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Dec 5, 2012 (4 years and 6 months ago)

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Dissertation Title:

Prospects of nanocarrier in cancer immunotherapy.



Selvi Tamil Selvam


This work is submitted in partial fulfilment of the
requiremen
t of MSc in
Cancer Immunology and Biotechnology,

The University of Nottingham.


Supervisor: Dr. Judith Ramage.

Date:
10.03.2011.


Contents:

Abstract

Introduction

Problems associated with cancer vaccine

Motivations for using Nanocarriers in
delivery of vaccines


Size matters

Internalization of nanoparticles depends on its size limitations

Lymph node as an target

Cross presentation

Complement activation

Enhanced permeability and retention effect

Classification of Nanocarriers

Enhancers for
Nanocarriers

Conclusion, opportunities and challenges

References

Abbreviations










Abstract
:


The dissertation focuses on new field nano
-
biotechnology, which has worked out on
description of how to fabricate nanodevices that mimics salient features of
natural
biological environment
which can be used
as a platform to treat cancer using
immunotherapy based
product vaccines.
This
study

examines the nanocarriers which has
been exploited as a platform for delivery of biotheapeutic or drugs which are well
est
ablished over the decades and claims some of the challenges in decoding them to an
effective nano
-
based cancer vaccine delivery system
.

In addition the dissertation
investigates whether NP can be used as a vaccine delivery system by means of targeting
lymp
h nodes, APC, activation of immune components and their role as an adjuvant.


Introduction:

The immune
organization

has the
capability

to
attack

the cancer and pre cancer cells. On
the other hand tumour cells have evolved to learn the escaping mechanism of
immunoselection. Cancer immunotherapies have developed the
strategy

of cancer
vaccines which can be applied in treatment with the existing canc
er.

Modern

science
has
helped us in unravelling more and more
secrets about

the human body and the
roles of
our system and currently
material science is
exploring

into the molecular level. The
marriage of these two
fields

is considered to be nanotechnolog
y.
Nanotechnology has
revolutionize
d the

medical diagnosis

and therapy for various
diseases
. Developments

in
these fields
have weighed in to innovative
nanoscale
-

an

methodology

that

brings

new
hopes

to cancer patients.

This dissertation is based on the mo
st salient hypothesis that
nanotechnology based

fabrication are
effective in vaccine development as the
nonsoluble nanoparticles

could provide creating a depot

at the site of injection ,controlled,
slow release of antigens, and safeguard at
in vivo

environment
.(24,25).

Superlatively
, a nanocarrier applicable for cancer delivery system
should crew in
good
physical stability within the biological
milieu

w
here it has to travel,
be
biocompatible,
and
should display good

affinity and selectivity for its
fina
l destination
.

The primary goal for
an

effective

cancer

vaccine is the delivery of antigen epitopes to antigen
-
presenting cells,
followed by processing, presentation and finally induction of an immune response
against target.

The

antigen presenting cel
ls

are dedicated
to take up and process antigen
for the induction of CD8 and 4

cell.

Presence of an adjuvant is also an important criteria.

Hence the

uptake
of antigens by APC

followed
by
gathering

of these cells

to t
he lymph
nodes and
sparking off
the

APC

to mature are the
vital steps

for the initiation
of potent
immune responses
.
Thus these are the
steps that call

for
attention

to be evaluated
for
vaccine

delivery system.

Problems associated with vaccine delivery:

Some of the major problems associated
with delivery of vaccines are :




Nonspecific

interaction with cell membrane,



Degradation of the vaccine due to proteases enzymes,



Antigen processing and presentation,



Duration of vaccine circulation,



Retention at the site of administration,




Acute or chro
nic toxicity,



Challenges in the aspects of safety and utility of viral vectors in regards to DNA
vaccine
,



Potential immunological risk with a targeted tissue for adenovirus or
insertional mutagenesis for retroviruses when they are used as
transfection tool

is witnessed.



