreal injection

Steve Ryan Laser Model

Drug Did it work in Non-Human Primates? Did it work in Rodents?

Ranibizumab Yes in laser induced CNV (Krzystolik 2002, Husain 2005)

Mixed. Yes in mice expressing human VEGF (Miki 2009); No in rats with laser induced CNV (Lu 2009)

Bevacizumab Yes in laser induced CNV (Lichtlen 2010, Goody 2011)

Mixed. Yes when administered IVT in laser induced CNV in mice (Hollanders 2014, Davis 2012); No in rats with laser induced CNV (Lu 2009); No when

administered intraperitoneally in laser induced CNV in mice (Yu 2008); Yes in mice expressing human

VEGF (Miki 2009); Yes in IVT VEGF induced CNV in rabbits (Ameri 2007)

Aflibercept Yes in laser induced CNV (Nork 2011) Yes. Yes in laser induced CNV in mice (Saishin 2003); Yes in subretinal CNV in mice expressing VEGF

(Saishin 2003); Yes when administered subcutaneously in subretinal matrigel induced CNV

in rats (Cao 2010); Yes when administered intraperitoneally in injury induced inflammatory NV

in mice expressing VEGF (Cursiefen 2010)

Pegaptanib No data Mixed. No in rats with laser induced CNV (Lu 2009); Yes in rat model of proliferative retinopathy

(hypoxia induced) (Ishida 2003). Prevents leukostasis and BRB breakdown in diabetic rats (Ishida 2003); prevents vascular permeability in

guinea pigs (Eyetech 2002), prevents VEGF induced corneal angiogenesis in rats (Eyetech 2002),

prevents NV in retinopathy of maturity in mice (oxygen induced) (Eyetech 2002)

Laser-induced CNV model in nonhuman primates- Rx-Gen (Matt Lawrence)

9 laser spots over the macula (1 next to fovea)

Lesions with various degrees of neovascularization and leakage

Lesions scored by fluorescein angiography and graded (I-IV) according to onset and

persistence of fluorescence over time

Only grade IV lesions considered as representative of CNV

1

2

3

4

5

6

7

8

9

Representative fundus fluorescein angiogram

Vehicle_IVT

70 d post treatment

14 d post lesion

4

4

4

2

2

4

4

4

4

Experimental Design

Day 0

• Intravitreal injection

• Vector: 2x1012 vg/eye (100 µL)

or

• Vehicle (100 µL)

Wk 8 Wk 12

• Fluorescein angiography

• Scoring of Grade IV CNV lesions

Wk 20

• Laser photocoagulation

• Standard of care in

positive control groups

Wk 10

• Termination

• Retinal punches for PK

Long-term Efficacy Study Design

Group Treatment N Dose (IVT; OU) Treatment deliveryLaser (OU) relative to

Day 0

1aADVM-022

2M/2F 2x1012 vg/50µL Day 0 13 month

1b 2M/1F 2x1012 vg/50µL Day 0 Not lasered

2aVehicle

2M/2F 50µL Day 0 13 months

2b 2M/1F 50µL Day 0 Not lasered

3 Aflibercept 2M/2F 1.2 mg/30µL Day of laser 13 months

Baseline 3 mo 7 mo 9 mo

Study TimelineLaser-induced

CNV

13 mo

Day 0: IVT dosing,

ADVM-022 and vehicle

16 mo

CNV assessment by FA and

OCT, 2 and 4 weeks post

laser

Aflibercept dosing

Termination of laser-treated animals

Biological sample collection

Termination of “observation”

groups

19 mo 21 mo

• One ADVM-022 animal died at Month 19 due to GI and kidney conditions unrelated to test article

18Confidential

Single dose IVT ADVM-022 Significantly Reduces the Incidence of Grade IV Lesions When Administered 13 Months Prior to Laser-induced CNV

Representative Fluorescein Angiography * P < 0.0001

19

ADVM-022 Delivered Intravitreally Results in the Long-Term Sustained High Levels of Aflibercept Expression in the Eye

