1. Drug Targeting
Dr. Suresh Bandari

2. The main complications currently associated
with systemic drug administration are
• Even biodistribution of pharmaceuticals throughout
the body
• The lack of drug specific affinity toward a
pathological site
• The necessity of a large total dose of a drug to
achieve high local concentration
• Non-specific toxicity and other adverse side-effects.
Drug targeting may resolve many of these problems

3. Drug targeting is the ability of the drug to accumulate
in the target organ or tissue selectively and
quantitatively, independent of the site and methods of
its administration.
Drug administration protocols may be simplified;
Drug quantity may be greatly reduced as well as the cost
of therapy;
Drug concentration in the required sites can be sharply
increased without negative effects on non-target
compartments.

4. MAGIC BULLET CONCEPT OF
PAUL EHRLICH
•Drugs would be targeted by virtue of groups having affinity
for specific cells
• A ligand would confer specificity on a non-specific reagent

5. ‘‘MAGIC BULLET’’ Two components :
•The first one is recognizes and binds the target
•The second one provides a therapeutic action in this
target
Currently, the concept of magic bullet includes a
coordinated behavior of three components:
(a) drug;
(b) targeting moiety;
(c) pharmaceutical carrier;

6. The principal schemes of drug targeting include
•Direct application of a drug into the affected zone,
•Passive drug targeting (spontaneous drug accumulation in the areas
with leaky vasculature, or Enhanced Permeability and Retention-
EPR-effect),
•Physical targeting (based on abnormal pH value and/or temperature
in the pathological zone),
•Magnetic targeting (or targeting of a drug immobilized on
paramagnetic materials under the action of an external magnetic
field), and
•Targeting using a specific ‘ vector’ molecules (ligands having an
increased affinity toward the area of interest).

7. a) Macro molecular conjugates, b) Particulate drug carriers

8. Pharmaceutical carriers
• polymers
• microcapsules
• microparticles
• Nanoparticles
• lipoproteins
• liposomes
• micelles

9. Targeting Moieties
•Antibodies
•Lectins and other proteins
•Lipoproteins
•Hormones
•Charged molecules
•Polysaccharides
•Low-molecular-weight ligands

10. Brain Targeting
Delivery of drugs to the brain is a major challenge because it is
tightly segregated from the circulating blood by a unique
membranous barrier, the blood–brain barrier (BBB).
The brain and spinal cord are lined with a layer of special
endothelial cells that lack fenestrations and are sealed with tight
junctions that greatly restrict passage of substances from the
bloodstream than endothelial cells in capillaries elsewhere in the
body. These endothelial cells, together with perivascular elements
such as astrocytes and pericytes, constitute the BBB.
BBB is often the rate-limiting factor in determining permeation of

14. Characteristics of the BBB are indicated: (1) tight junctions that seal the pathway
between the capillary (endothelial) cells; (2) the lipid nature of the cell membranes of
the capillary wall which makes it a barrier to water-soluble molecules; (3), (4), and (5)
represent some of the carriers and ion channels; (6) the 'enzymatic barrier 'that
removes molecules from the blood; (7) the efflux pumps which extrude fat-soluble
molecules that have crossed into the cells

15. The factors affecting particular substance to cross BBB
Drug related factors at the BBB
•Concentration at the BBB and the size,
•Flexibility,
•Conformation,
•Ionization (nonionized form penetrates BBB)
•Lipophilicity of the drug molecule,
•Cellular enzyme stability and cellular sequestration,
•Affinity for efflux mechanisms (i.e. P-glycoprotein),
•Hydrogen bonding potential (i.e. charge),
•Affinity for carrier mechanisms, and
•Effect on all of the above by the existing pathological conditions

16. The physicochemical characteristics
•Log Po/w of the therapeutic agent, the rule of 2 is generally
accepted i.e. the value of log Po/w nearing 2 is considered optimal.
•However, increasing the lipophilicity with intent to increase
permeability would increase the volume of distribution (Vd) and
also the rate of oxidative metabolism by cytochrome P450
•Peripheral factors including systemic enzymatic stability,
•Plasma protein binding affinity,
•Uptake of the drug into other tissues,
•Clearance rate, and
•Effects of existing pathological conditions are also important.