Deaths are actually reported in clinical trials due to viral vectors which are
a major setback.





Physical
method of delivery possesses

the problems of tissue damage.

Motivations for using Nanocarriers in delivery of vaccines
:

The principal attribute of
NP

system is their prospective quality to enhance the
accumulation of therapeutic agent in tumour
cells, protecting

the agents and not
disturbing the healthy cells.
The p
aradigm of using
NP

in

pharmaceutical
is
to increase
the inv
ivo efficie
ncy of the anti
-

cancer
drugs the

same
can

be
mimicked

for efficient
deliv
ery of cancer vaccines as wel
l. The ma
jor drive behind using
nanocarriers:



They

possess

the capability of prolonged circulation in the blood hence it
increases the chance
of efficient

release
vaccine
in close propinquity

of tumour
.





H
as the capability to attach to the
cell membrane
or

target tissue via s
urface
attached specific ligand (Skubitz KM et al,2004)



Release of the entrapped vaccines in response to local stimuli fo
r eg: Under
abnormal pH or temperature at cancer site and this property can be provided by
surface attached sensitive components.



It has the ability to bypass the lysosomal degradation by tailoring the outer
surface with cell penetrating peptides and hence

reduction of toxicity.



T
he size is less than 200 n
m that
aids in efficient delivery to the target cell




Since

its smaller in size, has enormous surface area.


Nano carrier based products which had
successful

interaction with our immune
system:

The below
table consists of nan
obased delivery system with encapsulated

drugs or other bio therapeutics but have successfully targeted immune
system, less

toxicity

and have even entered clinical trials.
Since they were successful in targeting
immune system,
nanopar
ticles pledge

to be an ideal candidate for targeted delivery of
cancer vaccine
.







Product

Name

Material

Size

(nm
)

Treatme
nt

Administr
ation

Route

Phase

and

Biotech
company

Target of
Immune
system

Doxil




Biovant

Pegylated

Lysosome



Nano sized
calcium
phosphate

75
-
80



20
-
50

Metastatic

Ovarian
cancer
.


Vaccine
developm
ent

i.

v




i.m

Phase III
Approved

Ortho
biotech

Phase I
completed


Biosante
pharmaceut
icals

Complement
activation.

(
1)


It has adjuvant
properties
Elicits immune
response to
DNA and
protein based
antigens
stronger.


It has less
hypersensitivity
reactions.


CYT
-
6091
(Aurimmu
ne)




Bioconjug
ated

nanoparti
cles





DF1









C3









TNF
-
alpha
PEGlyated
colloid gold
particle
.


Quantum
dots with
tri block
copolymers




Dendritic
Fullerene








Fullerene

derivative

33





10
-
15







4.7
-
5







20
-
24

Solid
tumor





Cancer








Chemopro
tection








Chemopro
tection

i.v






Subcut








I.V

Phase
-
I

Cytimmune

S
cience
.



Preclinical

Emory
-
Georgia
Tech
Nanotechnol
ogy
Centre
.


Preclinical


Carbon
Nanotechnol
ogy.





Preclinical

Carbon
Nanotechnol
ogy

Decreases TNF
toxicity targets
only tumour
area but not
healthy area.
±


Accumulation
in spleen. No
toxicity was
found in
Immune cells.

(Unger ec et
al).

Inhibition of
collagen
-
induced platelet
aggregation;


toxicity to RBC
is not found
in
vitro
ΐ



Inhibition of
collagen
-
induced platelet
aggregation;


toxicity to RBC
is not found
in
vitro
ΐ






Numerous
amounts

of n
anoparticles have proven to be a

promising carrier for
resourceful targeted delivery to the lymphatic system, arterial wall, lungs, or liver
[Brannon
-
Peppas and Blanchette, 2004].
Many invitro studies have proven that these
nano carrie
rs are not affected by the serum and hence it can be administr
ated in to
systemic circulation
which does not lead to any blockage or aggregation in blood
capillaries owing to its size.


