Download ABSTRACT Graduate Category: Health Sciences

Graduate
Category: Health Sciences
Degree Level: Ph.D.
Abstract ID#: 1306
A LONG-LASTING INTRANASAL GENE THERAPY APPROACH FOR PARKINSON’S DISEASE
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Amirah E. Aly , Brendan T. Harmon , Ozge Sesenoglu-Laird , Linas Padegimas , Mark J. Cooper , and Barbara L. Waszczak
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Department of Pharmaceutical Sciences, Northeastern University, Boston, MA and Copernicus Therapeutics Inc., Cleveland, OH
ABSTRACT
We have developed the first intranasal gene therapy approach with the potential to stop the progression
of Parkinson’s disease (PD), and possibly reverse its symptoms. Available drugs on the market alleviate
symptoms of PD but do not get to the heart of the problem, which is the progressive loss of dopamine
neurons. Our lab has found a way to harvest the potential of glial cell line-derived neurotrophic factor
(GDNF) as a treatment for PD. GDNF is a protein that activates survival and growth-promoting pathways,
protects dopamine neurons from injury, and restores their function. However, GDNF does not cross the
blood-brain barrier (BBB), so its use would require surgical injection into the brain. We are investigating
intranasal delivery of DNA nanoparticles (NPs) encoding GDNF as a way to bypass the BBB and allow the
brain to continuously produce GDNF. We have previously shown that our DNA NPs, developed by
Copernicus Therapeutics, Inc., transfect pericytes, which are cells that enwrap blood vessels throughout
the brain.
The goal of our current study was to determine the duration of protein expression after intranasal
administration of these DNA NPs. Our findings indicated that protein levels were highest at one week
after intranasal administration, and persisted over at least 6 months. Protein expression was elevated
throughout the brain, including in the brain regions involved in PD. These studies reinforce the intranasal
delivery of these NPs as a non-invasive, long-lasting means of gene therapy for PD and other CNS
disorders.
Intranasal delivery:
Intranasal delivery
7 days
30 min
Section A = olfactory bulbs (OB) / frontal cortex
Section B = rostral striatum
Section C = central and caudal striatum
Section D = midbrain
Section E = hindbrain
Section F = cerebellum
Section G = brainstem.
At 1 week, 3 months, and 6 months after
intranasal administration of pUGG NPs,
protein expression in the rat brain is always
significantly higher than in age matched,
saline-treated controls.
* p<0.05
RESULTS
Intranasal pGDNF protects dopamine neurons in the rat 6-OHDA model of PD
Unlesioned SN
BACKGROUND AND SIGNIFICANCE
• A lesion was produced by injection of 6-OHDA into
the SN on the left side of the rat brain.
1. What is Parkinson’s Disease (PD)?
NaCl control treatment group
DA
Naked pGDNF treatment group
• Brown color represents staining for tyrosine
hydroxylase (TH), the enzyme which produces
dopamine.
0.6
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0.5
Protein levels in rat brain were highest at 1
week post-intranasal administration, and
persisted at ~40% of maximal expression at
both 3 and 6 months.
0.4
0.3
0.2
* p<0.05
0.1
0.0
1 w eek
3 m onths
6 m onths
• Animals treated with pGDNF NPs had higher TH
staining density in the lesioned SN compared to
controls.
DA
• This indicates that more dopamine is being
produced on the lesioned side in rats given
intranasal pGDNF NPs.
2. What are the current therapies available for PD?
• Current therapies replace dopamine and diminish symptoms, but their effectiveness decreases over time.
• They do not stop or slow progression of the disease.
• New therapies that prevent damage to dopamine neurons, or rescue dying neurons, could stop PD in its
early stages.
3. Why is GDNF a promising therapy for PD?
GDNF is a “neurotrophic factor” that occurs naturally in the brain.
GDNF is reduced in the brains of patients with PD.
GDNF acts on dopamine neurons in the SN to promote survival and growth.
GDNF is potent; only a small quantity could help stop the progression of PD.
GDNF is a large protein and cannot cross the BBB. To reach the SN and the
striatum of patients with PD, it would need to be injected into the brain.
• GDNF is readily broken down in the body and would require repeated
doses.
• A safe and non-invasive means of delivering GDNF to the brain is needed to
harness its potential.
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4. Can intranasal administration be used as a way to bypass the BBB?
• Intranasally delivered proteins, nanoparticles and even cells bypass the BBB
to reach the brain.
• Transport to the brain follows two nerve pathways originating in the nasal
cavity:
1) the olfactory nerve
2) the trigeminal nerve
• Transported molecules reach brain areas involved in PD, the striatum and SN.
5. Can intranasal administration of the gene for GDNF be used as a treatment of PD?
• Intranasal GDNF gene therapy is appealing because it would generate a
renewable source of GDNF in the brain using a non-invasive route.
• The gene (DNA plasmid) must be compacted into nanoparticles (NPs) in order to
improve uptake into cells.
• pGDNF refers to the DNA plasmid that produces GDNF.
• pUGG refers to a second plasmid that produces GDNF linked to a fluorescent
marker, eGFP, used to aid in its detection.
Intranasal pDNA NPs result in protein expression in rat brain that is highest at 1 week and persists
at ~40% of the maximal level for at least 6 months.
Brain regions assayed
Saline
Naked pUGG or pGDNF
pUGG or pGDNF NPs
Lesioned SN
• A chronic and progressive movement disorder
• More than one million individuals affected in the US alone
• Common symptoms include tremors, bradykinesia (slow movement),
rigidity and postural instability
• Caused by the death of dopamine neurons in a brain area called the
substantia nigra (SN) which project to a brain region called the striatum
• Symptoms result from dopamine deficiency in the striatum
• Symptoms do not manifest until about 70% of SN dopamine neurons are
lost
RESULTS
EXPERIMENTAL DESIGN
whole brain weighted average
above background
(pg eGFP/mg protein)
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At 1 week, 3 months, and 6 months post
intranasal delivery of pUGG NPs, protein
levels were significantly elevated in both
the midbrain and striatum, the brain regions
involved in Parkinson's disease.
pGDNF NP treatment group
Nearly all cells transfected by intranasal pUGG are located adjacent to capillaries.
They are found adjacent to neurons only when neurons are close to capillaries.
* p<0.05
Conclusion
More eGFP-positive cells are located
adjacent to NeuN-positive neurons in
striatum of rats given pUGG NPs.
More eGFP-positive cells are adjacent to
TH-positive neurons in the SN of rats given
pUGG NPs than naked pUGG.
Thus, pUGG NPs transfect a higher percentage of cells close to neurons in the
striatum and SN, the target areas for an intranasal pGDNF gene therapy.
1. Intranasal administration of GDNF DNA nanoparticles
protects dopamine neurons in a rat model of Parkinson’s
disease
2. Most cells that produce GDNF are adjacent to capillary
endothelial cells and close to astrocytes enwrapping
capillaries. When these cells are adjacent to dopamine
neurons in SN, or neurons in any region, they are also
located close to a capillary.
3. These results provide evidence that intranasal delivery of
Copernicus’ pDNA NPs provides significant and prolonged
protein expression in rat brain, with highest levels at 1
week and continued expression over at least 6 months.
4. These studies demonstrate that intranasal delivery of
pDNA NPs is a promising, noninvasive, long-lasting
therapeutic approach for increasing protein levels in the
brain for treatment of CNS disorders such as Parkinson’s
disease.