Outline briefly how the antigenic components of current viral vaccines are produced and assess critically the alternatives

Currently available viral vaccines fall into 2 categories;-

1. Whole virus vaccines - live or inactivated, constitute the vast majority of viral vaccines at present

2. Subunit vaccines - made from purified viral antigens

Inactivated whole virus vaccines - these were the easiest preparations to use. The original virus is grown by normal virus culture methods eg. tissue culture (polio Salk), eggs (influenza), mouse brain (rabies Semple vaccine). The virus is then inactivated by formalin or B-propiolactone so that the replicative function is destroyed.

Live whole virus vaccines - live vaccines are prepared from attenuated strains that are almost or completely devoid of pathogenecity but are capable of inducing a protective immune response. They multiply in the human host and provide continuous antigenic stimulation over a period of time. Several methods have been used;-

a. Use of a related virus from another animal - eg. the use of cowpox virus to prevent smallpox.

b. Administration of pathogenic virus by an unnatural route - the virulence of the virus is often reduced when administered by an unnatural route. This principle is used in the immunization of military recruits against adult respiratory distress syndrome using enterically coated live adenovirus types 4, 7 and 21.

c. Attenuation by passage of the virus in an "unnatural host" or host cell and/or the use the temperature sensitive mutants - the major vaccines used in man and animals have all been derived this way. eg. 17D vaccine of yellow fever was developed by passage in mice and then chick embryos, polioviruses were passaged in monkey kidney cells. The molecular basis of attenuation is now beginning to be understood.

Virtually in all areas, live vaccines are superior to inactivated vaccines. A lower dose is required as the vaccine replicates in the human host, and so there is less likelihood of an allergic response. Often, only a single dose is required as opposed to inactivated vaccines where multiple doses are required to achieve immunity. The duration of immunity induced is longer lasting than that of inactivated vaccines. In addition, live vaccines induce good cellular immunity as well as local immunity.

On the debit side, there is a possibility of live vaccines reverting to virulence as in the case of polio Sabin. Also, live vaccines are contraindicated for immunocompromized and pregnant patients.

Whole virus vaccines are not applicable where the virus cannot be cultivated eg. hepatitis B, parvovirus. They are also undesirable in the case of oncogenic viruses eg. EBV, and in case of extremely dangerous infections eg. HIV. Live vaccines are also undesirable for certain viruses which are known to reactivate eg. CMV
 
 

Subunit_vaccines

To date (1989), the only subunit vaccine licensed for use is for hepatitis B which is derived from purified 22nm HBsAg particles from the blood of chronic hepatitis B carriers. There are obvious concerns about the possibility of incomplete inactivation of HBV and other bloodborne agents but to date, there had been no recorded cases of infection transmitted through this vaccine.
 
 

Live vs inactivated vaccines

Feature Live Dead

Dose low high

no. of doses single multiple

need for adjuvant no yes

Duration of immunity many years less

antibody response IgG, IgA IgG

Cell-mediated immunity good poor

Reversion to virulence possible not possible
 
 

Potential safety problems

Live vaccines

1. Underattenuation

2. Mutation leading to reversion to virulence

3. Preparation instability

4. Contaminating viruses in cultured cells

5. Heat lability

6. Should not be given to immunocompromized or pregnant patients

Inactivated vaccines

1. Incomplete inactivation

2. Increased risk of allergic reactions due to large amounts of antigen involved
 
 

Alternative_approaches

Recent advances in molecular biology had provided alternative methods for producing vaccines. Listed below are the possibilities;-

1. Recombinant viral antigen subunit vaccines

2. Synthetic peptides

3. Recombinant whole virus vaccines

4. Anti-idiotype antibodies

Recombinant_viral_antigen_subunit_vaccines

Virus proteins have been expressed in bacteria, yeast, mammalian cells, and viruses. E. Coli cells were first to be used for this purpose but the expressed proteins were not glycosylated, which was a major drawback since many of the immunogenic proteins of viruses such as the envelope glycoproteins, were glycosylated. Nevertheless, in many instances, it was demonstrated that the non-glycosylated protein backbone was just as immunogenic. The obvious advantage of recombinant viral antigen vaccines is that they would be available in unlimited quantities and the production and quality control processes would be simpler than whole virus vaccines.

Synthetic_Peptides

The development of synthetic peptides that might be useful as vaccines depends on the identification of immunogenic sites. The best known example is foot and mouth disease, where protection was achieved by immunizing animals with a linear sequence of 20 aminoacids. Synthetic peptide vaccines would have many advantages. Their antigens are precisely defined and free from unnecessary components which may be associated with side effects. They are stable and relatively cheap to manufacture. Furthermore, less quality assurance is required. Changes due to natural variation of the virus can be readily accommodated, which would be a great advantage for unstable viruses such as influenza. Synthetic peptides are not applicable to all viruses. This approach did not work in the case of polioviruses because the important antigenic sites were made up of 2 or more different viral capsid proteins so that it was in a concise 3-D conformation.