20

Longitudinally assessed expression in vitreous humor up to 21 month after

ADVM-022 dose (7 animals)

Aflibercept levels in retina and choroid 16 months post ADVM-022 delivery

(n=8 eyes, laser-treated group)

R e t in a C h o ro id

1

1 0

1 0 0

Afl

ibe

rc

ep

t (

g/g

tis

su

e)

0 .12 4 6 8 1 0 1 2 1 4 1 8 2 0 2 2

0 .1

1

1 0

M o n th

Afl

ibe

rc

ep

t (

g/m

L)

0 3 7 9 1 6

L a s e r tre a tm e n t N o n - la s e re d e y e s (n = 6 , 3 a n im a ls ))

L a s e re d e y e s (n = 8 , 4 a n im a ls )

Long-term Expression of Aflibercept in Retina From ADVM-022 Does Not Affect Retinal Morphology

21

OD OS

OD OS

OD OS

OD OS

Vehicle control ADVM-022 Baseline

Month 21 post-dose

22

12.5-month Long-term Study Indicates Self-limiting Mild-to-Moderate Inflammatory Response in ADVM-022 (2X1012 vg/eye) Injected Eyes

n=14 eyes. Horizontal bars represent mean value

0 0 . 5 1 3 6 9 1 2 . 5

0

1

2

3

4

A q u e o u s c e l l s

M o n t h s

Ha

ck

et

-M

cD

on

al

d

Sc

or

e

0 0 . 5 1 3 6 9 1 2 . 5

0

1

2

3

4

A q u e o u s F l a r e

M o n t h s

Ha

ck

et

-M

cD

on

al

d

Sc

or

e

0 0 . 5 1 3 6 9 1 2 . 5

0

1

2

3

4

V i t r e o u s c e l l s

M o n t h s

Ha

ck

et

-M

cD

on

al

d

Sc

or

e

0 0 . 5 1 3 6 9 1 2 . 5

0

1

2

3

4

K e r a t i c P r e c i p i t a t e s

M o n t h s

Ha

ck

et

-M

cD

on

al

d

Sc

or

e

0 . 0 0 . 5 1 . 0 3 . 0 6 . 0 9 . 0 1 2 . 5

0

1 0

2 0

3 0

4 0

I n t r a o c u l a r P r e s s u r e

M o n t h s

IO

P (

mm

Hg

)

l o w l i m i t o f n o r m a l I O P r a n g e

h i g h l i m i t o f n o r m a l I O P r a n g e

0 0 . 5 1 3 6 9 1 2 . 5

0

1

2

3

4

L e n s C a p s u l e D e p o s i t

M o n t h s

Ha

ck

et

-M

cD

on

al

d

Sc

or

e

ADVM-022 Dosed Intravitreally Results in the Long-Term Sustained High Levels of Aflibercept Retina Expression

23

Aflibercept levels in vitreous and aqueous humors up to 22.5 month after ADVM-022 dose

0 2 4 6 8 1 0 1 2 1 4 1 6 1 8 2 0 2 2 2 4

0 .1

1

1 0

M o n th s

Afl

ibe

rc

ep

t (m

g/m

L)

