1. Professor M. Dominik Fischer, MD DPhil FEBOphth
University Eye Hospital, Eberhard Karls University of Tuebingen, Germany
Nuffield Laboratory of Ophthalmology, University of Oxford, UK
Gene Therapy & Genome Editing
for Retinitis Pigmentosa

2. | brief background on retinitis pigmentosa
| gene therapy for eye diseases
| gene editing for eye diseases
Agenda
| outlook & science fiction

3. progressive loss of light sensing cells
Patients with Leber congenital amaurosis (LCA) are severely visu- lar changes that include alterations in fundus autofluorescence
den Hollander et al.
J CIin Invest 2010

4. Retinitis Pigmentosa
in the Western world,
blindness know their ge
to cost-effectively ident
Conventional Sanger
ring variants is warran
(CEP290) c.2991+1655
LCA patients of Europ
gated channel β3 (CNG
of ACHM patients (14
diseases for which the m
or a few genes encomp
exons/amplicons, such
exon CNGB3 gene or t
α3 [CNGA3] gene), STG
ABCA4 gene), and XL
RPGR gene or the 5-exo
Analysis of all know
effectively using allele
which yields pathologic
This technique is availa
AR RP, for which 25%–
cone rod dystrophy
Stargardt disease
cone dystrophies
achromatopsia
Leber’s congenital amaurosis

5. | affects 1: 3-4000 (= 200,000 in EU)
| umbrella term for multitude of inherited eye diseases
| symptoms vary widely - vast majority needs visual aids
| 30% AR, 20% AD, 10% XL, 40% simplex (> 200 genes)
Retinitis pigmentosa

6. autosomal recessive RP
Patients with Leber congenital amaurosis (LCA) are severely visu- lar changes that include alterations in fundus autofluorescence
den Hollander et al.
J CIin Invest 2010

7. | considerable unmet medical need (blinding condition)
| large number of different genes and disease types
| progressive degeneration - what is lost is lost
| small organ, easy access, excellent monitoring ability, .
. . .immune privilege, internal control, good scientific basis
important aspects

8. | brief background on retinitis pigmentosa
| gene therapy for eye diseases
| gene editing for eye diseases
Agenda
| outlook & science fiction

9. Gene therapy
viral vector acts
as gene Taxi
therapeutic DNA is transferred
to the target cell (e.g. the sick
and dying photoreceptor).
The delivered DNA substitutes
the disease-causing, defective
gene and acts like a healthy
gene.

10. CHM mit Muta(on c.877C>T (p.R293X)
AAV2.CHM ohne Muta(on

11. 11
201020142016
REP1 mit Muta(on c.877C>T (p.R293X)
AAV2.REP1 ohne Muta(on

13. CNGA3 linked achromatopsiaREP1 associated choroideremia
PDE6A linked Retinitis Pigmentosa
RPGR linked Retinitis Pigmentosa
BEST1 linked Morbus Best
RS1 linked juvenile Retinoschisis

14. current open ocular gene therapy trials
ClinicalTrial.gov query on Oct. 28th 2016

15. | brief background on retinitis pigmentosa
| gene therapy for eye diseases
| gene editing for eye diseases
Agenda
| outlook & science fiction

16. CRISPR/Cas gene editing - the concept

17. CRISPR/Cas - for Retinitis Pigmentosa
| key differences between retinal tissue vs. cell culture
- challenging delivery of tri-compound
- no active cell cycle (no S/G2 phases)
- limited repair mechanisms
- DNA less accessible

18. CRISPR/Cas - for Retinitis Pigmentosa
| knock-out of genes with ad mutation (20%)
- independent of repair mechanisms
- may benefit from addition of healthy
copy of gene
- off target events (0.1%) problematic

19. CRISPR/Cas - for Retinitis Pigmentosa
| editing of ar mutations (30%)
- different mutations on the same gene
need individual approaches (in contrast
to gene supplementation strategies)
- each individualised sequence needs
testing and approval (5 x 1Mio EUR/y)
- remaining risk of mutagenesis (50%?)

20. Test run for gene editing in retinal ganglion cells

21. Molecular Therapy 2016, 24(8), 1388–1394.
Single guide RNA, template DNA and Cas protein injected into
mouse zygotes. Zygotes that survived injection were implanted.
Principle of gene editing in mouse model of RP

23. | brief background on retinitis pigmentosa
| gene therapy for eye diseases
| gene editing for eye diseases
Agenda
| outlook & science fiction

24. | ad retinal dystrophies will be treated with CRISPR-Cas
| first hype will settle and allow gene editing 2.0 to emerge
| in the next 100 years, ex vivo gene therapy/editing will
dramatically alter medicine, agriculture and ecosystem
science fiction
| ar and xl retinal dystrophies by AAV gene therapy

25. NATURE,Oct2016,vol538

26. Centre for Ophthalmology Tübingen
A Zhour, I Seitz, F Reichel, JS Bellingrath, A Rina, A Ochakovski, L Conte, P Angelova
B Wilhelm, T Peters, N Kahle, A Rindtorff, B Beier, R Grund
L Kühlewein, L Kontostaneou, Y Vaheb, C Kortüm, E Zrenner
KU Bartz-Schmidt & M Ueffing, B Wissinger, M Seeliger
N Weisschuh, S Kohl, F Paquet-Durand, D Zobor, R Mühlfriedel
LMU Munich Columbia University
S Michalakis & M Biel S Tsang
MPI Biological Cybernetics Tübingen
G Keliris, T Ethofer
Nuffield Laboratory of Ophthalmology Oxford
M Simunovic, T Edwards, J Jolly, AR Barnard & RE MacLaren
Acknowledgements