17. •The lipophilicity of a given drug is inversely related to the degree
of hydrogen bond formation that occurs with surrounding water.
•The presence of certain chemical moieties in drug like terminal
amide, primary amines or amides and hydroxyl group favors
hydrogen bond formation resulting in a decreased lipophilicity.
•Thus for a compound to be transported through the BBB, the
cumulative number of hydrogen bonds should not go beyond 8–10.
Therefore for small drugs increasing lipophilicity i.e.
decreasing hydrogen bonds has a positive impact on capillary
permeability and drug transfer to the brain and for large drug
molecules with molecular weight above 400 Da or for those with
strong polarity, the capillary permeability will remain low
regardless of the lipophilicity

18. Several specialized transport mechanisms of solute transfer across
endothelial cells and into the brain interstitium are also present within
the BBB
Carrier system for monosaccharides, monocarboxylic acid, neutral
amino acids, basic amino acid, acidic amino acids, amines, purine
bases, nucleosides, vitamins and hormones.
The more lipophilic substances that are present in the blood can
diffuse passively directly through the lipid of the cell membrane and
enter the endothelial cells and brain by this means.

19. These solutes, and in many cases their metabolites, are actively
removed from the CNS by efflux transporters.
Various efflux transport pathways like P-glycoprotein and
active organic acid present in choroids plexus may also be
active in brain endothelial cells efflux systems are present in the
BBB to remove unwanted substances,
On the other hand the presence of the tight junctions and the
lack of aqueous pathways between cells greatly restrict the
movement of polar solutes across the cerebral endothelium

20. The molecules that can freely diffuse through this capillary
endothelial membrane can passively cross the BBB, and this
ability is closely related to their lipid solubility (lipophilicity/
hydrophobicity).
Practically all drugs currently used to treat brain disorders
are lipid-soluble and can readily cross the BBB following oral
administration.
The BBB also has an additional, enzymatic aspect: solutes
crossing the endothelial cell membrane are subsequently
exposed to numerous degrading enzymes within these cells.

21. These cells also contain many mitochondria – metabolically
active organelles – and active transport can significantly alter
both inward and outward transport for compounds.
The BBB is highly efficient and makes the brain practically
inaccessible to lipid-insoluble compounds.
Brain-delivery of such compounds, therefore, requires a
strategy to overcome the BBB.
Delivery of compounds such as neuropeptides or
oligonucleotides is further complicated by their metabolic
lability.

22. Functions of the BBB
•Firstly, maintaining internal environment of the brain, i.e.
maintaining brain interstitial fluid (ISF) and the cerebrospinal
fluid (CSF) composition within extremely fine limits, far more
so than the somatic extracellular fluid, so that the neurones can
perform their complex integrative functions.
•BBB protects the brain from fluctuations in ionic composition
that can occur after a meal or exercise, which could disturb
synaptic and axonal signaling.
•The barrier helps to keep the centrally and peripherally acting
neurotransmitters separate.

23. •A major function of the BBB is neuroprotection. Over a lifetime
CNS will be exposed to a wide range of neurotoxic metabolites and
acquired xenobiotics, which may cause cell damage and death. As
neuronal replacement is virtually absent in the CNS of mammals,
any enhancement of neuronal death will result in accelerating
degenerative pathologies and advance natural debilitation with age.
•Finally the continual turnover and drainage of CSF and ISF by
bulk flow helps to clear larger molecules and brain metabolites,
thus maintaining brain microenvironment

24. Strategies for Brain Targeting
Mechanisms for drug targeting in the brain involve going either
"through" or "behind" the BBB.
Neurosurgical or Invasive Strategies
BBB disruption
Disruption of BBB by osmotic means (Hyperosmolar solutions),
Intraventricular drug infusion
Intracerebral Implants
Biodegradable implants,
Physiologic based strategies
Psuedo nutrients eg: L-dopa
Cationic antibodies: These undergo Absorption mediated
trancytosis through BBB owing to positive charge.
Chimeric peptides

25. Pharmacologic Strategies
Chemical Delivery system
Nanocarriers for active targeting of the brain
Liposomes
Polymeric micelles.
Polymeric nanoparticles
Lipid nanoparticles .
Biochemically by the use of vasoactive substances such as
bradykinin,
Localized exposure to high intensity focused ultrasound (HIFU).
Cell-penetrating peptides and Brain transport vectors

