Investor and Analyst Breakfast American Society for Gene & Cell - - PowerPoint PPT Presentation

Investor and Analyst Breakfast

American Society for Gene & Cell Therapy Annual Meeting

Washington, D.C. May 12, 2017

This presentation contains forward-looking statements. All statements other than statements of historical fact are forward-looking statements, which are often indicated by terms such as “anticipate,” “believe,” “could,” “estimate,” “expect,” “goal,” “intend,” “look forward to,” “may,” “plan,” “potential,” “predict,” “project,” “should,” "will,” “would” and similar expressions. Forward-looking statements are based on management's beliefs and assumptions and on information available to management only as of the date of this press

• release. These forward-looking statements include, but are not limited to, statements regarding the

development of our gene therapies, the success of our collaborations, and the risk of cessation, delay or lack of success of any of our ongoing or planned clinical studies and/or development of our product

• candidates. Our actual results could differ materially from those anticipated in these forward-looking

statements for many reasons, including, without limitation, risks associated with collaboration arrangements,

• ur and our collaborators’ clinical development activities, regulatory oversight, product commercialization

and intellectual property claims, as well as the risks, uncertainties and other factors described under the heading “Risk Factors” in uniQure’s 2016 Annual Report on Form 10-K filed with the Securities and Exchange Commission on March 15, 2017. Given these risks, uncertainties and other factors, you should not place undue reliance on these forward-looking statements, and we assume no obligation to update these forward-looking statements, even if new information becomes available in the future.

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• Welcome

Matt Kapusta

Chief Executive Officer

• Scientific Overview

Harald Petry, Ph.D.

Chief Scientific Officer

• AAV5-miHTT gene therapy for Huntington’s

Disease Pavlina Konstantinova, Ph.D.

Director, New Therapeutic Target Discovery

• Neutralizing antibodies on efficacy of AAV

delivery Harald Petry, Ph.D.

Chief Scientific Officer

• Repeated gene delivery in NHP with AAV5

through immune adsorption Valerie Sier-Ferreira, Ph.D.

Head of Immunology

• Detection of AAV vector DNA and transgene

RNA in liver tissue by FISH Valerie Sier-Ferreira, Ph.D.

Head of Immunology

• Questions and Discussion

Group

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• Circulating Anti-AAV5 Neutralizing Antibody Titers up to 1:1031 Do Not Affect Liver

Transduction Efficacy of AAV5 Vectors in Non-Human Primates (Poster 198).

• Successful Repeated Hepatic Gene Delivery in Non-Human Primates Achieved with

AAV5 by Use of Immune Adsorption (Poster 395).

• AAV5-miHTT Gene Therapy Demonstrates Broad Vector Distribution and Strong

Mutant Huntingtin Lowering in a Huntington’s Disease Minipig Model. (oral presentation)

• Detection of AAV Vector DNA and Transgene RNA in Liver Tissue by Fluorescent In

Situ Hybridization (Poster 567).

• Novel AAV Vector Reservoirs: peripheral Blood Cells and Hematopoietic Progenitors.

(collaborator presentation)

AAV5-miHTT gene therapy for Huntington’s Disease

Pavlina Konstantinova, Ph.D. Director, New Therapeutic Target Discovery

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≥ 40 CAG repeat HTT DNA Prolonged CAG repeat exon 1 HTT mRNA Expanded polyglutamine (polyQ) tract Protein aggregation Neuronal degeneration

• Neurodegenerative disorder
• Autosomal dominantly inherited
• Prevalence: 1:10,000-30,000
• Age of onset around midlife
• Symptoms:
• Motor problems/chorea
• Cognitive decline
• Psychiatric disturbances
• Genetic testing available
• Only palliative treatment

TRACK-HD

Sagittal MRI control Early HD

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≥ 40 CAG repeat HTT DNA Prolonged CAG repeat exon 1 HTT mRNA Expanded polyglutamine (polyQ) tract Protein aggregation Neuronal degeneration