V itre o u s h u m o r, O D

V itre o u s h u m o r, O S

A q u e o u s h u m o r, O D

A q u e o u s h u m o r, O S

PK Study Comparing ADVM-022 to Bolus Aflibercept Recombinant Protein

IVT administration of either ADVM-022 or aflibercept to NHPs

24

Group Treatment NDose (IVT,

OU)Administration

Sample collection

Day

1

Day

3

Day

7

Day

10

Day

14

Day

21

Day

28

Day

35

Day

42

Day

49

Day

56

1a Aflibercept 2(M/F) 1.2 mg Day 0

1b Aflibercept 2(M/F) 1.2 mg Day 0

1c Aflibercept 2(M/F) 1.2 mg Day 0

1d Aflibercept 2(M/F) 1.2 mg Day 0

1e Aflibercept 2(M/F) 1.2 mg Day 0

1f Aflibercept 2(M/F) 1.2 mg Day 0

2 ADVM-022 2(M/F) 2 x 1011 vg Day 0

3 ADVM-022 2(M/F) 6 x 1011 vg Day 0

4 ADVM-022 2(M/F) 2 x 1012 vg Day 0

5 ADVM-022* 2(M/F) 2 x 1012 vg Day 0

Terminal collection of aqueous and vitreous humor, retina, and choroid

* Oral treatment with Prednisone

Additional in-life vitreous humor collection days

Confidential

25

Dose-ranging Pharmacokinetics of IVT ADVM-022 in NHP

➢ IVT ADVM-022 provides ocular expression of aflibercept in all compartments including

retina and choroid where CNV occurs

• Levels comparable to aflibercept-injected eyes 21-31 days post-dose

Vitreous humor Aqueous humor

0 .0 1

0 .1

1

1 0

1 0 0

D a y

Afli

be

rc

ep

t (

g/m

L)

1 7 1 4 2 8 4 2 5 6

6 .7 g /m L

3 .6 g /m L

0 .0 1

0 .1

1

1 0

1 0 0

D a y

Afli

be

rc

ep

t (

g/m

L)

1 7 1 4 2 8 4 2 5 6

1 .7 g /m L

0 .5 g /m L

1

1 0

D a y

Afli

be

rc

ep

t (

g/m

L)

1 7 1 4 2 8 4 2 5 6

1

1 0

D a y

Afli

be

rc

ep

t (

g/m

L)

1 4 2 8 4 2 5 6

0 .0 1

0 .1

1

1 0

1 0 0

D a y

Afli

be

rc

ep

t (

g/g

)

1 7 1 4 2 8 4 2 5 6

7 .3 g /g

3 .5 g /g

0 .0 1

0 .1

1

1 0

1 0 0

D a y

Afli

be

rc

ep

t (

g/g

)

1 7 1 4 2 8 4 2 5 6

4 .5 g /g

0 .9 g /g

1

1 0

D a y

Afli

be

rc

ep

t (

g/g

)

1 4 2 8 4 2 5 6

1

1 0

D a y

Afli

be

rc

ep

t (

g/g

)

1 4 2 8 4 2 5 6

Retina Choroid

N=4 eyes/2 animals

26

Dose-ranging Pharmacokinetics of IVT ADVM-022 in NHP

➢ IVT ADVM-022 provides levels comparable to aflibercept-injected eyes 21-31 days post-dose

N=4 eyes/2 animals

27

Baseline assessment Treatment evaluation Follow-up

Day 1 244 8 12 16 20

Primary endpoint

(Safety) 24 weeks

104

Secondary endpoints (Efficacy)

Anti-VEGF rescue therapy administered if retreatment criteria met

Patients will be administered a tapering prophylactic corticosteroid regimen

Weeks

Cohort 1:

6x1011 vg

n=6

Cohort 2:

2x1012 vg

n=6

Cohort 3:

6x1012 vg

n=6

DMC Safety

Review

(4 weeks)

DMC Safety

Review

(4 weeks)DMC Safety

Review

(4 weeks)

OCT = Optical Coherence Tomography

Screening

Aflibercept

Injection

SD-OCT

Assessment

ADVM-022 Injection

ADVM-022 OPTIC Phase 1 Trial Design - Initiated 4Q18*

*First patient dosed mid November, 2018

KEY POINTS

▪ Non Human Primate Models Demonstrate:

▪Robust expression of aflibercept sustained 22.5 months after single IVT injection

▪ADVM-022 delivered aflibercept is sustained at levels similar to the aflibercept

recombinant protein-injected eyes 21-31 days post-dose.

▪ Human Phase 1 Trial ADVM-022 – OPTIC

▪ IND Active – August 2018

▪ Granted Fast Track – September 2018

▪ First patient dosed November 2018 – Preliminary Data expected 1Q20.

28