Lecture 18

Question 1

Features of Adeno-Associated Virus

Answer 1

• Small (25nm)
• Replication deficient
• Single stranded DNA genome
• approx. 5 Kb size
• approx. 120 human and non-human serotypes (capsids)

Question 2

What are the Pros of AAVs?

Answer 2

• Non-pathogenic
• Transduces dividing & non-dividing cells
• High lvl & stable long term expression
• Does not integrate
• Low immunogenicity
• Several different serotypes

Question 3

What are the Cons of AAVs?

Answer 3

• Small genome packaging capacity
• Cell/tissue specificity
• Pre-existing immunity

Question 4

How do you make an AAV vector?

Answer 4

- Make a transgene construct: 
o Promoter
o Transgene
o Elements (pA)
* Max 4.8 Kb

• Transfect cell with therapeutic construct above
  o Rep and Cap genes incorporated
  –> Cap: makes capsid
  –> Rep: replicates transgene inside cell

Question 5

What are the features of the Retina?

Answer 5

• At the back of the eye

- Made up of neurons and glial cells

Question 6

What are the features of Rods & Cones?

Answer 6

• Photoreceptors, light sensing neurons

- Capable of absorbing light and transducing electrical signals

Question 7

What are 5 examples of inherited retinal disorders?

Answer 7

• Colour vision defects
• Macular degeneration
• Cone-rod dystrophies
• Night blindness
• Retinitis pigmentosis

Question 8

What are the features of subretinal ocular treatment?

Answer 8

• technical difficult surgery
• smaller vol./dose of treatment
• strong targeting of photoreceptor cells
• regional treatment effect

Question 9

What are the features of intravitreal ocular treatment?

Answer 9

• Extremely easy injection
• Larger vol./dose of treatment
• Weak targeting of ONL and INL but good for ganglion cell
• Higher immune response

Question 10

What is Leber Congenital Amaurosis (LCA)?

Answer 10

= Rare congenital inherited eye disease
(1 in 40 000)

o Multi-factorial (genetic + environment)
o Current treatment = monthly eye injections
o 2 sets of clinical presentation:
- Non-exudative, atrophic/dry AMD
- Exudative/wet AMD

*RPE65 gene therapy restores vision

Question 11

Dry AMD

Answer 11

• associated with gradual loss of vision

- deposits of DRUSEN

Question 12

Wet AMD

Answer 12

• aggressive and responsible for 90% sever vision loss cases
• multifactorial with unknown cause
• abnormal blood vessels
• high levels of vascular endothelial growth factor (VEGF)

Question 13

What are the current treatment methods of Wet AMD?

Answer 13

• anti-VEGF therapy
• every 4-8 week injection
• high burden