Size matters:










Nanoparticles immunogenicity is gaining attention because in many studies they h
ave
improved the immunogenicity either being an adjuvant or being an antigen themselves
when conjugated with an weak antigen

owing to its size
.
The
dimension
of

the

antigens
is an important

cri
ter
ia for the

proficient
uptake by APCs

and it varies for different
vaccines
.
The matter of geometry, shape, size and kinetics
of antigen fluctuates as well.
The

protein or viral subunit antigens are the smallest of about < 10 nm.

They

form

a
large

constituent parts

or aggregates

once they are

added with adjuvants
. Well defined
super molecular

particulate antigens such as virus like particles and nanocarriers are
larger than 20
-
200 nm.

The presentation of antigens in milieu of nanocarriers,
microparticles, freuds adjuvant, oil in
water
(MF59

AD
JUVANT), alum adjuvants and
whole cell vaccines are the largest of about 100nm
-

20

μm .
It is found that efficiency of
APC can be lifted by
maintaining the

vaccine size between 10 to 10000nm which could
be
facilitated

by encapsulating the vaccine in a nano
carrier. To

support the claim

that
size really does matter a
comparative animal

model study demonstrated by using nano
polystyrene

particles

of was done
and, in

particular
40
-
50nm particle

could promote
better CD8 and 4 response when compared to 0.5
µm by
Theodora fifs et al(2004)
inducing hig
h amount of INF
-
G and Ab
titers

in mice.
Another important observation was
reported by them is that these size promoted DEC205+ CD40+CD86 DC/APC cells but
they have not incorporated any danger signals in the
NP
. We ca
n
hypothesis

from this
that danger signals might have been built by its own by
NP

and it might have prompted
DC to react appropriately owing to size in range of viruses.

Large number of studies once
again confirms that
NP

can be used as an adjuvant.
It was

observed by another group

eg
polyme
thylmethacrylate
NP

have

prompted

long
-
lasting ab titres in HIV2 whole virus
vaccine in mice

models
.

They have compared their vaccine study with traditional
aluminium sulphate (alum) as an adjuvant with
NP

encapsulated v
accine
without

any
adjuvant
and found that ab
response was 100 times higher against the disease
.
Target
compartment: DC
,
Lymph node
,
activation

of
complement
system, to tumour.

Toxic Compartment
.

Prolonged circulation of
vaccine, denature, cross
reaction, inefficient
targeting,etc




CANCER VACCINE


[steinfenker et al].

Hence by using n
anocarrier we can speculate because of
NP

size
and
taking a clue from various studies fabrication of
NP

in range of pathogens
may act as
a

driving force to be preferentially taken up by the DC and as well and could act as PAMPS
to activate DC.

Comparative analysis between
NP

and pathogens
:

1
100
10
,
000
100
,
000
nm
Nano
-
particles Delivery system
with Vaccines
Pathogenic agents
10
10
,
00
Liposomes and virosomes
Microparticles
VLPs
Soluble
antigens
Viruses
Bacteria
ISCOMs
Size in relations to Pathogens
Efficient entry into lymph
vessels
Efficient uptake by APCs


The n
anocarriers have large surface area and are catatonically
charged
.

They
possess

hydrophobic
charge
which mimics cell’s

receptor
-
interacting properties

and
this
phenomenon
augments healthier
communication
between APCs and nano
particles
.

Vaccine uptake
Effe
ct and Efficiency of
and transport:


NP

interacts with the tumour as in the fashion of ligand

receptor, and bound carriers are
internalized b
efore the vaccine is released.
The vaccine uptake and transport element of
the dissertation
projects on the
comparative

analysis with nano carrier which deals with
the gene or drug delivery kinetics. With supporting evidence from literature

with respect
to carrier size, charge, elicitation of immune response and delivery methods to tumour
site, this dissertation

proposes an initiative to use nano based carrier
for the targeted
delivery of vaccines
.