Advantages of defined viral antigens or peptides include:

1. Production and quality control simpler

2. No nucleic acids or other viral or external proteins, therefore less toxic.

3. Safer in cases where viruses are oncogenic or establish a persistent infection

4. Feasible even if virus cannot be cultivated

Disadvantages:

1. May be less immunogenic than conventional inactivated whole-virus vaccines

2. Requires adjuvant

3. Fails to elicit CMI.

Recombinant viral proteins and synthetic peptide antigens are usually less immunogenic than conventional inactivated whole-virus vaccines. This problem may be circumvented to some extent by the use of ISCOMS (immunostimulating complexes), where the antigen is presented in an accessible, multimeric, physically well defined complex. ISCOMS are composed of adjuvant and antigen held in a cage like structure by lipids. Such a multimeric presentation mimics the natural situation of the antigen on the virus.

Recombinant_virus_vaccines

An alternative application of recombinant DNA technology is the production of hybrid virus vaccines. The best known example is vaccinia; the DNA sequence coding for the foreign gene is inserted into the plasmid vector along with a vaccinia virus promoter and vaccinia thymidine kinase sequences. The resultant recombination vector is then introduced into cells infected with vaccinia virus to generate a virus that expresses the foreign gene. The recombinant virus vaccine can then multiply in infected cells and produce the antigens of a wide range of viruses. The genes of several viruses can be inserted, so the potential exists for producing polyvalent live vaccines. HBsAg, rabies, HSV and other viruses have been expressed in vaccinia.

Hybrid virus vaccines have all the advantages of live viral vaccines. They are stable and stimulate both cellular and humoral immunity. They are relatively cheap and simple to produce. Being live vaccines, smaller quantities are required for immunization. As yet, there are no accepted laboratory markers of attenuation or virulence of vaccinia virus for man. Alterations in the genome of vaccinia virus during the selection of recombinant may alter the virulence of the virus. The use of vaccinia also carries the risk of adverse reactions associated with the vaccine and the virus may spread to susceptible contacts. At present, efforts are being made to attenuate vaccinia virus further and the possibility of using other recombinant vectors are being explored, such as attenuated poliovirus and adenovirus.

Anti-idiotype_antibodies

The ability of anti-idiotype antibodies to mimic foreign antigens has led to their development as vaccines to induce immunity against viruses, bacteria and protozoa in experimental animals. Anti-idiotypes have many potential uses as viral vaccines, particularly when the antigen is difficult to grow or hazardous. They have been used to induce immunity against a wide range of viruses, including HBV, rabies, Newcastle disease virus and FeLV, reoviruses and polioviruses.
 
 

DNA Vaccines

Recently, encouraging results were reported for DNA vaccines whereby DNA coding for the foreign antigen is directly injected into the animal so that the foreign antigen is directly produced by the host cells. In theory these vaccines would be extremely safe and devoid of side effects since the foreign antigens would be directly produced by the host animal. In addition, DNA is relatively inexpensive and easier to produce than conventional vaccines and thus this technology may one day increase the availability of vaccines to developing countries. Moreover, the time for development is relatively short which may enable timely immunization against emerging infectious diseases. In addition,  DNA vaccines can theoretically result in more long-term production of an antigenic protein when introduced into a relatively nondividing tissue, such as muscle.

Indeed some observers have already dubbed the new technology the "third revolution" in vaccine development—on par with Pasteur's ground-breaking work with whole organisms and the development of subunit vaccines. The first clinical trials using injections of DNA to stimulate an  immune response against a foreign protein began for HIV in 1995. Four other clinical trials using DNA vaccines against  influenza, herpes simplex virus, T-cell lymphoma, and an additional trial for HIV were started in 1996.

The technique that is being tested in humans involves the direct injection of plasmids—loops of DNA that contain genes for proteins produced by the organism being targeted for immunity. Once injected into the host's muscle tissue, the DNA is taken up by host cells, which then start expressing the foreign protein. The protein serves as an antigen that stimulate an immune responses and protective immunological memory.

Enthusiasm for DNA vaccination in humans is tempered by the fact that delivery of the DNA to cells is still not optimal, particularly in larger animals. Another concern is the possibility, which exists with all gene therapy, that the vaccine's DNA will be integrated into host chromosomes and will turn on oncogenes or turn off tumor suppressor genes. Another potential downside is that extended immunostimulation by the foreign antigen could in theory provoke chronic inflammation or autoantibody production