Safety and Efficacy of Gene Transfer for Leber’s Congenital Amaurosis

1. Safety and Efficacy of Gene Transfer for Leber’s Congenital Amaurosis Ryan Smith October 20, 2008

2. Leber’s Congenital Amaurosis (LCA) • Rare inherited eye disease (1 in 80,000) • Symptoms first occur in early infancy • Irregular behavior, nystagmus • Progresses through time resulting in total blindness by 3rd or 4th decade of life • No treatment (until now?...) • Described by Theodore Leber in 19th Century • Amaurosis – Vision loss without any lesions • Congenital – Not acquired, present at birth

3. LCA2

4. Concept Virus Cell DNA of Interest Nucleus Insert DNA into genome Functional Protein

5. Adeno-Associated Virus (AAV) • Common human virus (80%) • Not known to cause a disease • Very mild immune response • Advantages • Integrates into the same place in the genome nearly every time • Elicits no clear cytotoxic response • Actually shown to fight cancer

6. Adeno-associated Virus • Disadvantages • Cannot hold a lot of DNA in the head • Efficiently packing it can require removing the viral DNA • This may cause it to not integrate in the proper location

7. Method • Identified three subjects (1 male, 2 female) age 19 to 26 with LCA2 and profound vision loss. • Put a cDNA of RPE65 gene with a chicken beta actin promoter into an AAV vector. • Injected AAV2.hRPE65v2 into subretinal tissue under general anesthesia. • Only in one eye (the worse based on objective and subjective measurement) – Right in all three • Other used as a control

8. Method • Patients’ vision and health evaluated before and after surgery • Subjective Vision tests: • Pupillary light reflex • Nystagmus testing • Objective Vision tests: • Visual Acuity tests • Goldmann visual-field examination • Ability to navigate an obstacle course

9. Surgery and Immediate Response • Injection caused localized retina detachment • Reattached 14 hours after surgery • All patients showed unremarkable retinas after surgery • Patient 2 • developed outer lamellar cyst in fovea • Also developed a macular hole

10. Macular Hole

11. Post Surgery – Response to Vector • Vector DNA only found in tear of patient 1 on day 1 after surgery • Vector was not observed to spread through the body • No humoral or cell-mediated immune response in any of the patients • Neutralizing antibody titers to AAV2 capsid observed in Patient 2, but went away over time

12. Pupillary Light Response

14. Pupillary Light Response • All patients showed an increased light sensitivity • The injected eye was approximately three times as sensitive to light than at baseline • The injected eye drove the light response in both eyes • In each situation the less functional eye became better than the previously better functioning eye

15. Before and After Surgery

16. Before and After Surgery cont.

17. Results • Patient 1 • HM -> 20/1050 (P<0.001) • Patient 2 • HM -> 20/710 (P<0.001) • Patient 3 • 20/640 -> 20/290 (P=0.002) • Visual Field size increased • Nystagmus was reduced • Ability to navigate obstacle course improved (for patient 2)

18. Discussion • All patients showed improvement for 6 weeks, then improvement slowed (did not go away) • Reduction in nystagmus may account for improvement in uninjected eye • No apparent local or systemic effects of AAV

19. Macular hole in patient 2 • No inflammation or acute retinal toxicity • Could be contraction of a preexistingmembrane stimulated by the surgery • Could be caused by the surgery • No reported vision loss by the patient • Definitely would cause vision loss to normally functioning retina

20. Future Work • These three patients will continue to be monitored • This was first of three experiment groups involving 9 people • First group got a small dose • Second group gets a medium dose • Third group gets a large dose • Possible improved response if patients are treated in childhood

21. Fin.