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RECOMBINANT VACCINE

INTRODUCTION Definition  Vaccine

may lived or weakened strains of viruses or bacteria which intentionally give rise to in apparent to trivial infections  Vaccines may also be killed or inactivated organisms or purified product derived from them

INTRODUCTION  Vaccines

involves deliberate exposure to antigen under condition where disease should not result.

 It  It

is used to induced protective immunity

should induce antibody formation in the body

TYPES OF VACCINES 

A. Traditional vaccines (i) inactive (killed) vaccine (ii) attenuated vaccine (iii) Subunit vaccine

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B. New generation vaccines (i) conjugate vaccine (ii) Recombinant vaccine (iii) DNA vaccine

INACTIVE VACCINES  These

are the type of vaccines which are prepared by micro organisms which are previously virulent that have been killed (inactivated) with heat or chemicals (formaldehyde) so that it can not produced disease in person who receives the vaccines Eg. Original Salk vaccine

ATTENUATED VACCINES  These

are lives micro-organs that have been cultivated under condition which disable their virulent properties  They typically provoke more durable immunological response & are preferred type for healthy adults  E.g.. Yellow fever ,measles, mumps

SUBUNIT VACCINE  Rather

than introducing a whole inactivated or attenuated micro-organism to an immune system, a fragment of it can create an immune response  e.g. is the subunit vaccines against HBV that is composed of only the surface proteins of the virus

CONJUGATE VACCINE  Conjugate

vaccines have been developed to pathogens whose polysaccharide capsules protect them from phagocytosis eg. S. pneumoniae, and Neisseria meningitidis  This polysaccharides outer coats is poorly immunogenic, by linking these outer coats to proteins ,the immune system can led to recognize the as if it were a proteins antigen

RECOMBINANT VACCINE  Recombinant

vaccines are those in which genes for desired antigens are inserted into a vector, usually a virus, that has a very low virulence property.

 The

vector expressing the antigen may be used as the vaccine, or the antigen may be purified and injected as a subunit vaccine

DNA VACCINE 

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DNA vaccines are the newest vaccines and are still experimental ,although to date they seem to be very effective and their safely record is good., DNA of pathogen is inserted in animal cell or human cell. some muscles cells express the pathogen DNA to stimulate the immune system Both humeral and cellular immunity have been induced by DNA vaccines.

WHAT IS RECOMBINANT TEC.  This

technique was develop in 1980  This is technique by which desired gene can be cloned, DNA can be re-sequenced, & recombinant proteins can be produce  variety of important molecules for life can be prepared by introducing desire gene in DNA.  This technology use for production of hormones

HOW DOSE IT WORK?  Each

cell of living organism contains DNA, which contains genetic information  This DNA(genes) is necessary for the product of a verity of essential molecules in the body e.g.. Proteins , fats,  It was discovered that gene could be transferred from one cell to another

 This

mean that cell could be reprogrammed to secrete a particular protein for eg.growth hormone by introducing gene coding for growth hormone

 These

modified cell – now call as recombinant cell- transmit new characteristics in daughter cell

Transfer and Cloning of the Insulin Gene

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Recombinant vaccine A recombinant vaccine contains either a protein or a gene encoding a protein of a pathogen origin that is immunogenic and critical to the pathogen function;  the vaccine is produced using recombinant DNA technology.  The vaccines based on recombinant proteins are also called subunit vaccines. 

 Proteins

are generally immunogenic, and many of them are critical for the pathogenic organism.

 The

genes encoding such proteins can be identified and isolated from a pathogen and expressed in E. coli or some other suitable host for a mass production of the proteins

Transferring a Gene from one cell to another

TYPES OF RECOMBINANT VACCINE

 Subunit

vaccine  Gene deleted vaccine  Vectored virus vaccine  Other

Subunit vaccine 

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A subunit vaccine contains only the proteins which stimulate the immune system to attack. By manipulating DNA that codes for these most stimulatory proteins, we can mass produce a purified solution and immunize with only the antigen we want and no extraneous antigens. Eg. The Genitival brand of Feline Leukemia Vaccine (not currently on the market) is an example of a subunit vaccine

Gene deleted vaccine 

Gene which is responsible for production of antigenic response is deleted or removed from pathogen ,

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Pathogen is then inserted in to the body

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Due to removal of antigenic gene could not able to produce infection & physical illness

Vectored virus vaccine 

The DNA for the stimulatory proteins are inserted into harmless viruses. The live harmless virus is able to provide a very natural immune stimulation and will express the stimulatory proteins native to a harmful virus. In this method, one gets the benefits of the modified live vaccine with the benefits of the subunit vaccine. Eg. Rabies vaccine

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This technology is currently in use to vaccinate wildlife against rabies The new PUREVAX rabies vaccine for cats is also a vectored virus vaccine.