The mode of administration of vaccines is usually subcutaneous or intramuscular. The
transport of the vaccine from the peripheral tissue to lymphoid organs is an impor
tant
strategy which has to be exploited in vaccine design. The size of nano carrier vaccine
delivery system also
disturbs

the kinetics of the lymph drainage system.

Internalization of nanoparticles depends on its size
limitations
:











The mode of
targeting

the
tumour

environment can be achieved by two modes either by
EPR
effect and

this process is a passive

and nonspecific mode of application. The second
mode of targeting is most advanced method which
are extensively

investigated in
oncology is by active targeting, by attaching specific ligand to the surface of
NP
. For
example many
cancer cells over expresses transferins and folins because of the
metabolic
demands, which makes conjugation
of these
over expressed

compounds or
antibodies
to these receptors

proves to be a successful targeting system (
26,27,28).

Usage of whole mAB’s is

more advantageous when compared to fragments since they
have two binding sites which can give rise to higher avidity. Fc region of the antibody
can bind to immune cell receptor as well as the receptor of normal cells like
macrophages. When it binds to the
macrophages it leads to increase in immunogenicity
and hence liver and spleen can efficiently take up the nano carriers. Although the
efficacy of their binding and targeting the cancer sites are proven their lethal side effects
are also been reported due t
o nonspecific binding by Arnold Dm.et al. Proteomics and
genomics

tools, though seems to identify the appropriate target, to date but yet no
effective candidate has been identified.



Lymph node

and DC

as an Target:

Many supporting evidence have shown that

p
articles of less than 20
nm reach the
lymphoid organs directly with in hour of administration.

Reddy St et al(2007) have used
nanoparticles
of varying size between 100 and 20
nm

with antigen and successfully
targeted the lymph node
with 20 nm
where there
was

large concentration of
dendritic
cells

.These DC


are phenotypically and functionally immature and hence can process the
antigen which is in contrast with targeting the p
er
ipheral DC
. Hence using nanocarriers
deems
to
be an

advantage for transporting t
ransport
vaccines to lymph node
directly
and avoid the premature antigen presentation

for Cancer vaccine
.

Ideally a vaccine with
clinical benefit has to acquire two components, one that encodes the specific antigen and
secondly it has to enhance the immune

response with the aid of APC.


Attention for
specific delive
ry of antigen to DC has been witnessed
only
lately
. The new strategy use
can be
shifted to nano carriers since they can confer longevity of the product which they
are encapsulated with as witness
ed in many pharmaceutical delivery system. It can also
increase the prospect f
or Dc uptake and processing. Many supporting evidence across
the
literature
have shown
that
polymers

for eg
PLGA could bind and encapsulate proteins,
besides

DNA
(24)

and

since
these particles are large enough
, they can be taken up

by
Mode

Size

Phagocytosis

M
icropinocytosis

Caveolar mediated endocytosis

Clathetrin mediated endocytosis



large particles

>1 micro
meter

60 nm

120 nm.



phagocytosis
(25,26,2
7). For the efficient targeting of DC, many engineered
nanoparticles are fabricated for eg ligand binding mannose receptors; controlling their
size and binding of specific antibo
dy specific to DC like anti
-
CD11c are being reported.
These discoveries appear
s

judicious

to hypothesize that
we can achieve directed role of
APC by NP carriers

and their
following

enhanced

achievement, to

encourage

immunity
response to vaccination when co
mpared to cancer vaccine without delivery vehicle.

Another interesting development was reported
by Omar

A Ali et al 2009

where they have used biomaterials alternatively to program specific cells without any
transplantation. They have releas
ed the cytokines
first from

PLGA
and have successfully
activated the DC and subsequently presenting the
tumour

antigens

to the target site
lymph node
. This approach has generated potent, antitumou
r responses where they
have reported 90% survival in animal
models wh
ich usually dies from cancer in span of
25 days. The tumour antigen used in this experiment consists of TAA which could dilute
material presentation to APC. Hence it may be advantageous to use purified antigen.

