Adeno-Associated Viral Gene Therapy for Retinal Disorders - Gene Delivery and Therapy for Neurological Disorders - page 208

the human clinical trials for the treatment of Leber's congenital
amaurosis to drive RPE65 gene expression [ 15 ]. Following sub-
retinal delivery, transgene expression in RPE cells, photoreceptor
cells, and ganglion cells has been shown with both the CMV [ 29 ]
and CAG promoters [ 13 ]. However, in other tissues, AAV-driven
transgene expression has been demonstrated to be 137 times
higher when using the CBA promoter compared to CMV [ 30 ]. In
the brain [ 31 ] and muscle [ 32 ], there is evidence that the CMV
promoter may be silenced due to methylation in the longer term,
leading to transient or reduced gene expression.
For certain retinal degenerations, limiting gene expression to a
specifi c cell type may have benefi ts with respect to safety and func-
tional effi cacy, although in some cases overall levels of transduction
may be lower than with the use of a ubiquitous promoter. Regarding
expression in the RPE, the 1.6 kb RPE65 promoter has been suc-
cessfully used to limit gene expression to the RPE following sub-
retinal delivery in Briard dogs [ 33 ] and has also been used in
human clinical trials, although with questionable effi cacy [ 14 ]. In
targeting photoreceptors, the 292 bp human rhodopsin kinase
( hGRK1 ) promoter has been shown to be effective in driving high
levels of gene expression and specifi cally target rods and cones in
both mouse models and nonhuman primates [ 34 ]. In conditions
such as achromatopsia, it may be useful to limit gene expression to
cone photoreceptors alone. For this purpose, the 2.1 kb PR2.1 red
cone opsin promoter has been shown to specifi cally drive the gene
expression in L/M cones in the normal canine retina [ 35 ], and the
human cone arrestin promoter has shown expression in S and M
cones in mice [ 36 ].
Optogenetic strategies for restoring vision in the degenerate
retina might be optimized through gene delivery to bipolar cells.
A 200 bp enhancer region from the murine GRM6 gene fused to a
203 bp SV40 (simian vacuolating virus 40) core promoter has
been demonstrated to restrict transgene delivery to rod and cone
bipolar cells following subretinal delivery to the rd10 mouse model
of retinal degeneration [ 37 ]. In specifi cally targeting ganglion
cells, the 2.8 kb human connexin 36 promoter produced a selec-
tive transduction of foveal ganglion cells following intravitreal
delivery to the macaque retina [ 38 ], although the large size of this
promoter may limit its use.

2.3 WPRE

The woodchuck hepatitis virus posttranscriptional regulatory ele-
ment (WPRE) facilitates nuclear export of viral RNA. Incorporation
of this sequence into the 3 untranslated part of the AAV genome

has been demonstrated to increase gene expression by a factor of
1.8 times in brain tissue [ 31 ] and a sixfold increase in gene expres-
sion in cultured primary human fi broblasts compared to a vector
which did not contain WPRE [ 39 ]. AAV vectors containing WPRE

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