2. 2
Neuronal Ceroid Lipofuscinoses (NCL)
o Group of severe autosomal recessive neurodegenerative disorders, affecting
children and young adults.
o Lysosome storage disease: accumulation of
autofluorescent, electron dense material in cells.
NCL foundation
3. 3
Juvenile NCL (JNCL) or Batten Disease
o Most common type of NCL.
o Appearing age 6-8 years, life expectancy between 20s-30s.
o Lysosomal storage material: subunit c of mitochondrial ATP synthase F0.
o Gene affected: CLN3 (16p12.1).
o 438-aa transmembrane protein (Mw=48kDa) of unknown function
and ubiquitous localization.
4. 4
Most common CLN3 JNCL causing mutation
Del exons 7-8: c.461-
280_677+382del966,
p.[Gly154Alafs*29,
Val155_Gly264del]
5. 5
o Objective: correct CLN3 deficiency.
o Hypothesis: Low CLN3 levels are required for cellular homeostasis.
o Method: Use of adenovirus (non-integrative, dsDNA, transducing replicative and
non-replicative cells) to introduce the transient expression of CLN3. Serotype
with CNS tropism, but not exclusively.
Gene delivery approach using scAAV9
Chicken
β-actin
promoter-GFP
Mouse
Methyl-CpG-binding
protein 2-GFP
Because CLN3 regulatory
elements are not defined
6. 6
o Cln3Δex7/8
o 1-month-old mice:
• To more accurately depict human age of onset based on
mouse-human age equivalent estimates.
• When disease manifest (although phenotype is proven
very modest in mice before).
o Systemic (intravenous) delivery (1X) to enhance virus
biodistribution. AAV9 crosses BBB and transduces neuronal
and non-neuronal cells.
o Mice sacrificed after 5 months.
Gene delivery approach using scAAV9
7. 7
Results
o Widespread GFP expression with
both constructs detected
throughout the brain, eye and
spinal cord.
o MeCP2 primarily drove expression
in neurons and β-actin in
astrocytes.
8. 8
Results
o To them, Cln3Δex7/8 developed robust and persistent motor deficits beginning
with 2 months of age.
o Rotarod test:
o AAV9/MeCP2-hCLN3 reverses motor deficits in Cln3Δex7/8 mice -as early as
1 month after injection- whereas AAV9/β-actin-hCLN3 is ineffective.
9. 9
Results
o In a previous study they showed that microglia in Cln3Δex7/8 mice are primed
to be proinflammatory.
o Only AAV9/MeCP2-hCLN3 can significantly reduce microglia reactivity.
10. 10
Results
o Similar results obtained while evaluating reactive astrocytes.
o Only AAV9/MeCP2-hCLN3 reduces astrocyte activation.
11. 11
Results
o Effect of AAv9/hCLN3 transduction on lysosomal pathology by
immunostaning for LAMP-1 and subunit c of mitochondrial ATP synthase
(SCMAS).
o Only AAV9/MeCP2-hCLN3 reduces LAMP-1 levels and there is a trend
towards lower levels of SCMAS (mice are injected when inclusions are
already present).
