1. Gene therapy for Parkinson’s
disease
Under the esteemed guidance of:
Mr. Bimalendu Chowdhury
Astt.Professor
Department of pharmacology
RIPS
Presented by:
Utkarsh Alok
M.Pharm, 1st sem
Department of pharmacology
RIPS

2. INTRODUCTION ON PARKINSON’S DISEASE:
Parkinson’s disease is a progressive neurodegenerative disorder
whose main histo-pathological feature is the loss of
dopaminergic neurons in the pars compacta of the substantia
nigra with secondary striatal dopaminergic insufficiency[1].
Parkinsonism is a clinical syndrome consisting of four cardinal
feature:
1. Bradykinesia (slowness and poverty of movement),
2. Muscular rigidity,
3. Resting tremor (which usually abates during voluntary
movement),
4. Impairment of postural balance leading to disturbances of
gait and falling.[2]
Cognitive and psychiatric dysfunctions like dementia and
depression also accompanies the clinical symptoms.[8]

3. WHAT CAUSES PARKINSON'S DISEASE?
•Progressive loss of brain cells (neurones) in a part of the brain
called the substantia nigra, which produces the chemical
dopamine.
•As the cells die, less dopamine is produced and transported to
the striatum, the area of the brain that co-ordinates movement.
•Symptoms develop as neurones die off and dopamine levels
drop.

4. ETIOLOGIC FACTORS
•Aging: The annual incidence of IPD increases with age from
about 20 per 100,000 persons in the fifth decade of life to about
90 per 100,000 persons in the seventh decade of life, with the
usual age of onset at about age 60[17].
•Environmental factor: rural living, drinking well water, and
heavy metal and hydrocarbon exposure have small but
demonstrable contributions . Exposure to MPTP is capable of
inducing Parkinsonism in humans [3].
•Genetic factors: several gene mutations that can cause the
disease directly. Some of these mutations involve genes that
play a role in dopamine cell functions. Parkinson’s has
developed at an early age in individuals with mutations in
genes for alpha-synuclein, Parkin, PINK1, DJ-1 etc.

5. PATHOLOGY
1.NEURON LOSS- even mildly affected PD patients have lost
about 60% of their DA neurons, and it is this loss, in addition
to possible dysfunction of the remaining neurons, that accounts
for the approximately 80% loss of DA in the corpus
striatum.[3]
2.LEWY BODIES- major antigenic feature of Lewy bodies is
the expression of cellular proteins involved in protein
degradation. Presence of these antigens has been hypothesized
to represent efforts on the part of the cell to degrade the
abnormal protein aggregate [3].

6. PATHOGENETIC MECHANISM
Free radical and deficits in energy metabolism:
This theory is also referred to as the oxidant stress hypothesis or the
endogenous toxin hypothesis- initiated by H2O2, O2, which are
produced as a byproduct of mitochondrial oxidative
phosphorylation.
Protective mechanism- superoxide dimutase (and catalase,
antioxidants like ascorbic acid, tocopherol etc.
Mutation in SOD cause accumulation of aggregates of misfolded
mutated SOD, this can cause degeneration of motor neurons.

7. •The pharmacological and surgical therapies described above
aim to improve the symptoms of PD but none are proven to
have a significant impact on the underlying disease process with
respect to either slowing disease progression or restoring the
affected dopaminergic neurons.
• Gene therapy has distinct potential advantages over
conventional treatment modalities for PD as it could
theoretically be used to preserve or restore dopaminergic
neurons affected by PD through the action of neurotrophic
factors[41] or alternatively increase the availability of enzymes
required for dopamine synthesis[43].
•Although the disease modifying properties of these therapies
remains to be proven, they could potentially target the
underlying patho-physiological imbalances and may result in
much less fluctuation in response and a lower prevalence of
dyskinesias than conventional pharmacotherapy for PD.
Why gene therapy?

8. What Are Genes?
• Genes are carried on chromosomes and are the basic
physical and functional units of heredity .
• Genes are specific sequences of bases that encode
instructions on how to make proteins.
• When genes are altered so that the encoded proteins are
unable to carry out their normal functions, genetic
disorders result.

9. Gene therapy is a new approach to treating medical conditions,
which can be described as the use of genes as drugs.
Gene therapy can also be used to treat disorders where the
genetic cause is not known, or may not be caused exclusively by
genetic defects, such as Parkinson’s[44]. The carrier particle or
molecule used to deliver genes are called as vectors.
DEFINATION: Gene therapy is the process of inserting genes
into cell to treat diseases, where newly introduced genes will
encode proteins and correct the deficiencies[45].
What Is Gene Therapy?

10. What Is Gene Therapy?

11. •In most gene therapy studies, a "normal" gene is inserted into
the genome to replace an "abnormal," disease-causing gene.
•A carrier molecule called a vector must be used to deliver the
therapeutic gene to the patient's target cells.
•The most common vector is a virus that has been genetically
altered to carry normal human DNA.
•Viruses have evolved a way of encapsulating and delivering
their genes to human cells in a pathogenic manner.
•Scientists manipulate the virus genome to remove disease-
causing genes and insert therapeutic ones.
•Target cells, such as the patient's liver or lung cells, are
infected with the viral vector.
How Does Gene Therapy Work?