The major drawback is

that we have n
ot completely understood the mechanism of
NP

in
regards with increase in antigenicity or property of adjuvants.


Studies also put forward
that nanoparticles can intensify allergic
reactions (
29
) and
, hence
, careful preclinical
categorization is essential
.
Broad
NP

activity re
lationship research can aid
further
to
comprehend the

limits

that
can regulate

the antigenic and adju
vant
belongings

of NP.

Cross presentation

is
important
phenomenon

which are

being highly used in
therapeutic cancer vaccine

where DC ar
e involved in presentation of
foreign

antigen on
MHC I

to induce the priming of CD8 T cells
. The processes of
presenting

an exogenous
peptide on to MHC I is cross presentation

(19
, 20
)
.

It has been validated that
cross
presentation is highly feasible by
nanocarrier and this claim is proven by Emori t el
al(2004) where they have shown

the release
of antigen which were encapsulated in NP are
released

by either leucocytes or epithelial cells
.


These

antigen bearing
nano platform could
successfully
elicit th
e strong CTL response

(11,12)

.

However transferring of antigens in
draining lymph node is not clear
from their
experiment,

we can hypothise that by usisng
inert
NP
, it could be facilated by two possiblites either



By direct contact between DC with lymph n
ode DC or



Release of
NP

release of
NP

from lysosomal compartment that are
taken up by
resident Dc

This is true for a large number of biodegradable materials such as liposomes,
(15)

ISCOM,(16) and
polystyrene

materials (13,14).


Complement activation:

Activation of complement by nanocarriers based delivery of
vaccine may be beneficial or an limiting factor depending on its proposed application.
Complement activation by systemically administrated nanocarriers may lead to
hypersensitivity reactions, hence

it is important to tailor the surface properties to reduce
the reaction to acceptable level.
Chanan khan

and colleagues

group has reported that
by using nano liposomes in their clinical studies,
particles have

activated complement
pathway that lead
to hy
persentivity

reaction. Another study states that nanoparticles
carried vaccine which were up taken by dendritic cells followed by activation of Tcells
and generation of antigen immune response was dependent on complement activation
by nanoparticles by
Redd
y st et al(2007).

Th
us both studies and report are
contradictory

to each other but a notable phenomena is that they have used different mode of
strategies for injecting NP.


Most of the research have typically avoided the complement activation in order to
minimize the implant rejection which could clear the systemic drug delivery carrier . But
in delivery of cancer vaccine we can attempt the opposite so that nanocarriers can act as
an adjuvant and generate danger signal. Nanocarriers like



Polypropylene sulf
ide nanoparticles,



Lipid
Nano capsules
,



Cyclodextrin
-
containin
g polycation
-
based
NP




Polystyrene

Nano spheres

have proven that charged particles are more promising
in activation of

complement

when compared to neutral part.



The nanoparticles can be explored to validate the above mentioned concepts of
delivering vaccines, acting as an adjuvant to lymph node and complement activation to
mature the cells due to its ease of fabrication. Despite of the advantages, we need to
qu
estion in regards to toxicity, elimination or the molecular interaction has to be to
addressed in order to use nanocarriers as an effective platform.










Enhances Bcell and
Nonspecific

clearance of pathogens.


Hypersentivity reaction



Tcell activation.



Dc activation.








Humoral and



Cellular Immune response










Enhanced permeation and retention:

Complement activation

Nano based carrier of vaccine

Systemic
administration
Avoid

May
be beneficial.

Sc
or Id routes.



When vaccines are administrated intravenously, it is been transported by the blood to
heart . From the heart it goes to lungs where blood is oxygenated and now vaccines will
be transported to arteries. Since arteries carries the vaccines and circulates eve
rywhere
in the body it’s important to target the vaccines preferentially to tumor prone area. The
efficient delivery of vaccines can be done at the level of capillaries when the blood starts
flowing from arteries to capillaries. Only at the molecular leve
l of capillaries the
molecules from blood can leak out. The
endothelial

cells are genetically stable and do
not become resistant to the
therapy [
Bohem et al 2007] hence we can take advantage of
the leaky vasculature system.

