Definition of DNA Vaccines and Basic

Concept:

 Genes encoding antigen(s) specific to a

particular pathogen are cloned into a plasmid
with an appropriate promoter, and the plasmid
DNA is administered to the vaccine recipient.
 The DNA is taken up by the host cells and the
gene is expressed. The resultant foreign protein
antigens is produced in the cell and then
processed and presented appropriately to the
immune system.
 How Does DNA Vaccines Work:
 DNA vaccines elicit protective immunity against
an infectious agent or pathogen primarily by
activating two branches of the immune sysem: the
humoral arm, which attacks pathogens outside of
cells, and the cellular arm which eliminates cells
that are colonized by an invader. Immunity is
achieved when such activity generates long lasting
memory cells.
 Vaccines induction of immunity begins with
the entry of a DNA vaccine into a targeted cell,
such as muscle and the subsequent production
of the antigens normally found on the
pathogen of interest.
Successful DNA vaccination has been
demonstrated via a number of different routes
including:
 - intravenous
 - intramuscular
 - intra epidermal
 - intra spleenic
 - intra hepatic

with the majority of DNA vaccines so far being

administered through skin or muscle.
 Studies in rodents on the transfection efficiency
of injected DNA have demonstrated that muscle
is 100-1000 times more permissive than other
tissues for the uptake and expression of DNA.
 Tissues are also differ in the efficiency with which
they present antigens to the immune system.
 Tissues such as skin and the mucosal linings of the
respiratory tract and the gut that serve as barriers
against the entry of pathogens have associated
lymphoid tissues that provide high levels of local
immune surveillance.
 These tissues also contain cells that are
specialized for MHC class II restricted
presentation of antigens to helper T-cells. So it is
apparent that:
 muscles rout of administration supports efficient
transfection.
 Intraperitonal and subcotaneous , are the traditional
routs of administration, however they do not
support efficient transfection.

 Skin and muscle tissues, support efficient

transfection but deliver DNA to tissues with immune
surveillance.
 Methods of Administration

 Plasmid delivery at these sites is usually

accomplished by one of two methods:

 1- needle injection of DNA suspended in saline

 2- Gene gun, this method has more commonly used for
epidermal rather than intramascular administration.
 Several researchers have reported that the gene
gun mediated immunization is far more efficient
than needle injection, eliciting similar levels of
antibody and cellular responses with 100-5000 fold
less DNA.

 It was reported that as little as 16 ng of plasmid

DNA delivered epidermally via gene gun could
induce antibody and CTL responses in mice,
wherase intradermal injection of the same plasmid
requires 10-1000 µg of DNA to elicit comparable
responses.
 With regard to the immunization regimens, there
has not been any regimen that is shown to be
superior to others, it seems that each disease and
each vaccine construct differs from the other,
therefore, the best regimen of DNA vaccine
administration yet to be determined.
Enhancement of DNA vaccines action

 The most promising method of vaccine

enhancement is the co-administration of plasmid
encoding cytokines along with a plasmid encoding
an antigen.
IL-2 : a potent stimulator of cellular immunity that
induces proliferation and differentiation of T cells
as well as B cell and NK cell growth.

 It is reported a five fold increase in antibody

response when IL-2 plasmid was co- injected
with the plasmid encoding the antigen.
 IL-4:

induces differentiation of T-helper cells into Th2

subtype, enhances B cell growth, and mediates
Ig class switching. It was reported that injection
of a plasmid encoding IL-4 3 days before
immunization with a protein antigen increased
Ag specific antibody levels compared to protein
immunization alone.
 Granulocyte-monocyte colony-stimulating
factor (GM-CSF):

This cytokine increases production of granulocytes

and macrophages and induces maturation and
activation of APCs such as dendritic cells.
 Advantages and properties of DNA
vaccines
 Plasmid vectors can be constructed and tested
rapidly.
 Rapid and large-scale manufacturing procedures
are available.
 DNA is more temperature stable than live
preparations.
 Microgram quantities of expression vector can
induce immune response.
 Unlike killed vaccines, DNA vaccines can
produce diverse and persistent immune response
(both humoral and cellular arms of the immune
response)
 Protection can be achieved in large primate
models of human infections
 Multiple vectors encoding several antigens can
be delivered in a single administration
 Unlike the live attenuated vaccines, who posses
the risk of reversion to pathogenic state while
replicating inside the host, DNA vaccines are
safe and do not encode for genes that cause
diseases

 Unlike the killed vaccines, who induce short

immunity and need frequent boosting, DNA
vaccines cause long lasting immunity with
minimum boosts
HOW DNA VACCINE IS MADE

Viral gene
Recombinant DNA
Technology Expression
plasmid

Plasmid with foreign gene

 Transform into
bacterial cell

Plasmid
DNA
Bacterial cell
Plasmid DNA get
Amplified
Plasmid DNA
Purified

Ready to use
 METHODS OF DELIVERY
 Syringe delivery:-

Either
intramuscularly
or
Intradermally
 Contd..
 Gene gun delivery:-

Adsorbed plasmid DNA

into gold particles
Ballastically accelerated
into body with gene gun.
 HOW DNA VACCINE WORKS

BY TWO PATHWAYS
ENDOGENOUS :- Antigenic Protein is presented by
cell in which it is produced
EXOGENOUS :- Antigenic Protein is formed in
one cell but presented by
different cell
 HOW DNA VACCINES WORK

Muscle Cells
+ Plasmid DNA
 ENDOGENOUS PATHWAY

Nucleus

Plasmid
DNA

mRNA MHC-I

Antigenic
Peptides

Antigenic
Protein
 T- Helper Cell

Mu
ltip
ly

Memory T cells
EXOGENOUS PATHWAY

Antigenic Protein come outside

 s e d
oc yto
h ag
P

Antigen Presenting Cell

Antigenic Peptides
Memory
T- Helper Cell
Antibodies
Cytokines
Plasma B-Cell

MHC-II
Activated B-Cell Memory B-Cell
 ADVANTAGES

Elicit both Humoral & cell mediated

immunity
Focused on Antigen of interest
Long term immunity
Refrigeration is not required
Stable for storage
 DISADVANTAGES

Limited to protein immunogen only

Extended immunostimulation leads
to chronic inflammation
Some antigen require processing
which sometime does not occur
 Genetic Toxicity

Integration of DNA vaccine into host Genome

Insertional mutagenesis
Chromosome instability
Turn ON Oncogenes
Turn OFF Tumor suppressor genes
Over Expression of DNA vaccine

Acute or chronic inflammatory responses

Destruction of normal tisues

Generation of Autoimmune
diseases

Anti DNA Antibodies

Autoimmune diseases

Autoimmune Myositis
 Antibiotic Resistance

Plasmid used is resistance to

antibiotics for selection

Raise the resistance to same

antibiotic in the host
 FUTURE PROSPECTS

 Plasmid with multiple genes provide immunity

against many diseases in one booster

 DNA vaccines against infectious diseases such

as AIDS, Rabies, Malaria can be available
Thank you