26. Chemical Delivery Systems
Brain-targeted chemical delivery systems (CDSs) represent a rational
drug design approach that exploits sequential metabolism not only to
deliver but also to target drugs to their site of action.
By localizing drugs at their desired site of action, one can reduce
toxicity and increase treatment efficiency.
The CDS concept evolved from the prodrug concept in the early
1980s, but was differentiated by the introduction of target or moieties
and the use of multistep activation.
The cunning aspect of these brain-targeted systems is that, in addition
to providing access by increasing the lipophilicity, they exploit the
specific bidirectional properties of the BBB to ‘lock’ inactive drug
precursors in the brain on arrival, preventing exit back across the
BBB

28. CDSs are inactive chemical derivatives of a drug, being obtained by
one or more chemical modifications.
The introduced bioremovable moieties can be categorised into two
types.
A targetor (T) moiety is responsible for targeting, site-specificity and
lock-in; whereas modifier functions (F1...Fn) serve as lipophilizers,
protect certain functions, i.e., necessary molecular properties to
prevent premature, unwanted, metabolic conversions.
The CDS is designed to undergo sequential metabolic conversions,
disengaging the modifier function(s) and finally the targetor, after the
moiety has fulfilled its site- or organ-targeting role

29. Lock in mechanism of E2-CDS provided by introduction of a targetor moiety that exploits a 1,4-
dihydrotrigonelline (green) Trigonelline (red) type conversion. On hydrolysis trigonelline converts to
active drug.

30. During the past decade, the system has been explored with a wide
variety of drug classes, and considerably increased brain exposure
as well as brain targeting (i.e. brain vs systemic exposure) have
been obtained in several cases; for example, 3’-azido-3’-
deoxythymidine (AZT)-CDS, ganciclovir-CDS and
benzylpenicillin-CDS.
AZT-CDS administration in rats simultaneously increases brain
exposure 32-fold and decreases blood exposure threefold as
compared with AZT administration.
Among all CDSs, the estradiol chemical delivery system (E2-CDS)
is in the most advanced investigation stage. Following earlier
clinical trials (Phase I and II),

31. Molecular packaging: brain delivery of Neuropeptides
Delivery of peptides through the BBB is even more challenging than
delivery of other drugs, because peptides tend to be rapidly inactivated
by the ubiquitous peptidases.
For a successful delivery, three issues have to be solved simultaneously:
enhance passive transport by increasing the lipophilicity,
ensure enzymatic stability to prevent premature degradation, and
exploit the lock-in mechanism to provide targeting.
Successful brain deliveries have already been achieved using this
strategy for a Leu-enkephalin analog, thyrotropin-releasing hormone
(TRH) analogs and kyotorphin analogs

32. It is of particular significance for TRH delivery because
the corresponding process might require up to five or six
consecutive metabolic steps.
Therefore, selection of a suitable spacer moiety, which is
inserted between the targetor and peptide units to ensure
correct timing for targetor release, proved important for
the efficacy of TRH-CDSs.

34. HO NH2
HO
Dopamine
• Dopamine is also classed as a monoamine neurotransmitter and is
concentrated in very specific groups of neurons collectively called the
basal ganglia. Dopaminergic neurons are widely distributed
throughout the brain in three important dopamine systems (pathways):
the nigrostriatal, mesocorticolimbic, and tuberohypophyseal
pathways. A decreased brain dopamine concentration is a contributing
factor in Parkinson ﾕ s disease, while an increase in dopamine
concentration has a role in the development of schizophrenia.

35. The first group regulates movements: a deficit of dopamine in this
(nigrostriatal) system causes Parkinson's disease which is characterized
by trembling, stiffness and other motor disorders, while in the later
phases dementia can also set in. The second group, the mesolimbic, has
a function in regulating emotional behavior. The third group, the
mesocortical, is involved with various cognitive functions, memory,
behavioral planning and abstract thinking, as well as in emotional
aspects, especially in relation to stress. The earlier mentioned reward
system is part of this last system. Disorders in the latter two systems
are associated with schizophrenia.