The dimensions of capillaries a
re 7
-
15 microns which
allows the red blood cells to flow through in normal
hierarchy.
But in tumor capillary the
endothelial cell linings have breaks which can allows the vaccine to leak out and hence it
acts as a major advantage for efficient delivery o
f vaccine.
Delivery of therapeutics by
these phenomena

is called enhanced permeation and retention was demonstrated by


Matsumura and
Maeda and

these phenomena

are

highly applied in pharmaceutical
carriers
.

Since these have given promising
results,

we can

take advantage of
this proven methodology

for targeted delivery of vaccine in solid tumour area

and subsequent cross presentation is
possible
.
The literatures have shown that usually these breaks are in range of 0.5 microns
and hence a nano carrier with
vaccine can seep out efficiently where red blood cells
cannot.


Though EPR effect is an important criteria used in
pharmacology

NP
, still the question
remains to what extent accumulation of
NP

in tumours accounts for the improvement in
antitumor activity.
Several

speculative mechanism , one such possibility

we can
hypotheise that
NP

release of vaccine is possible by the interstiti
al fluid surrounding the
tumour as these fluids contains oxidizing agents, lipases etc.




EPR EFFECT











Nanocarriers


























Normal Tissue


























The nanocarrier are not able
seep through

the normal vasculature system where as in
the below diagram
NP
s gather because leaky vasculature which leads to

EPR eff
ect.



Leaky
vasculatur
e







Oral delivery

of vaccines would be a beneficial one if the patients require
repetitive,
constant

delivery of vaccines.

But for the vaccines to remain effective, it has to survive
the p
H conditions. Many research

are being carried out in developing the smart carrier
system which are pH sensitive and that take advantage of the pH change in the body
especially in gastrointestinal tracts. Kim B et al
2009
have reported pH sensitive
hyrdogels made of poly methyl acryl
ic acid grafted wit
h ethylene glycol
where the gel
shrinks
at low pH of 2 due to the establishment

of interploymer complexes, whereas at

physiological pH 7 the NP
swell 3
-
25 times and releases its cargo
at targeted site
.


Size is considered to be of utmos
t importance and it was demonstrated repeatedly that
NP

are processed more efficiently in to cell

[Panyam and Labhasetwar, 2003 a] or
accumulated in tissue [Jani et al. 1990; Lamprecht et al., 2001] than larger particles.
Uptake and delivery of the vaccine
s could be classified
as passive

targeting based upon
their enhanced permeability retention and active targeting which makes use of
tissue/cell specific targeting which leads to accumulation of nanocarriers at specific hot
spot [Kim and Nie,2005].

Nano car
rier
fate is

determined by the immune system
depending on their composition. Biodegradable materials

are digested and cleared from
the body
,

(8,9)

while

nonbiodegradable particles accumula
te
(10).

NP

tend t
o aggregate themselves and are

capable

of forming m
icelles by themselves
instead of just

congregatition.

Choosing of amphiphilic component has several
advantages

as it is not degraded by proteases.

The
NP

can be spun in any orientation or
size depending on our vacc
ine and target
. Biodegradation

of the car
riers usually appears
to happen at very slow rate

and

they appear not to interfere with the normal cell
function to a greater significant effect.



Possible mode of release of Vaccines from nanocarriers:



Owing to polymer erosion and

degradation.



Diffusion
through pores of particles.



Discharge

from the surface of carriers.



Application of magnetic or sonic field can yield pulsed delivery.


Classification of NANO
CARRIER system:

Varieties of carrier systems are in development for efficient treatment in cancer
and
much of the biomedical research is done in delivery of drugs. The list brings in to group
of

few

nano particles which can be engineered to the level of reality to microbial
pathogens for efficient delivery of cancer vaccines. Many research studies have

even
validated that immature DC has a potential to uptake a variety of nano particles such as
latex, polystyrenes, PLGA and liposomes.