12. How Does Gene Therapy Work?

13. Approaches for gene therapy
There are two approaches to achieve gene therapy:-
1. Somatic cell gene therapy.
2. Germ cell gene therapy.
Somatic cell gene therapy:
The therapeutic genes are transferred into the somatic cell or
body, of a patient. Any modifications and effects will be
restricted to the individual patient only, and will not be
inherited by the patient's offspring or later generations.
Somatic gene therapy represents the mainstream line of current
basic and clinical research, where DNA is used to treat a disease
in an individual[47].

14. Germ cell gene therapy.
Germ cells, i.e. sperm or eggs, are modified by the introduction
of functional genes, which are integrated into their genomes.
This would allow the therapy to be heritable and passed on to
later generations. Although this should, in theory, be highly
effective in counteracting genetic disorders and hereditary
diseases.
The genetic alteration in somatic cell is not carried to the next
generation. Therefore , somatic cell gene therapy is preferred
and extensively studied with an ultimate objective of correcting
human disease [45].

15. •Problems With Gene Therapy.
•Short-lived nature of gene therapy- patients will have to
undergo multiple rounds of gene therapy.
•Immune response- risk of stimulating the immune system in a
way that reduces gene therapy effectiveness is always a potential
risk.
•Problems with viral vectors- viruses, the carrier of choice,
present potential problems to the patient, like toxicity, immune
and inflammatory responses, and gene control and targeting.
•Multi-gene disorders- most common disorders, such as heart
disease, high blood pressure, Alzheimer's disease, arthritis and
diabetes, are caused by the combined effects of variations in
many genes.
•Chance of inducing a tumor (insertional mutagenesis) - If
the DNA is integrated in the wrong place in the genome, for
example in a tumor suppressor gene , it could induce a tumor.

16. As such, gene therapy involves a great risk. There are several
regulatory agencies whose permission must be sought before
undertaking any work related to gene therapy. Recombinant
DNAAdvisory Committee (RAC) is the supervisory body of
the National Institute of Health (NIH), USA, that clears
proposals on experiments involving gene therapy[45].

17. TYPES OF GENE THERAPY
1.Ex.vivo gene therapy.
2.In-vivo gene therapy.
Ex.vivo gene therapy: Cells are modified outside the body and
then transplanted back in again. Ex vivo gene therapy uses gene
transfer techniques to produce genetically modified primary
cells or cell lines for transplantation into the brain of
Parkinsonian individuals.
This procedure involves patients own cell for correction and
then their return back to the patient. This technique is therefore,
not associated with adverse immunological responses after
transplanting cells. Ex-vivo gene therapy is efficient only, if the
therapeutic gene is stably incorporated and continuously
expressed, this can be achieved by use of vectors.

19. In vivo gene therapy: In vivo gene therapy for the CNS
consists of the direct intra-cerebral injection of genetic
material using appropriate vectors[54].
The success of in-vivo gene therapy depends upon the
following parameters[45]:-
•The efficiency of the uptake of the remedial (therapeutic gene)
by the target cells.
•The expression capability of the gene.
Gene transfer to the brain by using viral vectors offers the
advantage of being less invasive than transplantation techniques,
leaving the striatal circuitry undisturbed by cellular implants
and eliminating risks of unwelcome host immune responses or
tumour formation[53].

21. Vectors for gene delivery
The carrier particle or molecule used to deliver genes are called
as vectors.
The ideal vector system would have the following characteristics:
 an adequate carrying capacity;
 to be undetectable by the immune system;
to be non-inflammatory;
 to be safe to the patients with pre-existing lung inflammation
to have an efficiency sufficient to correct the cystic fibrosis
phenotype
to have long duration of expression and/or the ability to be
safely re-administered.

22. There are two categories of delivery vehicle (‘vector’)
1.Non-viral vectors
2.Viral vectors
Retrovirus Herpes
Simplex V
Adenovirus AAV Liposo
me
DNA Polymer
Integration Yes Non Non Yes Non
Expression Stable Transient Transient Stable Transient
Transfection Efficient Efficient Efficient Low Low
Immune Response No Yes High No Yes Yes or No No
Generally, viral vector system show higher gene transfer
efficiency than non-viral gene carrier system, but viral
systems have potential risk of wild type virus regeneration,
immunogenecity and cancer formation.

23. Vectors associated with gene therapy for parkinsons:-
Overexpression of GDNF mediated by the lentiviral vector has
conferred some protection of the nigrostriatal dopamine
terminals against toxic insults[63].
Ad-GDNF can protect dopaminergic neurons and improve
dopamine-dependent behavioral function in young rats with
progressive 6-OHDA lesions of the nigrostriatal projection[71].
 AAV-2, when administered locally, it transduces only neurons
within the central nervous system and is particularly efficient in
brain regions known to be involved in the pathophysiology of
PD, such as the globus pallidus and substantia nigra[75].

24. Strategies for the rationality in the gene therapy trials in
Parkinson’s disease
PD has a complex pathophysiology that is by no means fully
understood and involves multiple brain structures and signalling
pathways. There are three broad approaches to selection of a
therapeutic target[81].
1. Restoration of DA synthesis in the dorsal striatum;
2. Modulation of activity in the basal ganglia downstream of the
striatum; and
3. Modification of disease progression by neuroprotection.