• Neurodegenerative disorder
• Autosomal dominantly inherited
• Prevalence: 1:10,000-30,000
• Age of onset around midlife
• Symptoms:
• Motor problems/chorea
• Cognitive decline
• Psychiatric disturbances
• Genetic testing available
• Only palliative treatment

TRACK-HD

Sagittal MRI control Early HD

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Adapted from Ross et al., Nat. Rev. Neurol. 10, 204-2016 (2014)

 function (%) Age 

presymptomatic prodromal early moderate advanced 1 2 3 4 5

Motor diagnosis

Premanifest

AMT-130

slowdown disease progression

Manifest

45 100 65 25

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ITR

polyA CAG promotor

miHTT-451

ITR

AAV5-miHTT (AMT-130):

• Replication deficient
• Adeno-associated virus, serotype 5
• Designed to deliver engineered miHTT
• Reduction of huntingtin expression
• Low potential off-target effects

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Miniarikova et al., Molecular Therapy NA 2016, Samaranch et al., Gene Therapy 2017

m u t a n t w i l d - t y p e m u t a n t w i l d - t y p e m u t a n t w i l d - t y p e m u t a n t w i l d - t y p e 5 0 1 0 0

S t r i a t u m

H T T a l l e l e

H T T p r o t e i n l e v e l ( % )

* * * *

P B S + 5 % S u c r o s e 5 . 2 x 1 0

9

2 . 6 x 1 0

1 0

1 . 3 x 1 0

1 1

g c / m o u s e A A V 5 - m i H T T

m u t a n t w i l d

• t

y p e m u t a n t w i l d

• t

y p e m u t a n t w i l d

• t

y p e m u t a n t w i l d

• t

y p e 5 0 1 0 0

C o r t e x

H T T a l l e l e

H T T p r o t e i n l e v e l ( % )

* *

Control AAV5-GFP injection

Striatum

gc/mouse AAV5-miHTT

Cortex

gc/mouse AAV5-miHTT

AAV5-miHTT

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H T T a g g r e g a t e s

P B S + 5 % s u c r o s e 6 . 5 x 1 0

1 0

g c A A V 5 - G F P 6 . 5 x 1 0

1 0

g c A A V 5 - m i H T T 1 0

1 0

1 0

1 0

* * * * *

AAV5-miHTT PBS+ 5%sucrose

D A R P P - 3 2 l e s i o n ( m m

3 )

P B S + 5 % S u c r o s e 6 . 5 x 1 0

1 0

g c A A V 5 - G F P 6 . 5 x 1 0

1 0

g c A A V 5 - m iH T T

0 . 0 0 . 5 1 . 0 1 . 5 2 . 0 * * * * * *

AAV5-miHTT PBS+ 5%sucrose

Prevention of neuronal dysfunction Suppression of mutant huntingtin aggregation

Miniarikova et al., Gene Therapy, accepted

M A Y 1 2 , 2 0 1 7 | 13 Evers MM, ASGCT 2017 presentation 536

striatum thalamus

1. PBS + 5%sucrose 2. 1x1013 gc AAV5-miHTT 3. 3x1013 gc AAV5-miHTT

striatum thalamus

GFP GFP

* * * * * * * * * *

4. 1x1013 gc AAV5-GFP:

M A Y 1 2 , 2 0 1 7 | 14 P u t a m e n C a u d a t e T h a la m u s C o r t e x

1 0

3

1 0

4

1 0

5

1 0

6

1 0

7

1 0

8

V e c t o r g e n o m e c o p i e s

p e r  g D N A

L L O D

P u t a m e n C a u d a t e T h a la m u s C o r t e x

0 . 1 1 1 0 1 0 0

M a t u r e m i H T T

m o l e c u l e s / c e l l

vector DNA microRNA

1 x 1 0

1 3 g c A A V 5 - m iH T T

3 x 1 0

1 3 g c A A V 5 - m iH T T

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P B S + 5 % S u c r o s e 1 x 1 0

1 3 g c A A V 5 - m iH T T

3 x 1 0

1 3 g c A A V 5 - m iH T T

P u t a m e n C a u d a t e T h a l a m u s C o r t e x 5 0 1 0 0 1 5 0

h H T T m R N A e x p r e s s i o n ( % )