Many NP

have shown to be an promising tool
for the presentation of antigen by both MHC class I and II

on surface o
f DC
for example Y
waecklerle et

al in 2005 PLGA encapsulated antigen was processed in two
independent way with final
outcome of simultaneous activation of both CD8 and CD4 T cells.


Nanoparticles

Material


Merits

Demerits


PLGA/PLA










Liposomes

Composed
varying
ratios of lipid and
cholesterol.






Dendrimers

It is an artificial
polymer molecules
that resembles a
foam ball with tree
like structures.




Most commonly used in
biomedical industry for past 20
years. Biodegradable,
biocompatible and FDA
approved.


Proven results of stimulation of
immune cells as a measure of
increased result in IL2 and INF
-
gamma in spleen homogenate.


Can be efficiently engineered

to act as an controlled delivery
system and hence can
potentially
release the antigens
as well as adjuvant to the
target, at a definitive rate and
thus enhancing the immune
response over a sustained
period of time.


It is a bilayer vesicle which aids
in determining the cellular
interaction between liposomes
and cells.


L
iposomes can deliver the
vaccine by sticking on the cell
surface, slowly diffusing across
the membrane and hence
achieving a controlled release
of the vaccine.





Like proteins they are held by
chemical bond. Contains a
great number of voids or
packets to

carry the
encapsulated materials in
them. They can be engineered
and designed to swell and


Coating process to escape
the scavengers is tedious










Colloidal suspension
stability.









Unfortunately the body
can identify the liposomes
as foreighn intruder.

In

response researchers
have developed stealth
liposomes, coating with
large neutral polymers and
these barriers interact
weakly with antibodies
that attack .








Enhancers for nano carriers:

Surface modification

of nano carrier
:



The
NP

might be

picked

up by macrophages or

phagocytic cells and could result in
undesirab
le reaction and
some inflammatory or toxic reactions

as we have already stated
.
For example granuloma developments were
reported in lung and skin when the animals
were exposed to carbon nanotubes nanoparticles

(21,22)
.
Hence It calls for the attention
to modify the nanoparticles
to make them recognize as
self. The

surface chemistry and
natural features on nano materials
are
important

parameters which has a profo
und effect
in absorption by proteins
,
cell interaction and response with the
host. For example our
claim has been

reported by
Collier To et

al
2009

that

surface chemistry has been
revealed

to
amend

monocyte adhesion in
vitro
.

The
advances in nanocarrier have

yielded
;



Traditional
NP

which had drugs alone.



Targetted
NP

immunocarrier with ligand or ab.



Magnetic
NP

which are fabricated in contrast to MRI, magnetic particles loaded
with therapeutics which are sensitive to ex
ternal magnetic field.



Prolonged surviving and circulating
NP

in which are attached with PEG .



Cell penetrating
NP

with CPP ligand which allows enhanced uptake by cells.










Chitosans






liberate their contents only
when the appropriate trigger
molecules are present.
Dendrimers are found to
bypass the immune defenses
and slip the vaccine to target

area .

Has excellent mucoadhesive
property and biocompatible,
biodegradable.






Agglomerates may form
during synthesis.


Interaction with plasma proteins:


PEG

is a widely studied non fouling molecule and these molecules are found to be highly
resistant to protein and scavenge
rs of the body.
It has been hypothesized that PEG
creates a steric shield around the coated particle in drug delivery system, effectively
p
reventing plasma proteins from adhering to the particle and hence following the same
principle coating our nano based vaccine carrier
would
prevent uptake

by

any

mononuclear
phagocytes.
PEG also

imparts hydrophilic nature which can help the carrier
to shie
ld themselves from the immune system

(23)
.

