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Syeda Hira 08-arid-1152Ph.D (Biochemistry)

DNA VACCINES

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Contents

• Introduction

• History

• Difference

• Production of DNA vaccines

• Method of Delivery

• Mode of Action

• Advantages and Disadvantages

• Future Perspective

• Conclusion

Introduction

Small circular piece of bacterial DNA.

Genetically Engineered to produce one or two specific protein

from microorganism.

The DNA Sequence code for antigenic protein of pathogen.

As this DNA inserted into cells it is translated to form antigenic

protein. As this protein is foreign to cells , so immune response

raised against this protein.

In this way ,DNA vaccine provide immunity against that pathogen.

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History In 1990, University of

Wisconsin, Jon Wolff found that

injection of DNA plasmids

produce a protein response in

mice

Since then concept of DNA

vaccines started

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Difference between DNA vaccine and Traditional vaccine

Traditional medicine Uses weak or killed form of an infectious agent Provide humoral immunity

DNA vaccines Uses only the DNA from the infectious organism Provide both humoral and cell mediated immunity

Preparation of DNA Vaccine

Viral gene

Expression plasmid

Plasmid with foreign gene

Recombinant DNA Technology

Bacterial cell

Transform into bacterial cell

Plasmid DNA

Plasmid DNA Purified

vaccine

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DNA Vaccines: Optimization strategies

1. Plasmid optimization

2. Gene optimization

3. Formulation adjuvant

4. Immune plasmid adjuvant

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Plasmid Optimization

Plasmid vectors for use in vaccination contain

• Promoter

• Enhancer

• Antigen-encoding

• Polyadenylation sequences

• Antibiotic resistance site

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Gene Optimization

• Selection Requirements

High GC content

Species-specific codon utilization

Consensus immunogens

Nuclear localization sequences

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Formulation Adjuvants

Microsphere/nanoparticle

Liposomes

Polymers

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Immune Plasmid Adjuvants

Cytokine

Chemokine

Co stimulatory molecules

Heat shock proteins

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Methods of Delivery

There are two methods

Intramuscular immunizations both directly transfected dendritic cells and

macrophages can present antigen

i.m. immunizations are largely independent of DNA expression in

the muscle target

intramuscular deliveries of DNA tend to raise type 1 T-cell help for

intracellular and plasma membrane antigens but type 2 T-cell help

for secreted antigens.

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Continue… Gene Gun Method

• Covering gold micro particles with recombinant DNA• Shoot them by gas pressure normally helium• gene gun immunizations directly transfected dendritic

cells present antigens• Gene gun immunizations depend on antigen expression

at the skin target • Gene gun immunizations tend to raise type 2 T-cell help

for both cell-associated and secreted antigens. 

The “gene gun”

The Helios Gene Gun is a new way for in vivo transformation of cells or organisms (i.e. gene therapy and genetic immunization (DNA vaccination)). This gun uses Biolistic ® particle bombardment where DNA- or RNA-coated gold particles are loaded into the gun and you pull the trigger. A low pressure helium pulse delivers the coated gold particles into virtually any target cell or tissue. The particles carry the DNA so that you do not have to remove cells from tissue in order to transform the cells.

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Mechanism of Action

The DNA vaccine works in 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.

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mRNA

Antigenic Protein

Antigenic Peptides

MHC-I

Plasmid DNA

Nucleus

ENDOGENOUS PATHWAY

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Multiply

Memory T cells

T- Helper Cell

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EXOGENOUS PATHWAY

Antigenic Protein come outside

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Phagocytosed

Antigen Presenting Cell

Antigenic Peptides

T- Helper Cell

Cytokines

Activated B-Cell Memory B-Cell

Plasma B-Cell

Memory Antibodies

MHC-II

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WHEN VIRUS ENTER IN THE BODY

Viral Protein

Memory T-Cell

Antibodies

Advantages

Elicit both humoral and cell mediated immunity Plasmid vectors can be constructed and produced

quickly  and the coding sequence can be manipulated in many ways.

Focused on antigen of interest Stable for storage Not require refrigeration Another important advantage of genetic vaccines is their

therapeutic potential for ongoing chronic viral infections

Disadvantages

Prolong immunostimulation may lead to chronic inflammation

Limited to protein Antigen only.DNA can be used to raise immune responses against pathogenic proteins, certain microbes have outer capsids that are made up of polysaccharides.  This limits the extent of the usage of DNA vaccines because they cannot substitute for polysaccharide-based subunit vaccines

Safety Issues

Genetic toxicity Integration of DNA vaccine into host Genome may result inChromosome instability, turn ON Oncogenes ,Turn OFF

tumor Suppressor Genes

Over expression of DNA vaccine may cause acute or chronic inflammation and normal tissues destruction

Generation of autoimmune diseases

Resistance to antibiotics

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

Conclusion

Simple ,safe and cheap alternative method for generation of humoral and cell mediated immunity

DNA Vaccines are in their early phase

DNA vaccines are going to be Vaccines of next generation

References• Widera, G., M. Austin, D. Rabussay, C. Goldbeck, S. W. Barnett, M. Chen, L. Leung,G. R. Otten, K. Thudium, M. J. Selby, and J. B. Ulmer. 2000. Increased DNA vaccine delivery and immunogenicity by electroporation in vivo. J. Immunol. 164:4635– 4640.• John J. Donnelly,* Britta Wahren, and Margaret A. Liu.2005. DNA Vaccines: Progress and Challenges. J. Immunol. ; 175: 633-639.•Donnelly, J. J., J. B. Ulmer, J. W. Shiver, and M. A. Liu. 1997. DNA vaccines. Annu. Rev. Immunol. 15: 617– 648.•Tang, D.; Devit, M.; Johnston, S.A.; Others. 1992. Genetic immunization is a simple method for eliciting an immune response. Nature 356 (6365): 152–154. •Michele A. Kutzler* and David B. Weiner. 2008. DNA vaccines: ready for prime time? Nature Reviews. 9: 777-788.•M. A. LIU. DNA vaccines. 2003. J. Internal Medicine. 253: 402–410.•Siddhesh D. Patil,1 David G. Rhodes,1 and Diane J. Burgess. 2005. DNA-based Therapeutics and DNA Delivery Systems: A Comprehensive Review. AAPS Journal. 7:61-77.

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