36. In Parkinson’s disease, there is degeneration of the substantia nigra which produces the
chemical dopamine deep inside the brain

37. Since PD is related to a deficiency of dopamine, it would be
appropriate to administer dopamine
Problem: Dopamine does not cross BBB, since it is too polar
HO NH3+
Polar groups Mostly protonated
HO
to the
corresponding
Dopamine ammonium salt

38. If dopamine is too polar to cross the BBB, how
can L-DOPA cross it?
HO NH3+
HO NH3 +
Polar groups Mostly protonated
Polar groups Mostly protonated to the corresponding
HO O O
HO
to the corresponding ammonium salt
ammonium salt H
Dopamine L-DOPA
Polar group
L-DOPA is transported across the BBB by an amino acid
transport system (same one used for tyrosine and
phenylalanine)

39. Once across, L-DOPA is decarboxylated to dopamine by Dopa
Decarboxylase.This is an example of a “prodrug”, that is, a molecule that
is a precursor to the drug and is converted to the actual drug at an
appropriate place in the body.
In actual practice, L-DOPA is almost always coadminstered together
with an inhibitor of aromatic L-amino acid decarboxylase, so it doesn’t
get converted to dopamine before it crosses the BBB.
The inhibitor commonly used is carbidopa, which does not cross the
BBB itself. The inhibitor also prevents undesirable side effects of
dopamine release into the PNS, including nausea. H
HO H+
N 3
HO N
N 2
H
HO O HC
3
C 2
OH
O HO
H
L O A
-D P C r id p
ab o a

40. Polymeric nanoparticles suitable delivery systems for brain.
The mechanisms for nanoparticle mediated drug uptake by the brain include:
• Enhanced retention in the brain–blood capillaries, with an adsorption on to
the capillary walls, resulting in a high concentration gradient across the
BBB.
• Opening of tight junctions due to the presence of nanoparticles.
• Transcytosis of nanoparticles through the endothelium.
Furthermore, coating of these polymeric nanoparticles with polysorbate has
been reported to improve the brain bioavailability. Some of the proposed
mechanisms by which the polysorbate coating is effective, include:
• Solubilization of endothelial cell membrane lipids and membrane
fluidization, due to surfactant effects of polysorbates.
• Endocytosis of polymeric nanoparticles due to facilitated interaction with

41. But, there are various problems associated with the use of these polymeric
nanoparticles
• Residual contamination from the production process, for example by
organic solvents,
• Polymerization initiation,
• Large polymer aggregates,
• Toxic monomers and toxic degradation products,
• Expensive production methods,
• Lack of large scale production method and
• A suitable sterilization method e.g. autoclaving.
Considering the success of nanoparticles to pass through the BBB
and their limitation(s) especially toxicity and stability, another suitable
option for drug delivery into the brain would be SLNs.

42. SOLID LIPID NANOPARTICLES
SLNs constitute an attractive colloidal drug carrier system.
SLNs consist of spherical solid lipid particles in the nanometer range, which are
dispersed in water or in aqueous surfactant solution. They are generally made
up of solid hydrophobic core having a monolayer of phospholipid coating.
Advantages of SLNs over polymeric nanoparticles (and other delivery systems
like liposomes)
The nanoparticles and the SLNs particularly those in the range of 120–200 nm
are not taken up readily by the cells of the RES (Reticulo Endothelial System)
and thus bypass liver and spleen filtration.
2. Controlled release of the incorporated drug can be achieved for upto several
weeks. Further, by coating with or attaching ligands to SLNs, there is an
increased scope of drug targeting.

43. 3. SLN formulations stable for even three years have been developed.
4. High drug payload.
5.Excellent reproducibility with a cost effective high pressure
homogenization method as the preparation procedure.
6.The feasibility of incorporating both hydrophilic and hydrophobic
drugs.
7. The carrier lipids are biodegradable and hence safe.
8. Avoidance of organic solvents.
9. Feasible large scale production and sterilization.

44. Use of ligands.
Ligands or homing devices that specifically bind to surface epitopes or receptors on the
target sites, can be coupled to the surface of the long-circulating carriers.
Certain cancer cells over express certain receptors, like folic acid (over-expressed in
cells of cancers with epithelial origin),
LDL (B16 melanoma cell line shows higher expression of LDL receptors) and peptide
receptors (such as somatostatin analogs, vasoactive intestinal peptide, gastrin related
peptides, cholecystokinin, leutanising hormone releasing hormone). Attaching suitable
ligands for these particular receptors on to the nanoparticles
would result in their increased selectivity
Allen et al. postulated that the presence of specific ligands on the surface of
nanoparticles could lead to their increased retention at the BBB and a consequent
increase in nanoparticle concentration at the surface of BBB. While attempting to prove
their assumption, they prepared coated nanoparticles from Brij 78, and emulsifying
wax, with thiamine ligand (linked to DSPE via a PEG spacer).