Opsonisation
: One of the major biological
barriers

to be controlled
for efficient

delivery
of vaccine is opsonisation.
In addition to PEG other materials which are being used are
phospholipids surfaces and sacchar
ide
surfaces
. Materials like PEG, polaxamers and
polaxamines can block the electrostatic as well as hydrophobic interactions of opsonin
with

nano carriers and hence these functional groups can imp
art a stabilized nano
particles [
csaeba et al 2006
].

M
oreira

et al 2001 have demonstrated

that usage of compounds li
ke growth factor
antagonist factors

in

therapeutic
encapsulated liposomes

has
better
kinetics and
biodistribution of nanocarriers in treatment for human small lung
cancer from

carrier
without antagoni
st compound.

They had better uptake and
specifically

recognized

by
H69 cell lines through receptor mediated process leading to
intracellular

accumulation
and efficient release of
therapeutics at target site.


Intergrins and other receptors which are
present on the cell membrane are responsible for the
cell adhesion to absorbed proteins. Hence it is very critical to control the protein adsorption
on the surface of the biomaterials. Several methods are being reported in literature and
examined such as i
ncorporation of short oligopeptides that exhibit that exhibit specific
binding on the surface of biomaterials. Furthermore the structure, confirmation and the
orientation of the oligopeptides used also plays an important role in modulation of
biomaterials
surface.


It is important to protect the vaccines from trafficking lysosomes when they enter in to
cell and protect them from degradation. Now to escape from this degradation process
the researchers have constructed polymers that take the advantage of pH g
radient
between endosome and cytoplasm. Lowman et al have reported new class of polymer,
where the therapeutic are conjugated to carrier at physiological pH. These complexes are
cross linked with acetyl linkers and hence when they enter the endosome these
acid
degradable linker molecules are hydrolysed and thus the therapeutics is released from
the core membrane of the carrier.

A considerable amount of research are carried on the
surface modification of nano carriers such that



They are resistant to biolog
ical intervention in natural environment and



Surface modification would aid better in delivery of vaccines at tumour area.







Conclusion, opportunities and
challenges:


It is evident from various
researches

as discussed in our dissertation, small nanoparticles
can drain freely to LN for delivery of antigens. Subsequent presentation and
elicitation

of
CTL
idealises the Np to be used as
a

suitable
bio nanotechnology

platform to deliver
cancer
vaccine.

The

majo
r research area on the biomaterials were on their chemical
structures but currently there is an increasing focus on other attributes of biomaterials
such as topological, mechanical and electrical attributes, in order to gain proficient
biological response,

that can be further enhanced in
favour

of cancer

vaccine.

Hence for
generating next generation of cancer vaccine using nanotechnology platform, the
researchers must turn to biology for longer term of solution.

Choosing appropriate

nanocarrier is quite amb
iguous, and existing studies seems to be difficult to interpret
because of many biological factors.

Improved targeted NP are been established in
various invivo model and currently more than 120 clinical trials are underway with ab
coated
NP (
37).



Since

nanotechnology is relatively a new field and major biotech companies have started
investing on NP for delivery of biologics
,

recently
,
there is a

major challenge is the
immunotoxicity analysis using NP. The analysis of new NP is not straightforward and
th
ere is a lack of universal guide for toxicity analysis. European standards instructs
application of immune function tests in chronic rodent as an initial test, US FDA
recommends such tests only if there is a pre
-
evidence of such toxicity. Asian country
als
o follows as
-
required
approach (
30
,31,32,33
).
Hence it calls for an immediate action to
validate a scientific approach to identify and

quantify the immunotoxicity related to NP.
In addition to providing warning of risks and setting boundaries, nanotoxicolog
y research
could help to provide potential insight to
design

more efficient nano
platform

for delivery
of cancer
vaccines
.






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Abbreviations:



PEG:

Poly ethylene glycol

NP
:

Nano particles

AB
: Antibody.

Apc: Antigen presenting cells.

DC: Dendritic cells.

LN: Lymph node.

EPR: Enhanced permeation

and retention.