Other vaccine 

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the "naked DNA or RNA" vaccine. Here, DNA from the infectious agent is injected into the host. No proteins. No putting the DNA inside any viruses. Just plain raw DNA. This DNA is taken up by the local muscle cells and soon the proteins coded for by this DNA are being produced. The immune system thus receives its stimulation without risking exposure to an infectious agent of any kind.

METHOD of PREPRATION 

The first step is to identify a protein that is both immunogenic and critical for the pathogen.

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The gene encoding this protein is then identified and isolated.

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The gene is integrated into a suitable expression vector and introduced into a suitable host where it expresses the protein in large quantities.

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The protein is then isolated and purified from the host cells

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It is used for the preparation of vaccine.

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The host organisms used for expression of immunogenic proteins to be used as vaccines may be anyone of the following.

1. A genetically engineered microorganism, e.g., yeast for the expression of hepatitis B surface antigen (HBsAg) used as vaccine against hepatitis B virus (approved for marketing in India).

2. Cultured animal cells, e.g., HBsAg expressed in CHO (Chinese hamster ovary) cell line and C-127 cell line. 3. Transgenic plants, e.g., HBsAg, HIV-l (human immunodeficiency virus-I) epitope (in experimental stages). 4. Insect larvae; the gene is integrated into a bacculovirus genome, which is used to infect insect larvae. Often a very high quantity of the recombinant protein is produced.

Recombinant vector vaccine 

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Genes encoding for major antigens of especially virulent pathogens can be introduced into attenuated viruses or bacteria. The attenuated organism serves as a vector, which replicates within the host and expresses the gene product of the pathogen. Various organisms have been utilized for vector vaccines and include adenoviruses, attenuated poliovirus, vaccinia virus, BCG strain of Mycobacterium bovis and attenuated strains of Saimonel/a.

 vaccine

virus, which has been used to eradicate smallpox is widely employed as a vector vaccine. This large and complex virus containing a genome of about 200 genes, can be manipulated to carry scores of foreign genes without any impairment in its capacity to infect host cells and replicates.

HBV recombinant vaccine 

DescriptionHepatitis B Vaccine (Recombinant) is a noninfectious subunit viral vaccine derived from hepatitis B surface antigen (HBsAg) produced in yeast cells. A portion of the hepatitis B virus gene, coding for HBsAg, is cloned into yeast, and the vaccine for hepatitis B is produced from cultures of this recombinant yeast strain according to methods developed in the Merck Research Laboratories.

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Vaccine must contain antibody against surface antigen(HBV Ag) It was first made from the serum of people who had been HBV infected and subsequently cleared the infection. Because these individuals naturally produced anti-HBsAg IgG and then cleared the infection, they were immune to reinfection with HBV. The serum from these individuals was taken and purified antibody was passively administered to others in order to induce immunity

Recombinant tec.  In

an infected individual HBsAg is present in two forms:  as a protein on the surface of the 42 nm virion (also called the Dane particle)  and as a secreted 22 nm particle that is a hollow sphere of surface antigen.

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gene encoding for this proteins is determine and is inserted in to the vector Two types of vector is used 1.

Yeast : sacchromyces cervices

3.

Chinese hamster overy

Yeast cell  yeast

assembled this 22 nm protein(s gene) in the same way that surface antigen assembles and is secreted in humans.  Therefore, the artificial surface antigen resembled the naturally occuring particle.

Chinese hamster ovary (CHO)  the

recombinant plasmid was transfected into Chinese hamster ovary (CHO) cells.  this plasmid was different than the recombinant yeast plasmid because it included an extra piece of the genome While the yeast plasmid included only the S gene of HBV, the CHO clones used the S gene plus the pre-S2 portion of the genome,

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creating a vector that began one start codon upstream in the genome.

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this case as well, the 22-nm particle containing the immunogenic HBsAg was produced by the artificial system as that of yeast .

 Data from one of many efficacy trials... image taken from Da Villa et al. Res Virol. 1998. 9.

Production of vaccinia vector vaccine 

The gene that encodes the desired antigen (organ) is inserted into a plasmid vector adjacent to a vaccine promoter

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When this vector is incubated in tissue culture , the antigen & promoter inserted in to the vaccinia virous genome