46. Gene targeting technology & gene therapy of the brain
Many serious disorders of the CNS that are resistant to conventional
small-molecule therapy could be treated, even cured, with gene therapy of the
brain.
Current approach include delivery of the therapeutic gene to the brain
by drilling a hole in the head followed by insertion of the gene incorporated in a
viral vector.
The advantage of craniotomy-based gene delivery is that the gene can
be expressed in a highly circumscribed area of the brain with an effective
treatment volume of 1–10 μl. This craniotomy based delivery does not enable
the expression of the therapeutic gene widely throughout the brain or even to a
relatively localized area such as a brain tumor, which could have a volume
greater than several milliliters.
Viruses have been the vector of choice because the virus-coat proteins
trigger endocytosis of the virus into the target brain cell. The two most
commonly used viral vectors are adenovirus or herpes simplex virus (HSV).
The problem with both these viruses is that, because they are common, humans
have a pre-existing immunity. This immunity generates an inflammatory
response

47. Gene targeting technology
Craniotomy and viruses are first-generation brain gene delivery systems.
Gene therapy of the brain use delivery systems that are both noninvasive and non-
viral. A brain gene delivery system should enable widespread expression of a
therapeutic gene throughout the brain following a simple intravenous injection.
First, the exogenous gene packaged in a non-viral plasmid vector is
interiorized within a nanocarrier, much like exogenous genes are packaged in the
interior of viruses. This protects the therapeutic gene from the endonucleases in
the body.
Second, the nanocarrier is non-immunogenic and formed by either
natural lipids or other non-immunogenic polymeric substances.
Third, the nanocarrier carrying the exogenous gene is stable in the
bloodstream with optimal plasma pharmacokinetics following an intravenous
injection. (The rapid RES uptake can be blocked by pegylation. The pegylated
liposomes are stable in the bloodstream and have long blood circulation times).

48. Fourth, the surface of the nanocarrier is modified that triggers
transcytosis across microvascular endothelial barriers such as the BBB and then
endocytosis into target neurons or glial cells in brain. (Targeting through the BBB
and neuronal plasma membrane is accomplished by tethering the tips of 1–2% of
the PEG strands with a targeting monoclonal antibody (MAb) to form an
immunoliposome).
Owing to expression of the transferrin receptor (TfR) on both the BBB
and the neuronal plasma membrane, the use of an anti-TfR MAb causes the
pegylated immunoliposome to undergo transport through both the BBB and the
neuronal plasma membrane in vivo.
The liposomal lipids fuse with the endosomal membrane inside neurons,
which releases the plasmid into the cytosolic space of target neurons, where it can
then diffuse to the nuclear compartment. The only immunogenic component of the
formulation is the MAb and the immunogenecity of murine MAbs in humans can
be eliminated with genetic engineering and ‘humanization’ of the MAb.

49. β-Galactosidase histochemistry of a rat brain removed 48 h after a single intravenous
injection of a β-galactosidase gene carried by a plasmid that is packaged in the interior of
85 mm liposomes.
The surface of the liposome is covered by thousands of strands of 2000 Da (PEG), and this
stabilizes the liposome in the blood and prolongs the circulation time in the plasma.
Approximately 2% of the PEG strands that project from the liposome surface are tethered
to a monoclonal antibody that targets the transferrin receptor. This receptor is expressed
both on the brain capillary endothelium, which forms the blood–brain barrier in vivo, and
on the neuronal plasma membrane. Targeting the immunoliposomes to the transferrin
receptor enables transport across both the blood–brain barrier and the neuronal plasma
membrane in vivo. The use of gene targeting technology enables widespread expression in
the brain of an exogenous gene following a single intravenous administration of a non-viral
gene formulation.

50. Nice presentation
All the credit goes to Dr. Suresh Bandari
Thank you.