Download Pharmaceutical Delivery of Antisense Oligonucleotides

Pharmaceutical Delivery of
Antisense Oligonucleotides
(ODNs)
Aws Alshamsan, RPh
Teaching Assistant
Department of Pharmaceutics
College of Pharmacy – King Saud University
Nucleic Acid Structure
Antisense ODNs?

They are sequenceselective nucleotide
sequences that can bind
to a target mRNA to
inhibit gene expression
i.e. to inhibit protein
synthesis.
Protein Synthesis
Antisense Strategy
Antisense Approach vs. Conventional
Therapy
Disadvantages of Conventional Therapy

Requires the non-rational approach i.e.
screening thousands of compounds to find an
active molecule.

Lacks specificity of action.
Why don’t we have an ODN for
every disease?

The main barrier to antisense strategy is now
achieving delivery of the ODNs in sufficient
quantities to the correct target sites of action
and for the desired time frame to achieve the
desired level of gene inhibition.

ODNs are polyanionic macromolecule (large
and charge).
Designing Biologically Stable ODNs
Designing Biologically Stable ODNs
Cellular Uptake



The cellular uptake of naked ODNs is
generally inefficient.
ODNs enter cells in vitro through pino- and
endocytosis.
The mechanisms of uptake depend on factors
such as ODN chemistry, length, conformation
and concentration, cell type, stage of cell
cycle, degree of cell differentiation and cell
culture condition.
Delivery Strategies
1.
2.
3.
4.
5.
6.
Liposomes and Lipoplexes.
Dendrimers.
Receptor-mediated endocytosis.
Local delivery to the brain.
Polymers.
Oral delivery.
Liposomes

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Cationic liposomes are the
most widely used system for
ODNs delivery.
They afford ODNs
protection and enhance
cellular uptake.
A significant optimization in
terms of charge ratio
between cationic lipid and
ODNs at a given dose is
essential for effective
delivery and activity.
Lipoplexes



They are lipid
complexes with ODNs.
Cationic lipoplexes are
preferable for ODNs
delivery.
The toxicity and serum
sensitivity of ODNLipoplex systems might
limit their widespread
use in vivo.
Liposomes and Lipoplexes

In vivo studies have shown that
they are rapidly cleared from the
circulation by the macrophages.

Modification with PEG (Stealth
Liposomes) reduces opsonization
and phagocytosis and resulted in
long circulating drugs.

Such systems may allow
enhanced passive targeting of
tumours, which have leaky
vasculature that is permeable to
small-size liposomes.
Dendrimers (DEN)



An advantage of DEN is the very
defined polymerization reactions
yielding monodisperes,
reproducible products.
Most of studies to date regarding
dendrimer delivery of ODNs have
been done using the
polyamidoamine (PAMAM)
starburst DEN.
DEN-ODNs complex have shown
reduce metabolic degradation and
enhance cellular uptake.
Percentage intact in serum was
10% naked ODNs to 85% DENODNs complex.
Receptor-Mediated Endocytosis




This process has the potential for
the effective targeting of ODNs.
Tumour cells has been a good
candidate for such targeting due to
its increased requirement for
essential nutrition.
Conjugating ODNs to a
transferrin receptor antibody
demonstrated a 3-fold increase in
cellular uptake compared to naked
ODNs.
Transferrin receptor is highly
expressed in brain capillary
endothelium, this technique is
useful for ODNs CNS delivery.
Local Delivery to the Brain


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The in vitro success of the previous technique targeting the
brain has not been transferable to the in vivo model.
This is may be due to the significant binding of ODNs to
plasma proteins forming a high molecular mass complex
which will be easily excluded from crossing BBB.
Direct intracerebroventricular injection is considered for such
cases but brain tissue toxicity may occur.
A promising development is the use of biodegradable
microspheres, which may overcome the toxicity problems and
provide region-specific and sustained delivery of ODNs.
Polymers
Advantages of Polymers Usage for ODNs Delivery:

1.
2.
3.
4.
Afford protection to ODNs.
Control the rate of the release of the encapsulated drug.
Can be fabricated as implantable devices for local
application.
Could be administered parenterally.
Factors Controlling Release Profile:

1.
2.
3.
The size of the microsphere.
The length of ODNs.
The molecular weight of the polymer.
Polymers


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Polylactides and co-polymers
of lactic acid and glycolic acid
P(LA-GA) have been evaluated
for ODNs delivery.
Entrapment of ODNs within
P(LA-GA) microsphere matrix,
provides nuclease protection.
Release of ODNs from the
microspheres in vitro showed
that ODNs were still able to
function.
Polymers


The release profile has been
shown to be “triphasic”.
Phase 1: “burst effect” after
48h.

Phase 2: More sustained
release.

Phase 3: Additional increased
release results from bulk
degradation of the microsphere.
Polymers

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Polyalkylcyanoacrylate (PACA)
nanoparticles have been
investigated for ODNs delivery.
ODNs are adsorbed onto the
charged (cationic) surface of the
PACA nanoparticles providing
efficient protection and cellular
uptake.
The toxicity of hydrophobic
cation and the production of
formaldehyde after degradation
may limit the in vivo use of this
system.
Oral Delivery

ODNs are rapidly degraded in GIT.

However, some studies have shown that some
P.O. ODNs are as effective as I.P. ODNs.

Some studies suggest slow passive diffusion
and transcytosis as possible mechanisms for
oral bioavailability.
Oral Delivery

ODNs are rapidly degraded in GIT.

However, some studies have shown that some
P.O. ODNs are as effective as I.P. ODNs.

Some studies suggest slow passive diffusion
and transcytosis as possible mechanisms for
oral bioavailability.
Fomivirsen Sodium (Vitravene)®



For the local treatment of CMV retinitis in AIDS
patients.
Dose 150-330 μg intravitreal injection.
Cleared locally by exonucleases 1-2 hr after injection.