( n o r m a l i z e d b y s s G A P D H , r e l a t i v e t o c o n t r o l )

* * * * * * * * * *

mutant HTT mRNA

P u t a m e n C a u d a t e T h a l a m u s C o r t e x 5 0 1 0 0 1 5 0

m u t a n t h u n t i n g t i n p r o t e i n ( % )

( r e l a t i v e t o P B S + 5 % s u c r o s e )

* * * * * *

mutant huntingtin protein

P B S + 5 % S u c r o s e 1 x 1 0

1 3 g c A A V 5 - m iH T T

3 x 1 0

1 3 g c A A V 5 - m iH T T

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• Dose-dependent reduction of HTT in HD rodent and tgHD minipig models translates

in therapeutic benefit.

• Widespread vector distribution upon (MRI-guided) CED delivery in NHP and tgHD

minipigs supported selection of striatum as the target brain structure.

• Long-term expression, tolerability and efficacy supports further clinical

development of HTT-lowering gene therapy for HD with AMT-130.

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Effective liver-directed gene delivery, despite the presence of neutralizing antibodies in non-human primates

Harald Petry, Ph.D. Chief Scientific Officer

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Approach:

• NAB impact on AAV5 transduction was tested in 14 NHP
• Sera of those 14 NHP all had pre-existing anti-AAV5 NAB titers ranging from 1:57 to 1:1031
• Those 14 NHP were injected intravenously with increasing doses of AAV5-hFIX:
• 5e11 gc/kg (n=3)
• 5e12 gc/kg (n=5)
• 2.5e13 gc/kg (n=3)
• 9.3e13 gc/kg (n=3)
• Transduction efficiency was assessed by measuring:
• Circulating FIX protein levels in plasma 7 days after vector infusion.
• Vector DNA in the liver 6 months after vector infusion (post mortem).

Impact of neutralizing antibodies (NAB) directed against AAV5 on efficacy of liver directed gene delivery in non-human primates (NHP)

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• Demonstration that successful AAV5-based liver-directed gene delivery can be

achieved in NHP, despite the presence anti-AAV NAB titers up to at least 1:1031.

• Poses question whether patients with pre-existing anti-AAV5 antibodies could benefit

from AAV5-based gene therapy.

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Successful repeated hepatic gene delivery in non-human primates achieved with AAV5 by use of immune adsorption

Valerie Sier-Ferreira, Ph.D. Head of Immunology

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Background:

• Presence of circulating neutralizing antibodies (NABs)

against AAV vector capsids impair transduction of the target cells and therapeutic efficacy.

Goal:

• To overcome anti-AAV pre-existing antibodies due to

exposure to wild type AAV

• To increase the number of patients eligible for the therapy
• To overcome anti-AAV antibodies raised after exposure to

AAV therapy

• To facilitate re-administration of AAV gene therapy

Approach:

• Immuno-adsorption procedure

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• Experimental set-up proof of concept in NHPs
• Mean Reduction levels NABs by immuno-adsorption: 11 times

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• SEAP and hFIX transgenes expression
• Proteins levels
• mRNAs levels

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• Data demonstrate that the use of an immune adsorption procedure enables

successful re-administration of an AAV5-based gene transfer in NHPs.

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Fluorescent in situ hybridization (FISH):

A powerful method to determine DNA/RNA distribution following AAV-based gene delivery

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“haat-GFP” vector DNA/RNA is visualized in green (FISH), Albumin RNA in purple (FISH), GS protein in blue (IHC)

Why FISH and IHC?

• To determine distribution
• To determine the cell specificity
• To quantify on the level of “a cell”

Following liver targeted AAV gene delivery

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FISH IHC

Image analysis (confocal microscopy, HALO program)

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• The combination of FISH and IHC permit to assess the physiological transduction

profile of AAV in the liver which is a valuable tool to further optimize AAV-targeting.

• To develop AAV-based gene therapies with increased efficiency and selectivity.

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Closing Remarks

and

Discussion