While relatively simple in concept, the goal of putting genes into patientsí cells has been elusive due to a variety of technical problems which must be overcome before gene therapy can be used. These technical challenges include developing the ability to efficiently transfer the DNA which codes carries the therapeutic gene into target cells of patients and, once the gene is in place, to get it to express (make the protein it codes for) at high enough levels to be of therapeutic benefit. Over the past several years significant progress has been made in overcoming these and other technical hurdles to successful gene therapy. However, the very discoveries which will make gene therapy a viable strategy in the near future, may also be applied to the development of novel biological weapons or the "upgrading" of current weapons so that they are able to circumvent current defensive strategies. Conversely, gene therapy strategies may also be applied to protect targets from specific bioweapons. Specific examples of such strategies are presented below.

A paradigm for biological weapons is the use of pathogenic viruses or bacteria to infect targets. Potential defenses against such agents include antibiotics or vaccines to suppress the development of infections by these agents or the use of immunologic or pharmacologic agents to suppress the effects of toxins that might be produced by the pathogens.

1. Use of drug resistance genes.

A strategy used in the gene therapy of cancer is to transfer genes which confer resistance to certain toxic drugs (i.e., chemotherapeutic agents) to the normal cells of a patient. For example, if the dose of a certain chemotherapeutic agent is limited by its toxicity to blood cells, then a gene which protects cells from the agent could be put into all blood cells. Therefore the blood cells would now be resistant to the chemotherapy drugs so that higher doses could be used. The higher doses might then allow for more effective killing of cancer cells. Examples of proteins which protect cells from chemotherapy drugs include enzymes which break down the drug inside cells, pumps which are able to pump the3 drugs out of cells and proteins which allow the cells to keep growing despite the damaging effects of the drug. Technology currently available for gene therapy and molecular biology could easily be adapted to transfer protective genes to pathogenic bioweapons such as bacteria and viruses, thus making them, or the cells they infect, resistant to drugs which might combat the warfare agents.

2. Alteration of toxin genes to potentiate biologic damage.

Another strategy in gene therapy is to replace genes which code for abnormal proteins with genes which code for proteins with normal or even improved functional properties. Genes coding for toxins of pathogenic microbes could be isolated and engineered ex vivo to produce proteins with altered properties. One example might include toxins which bind more strongly to a cellular target and thus produce a more potent response. Another might involve a toxin for which a specific pharmacologic inhibitor had been designed. The gene, and therefor the protein structure, of the toxin could be altered so that it was now resistant to the antidote but was still able to carry out its toxic function. Using gene therapy-derived gene transfer techniques these genes could then be returned to the parent microorganisms making them more effective biological weapons.

3. Alteration of genes to help microorganisms elude vaccine strategies.

One current defensive strategy against infectious bioweapons is to vaccinate potential targets so that an immune response is developed to the potential agents. These strategies result in the production of antibodies which can bind to and inhibit the function of biotoxins or kill microorganisms. Similarly, vaccines can also lead to the development of specific immune system cells (lymphocytes) which destroy invading microorganisms. Vaccines to specific organisms or the toxins they produce can potentially be administered to persons to elicit immune responses to these agents. The antibodies and lymphocytes which mediate these responses specifically recognize structural features of the microbe or toxin and destroy it. Using molecular biological techniques the genes for these immunologic targets can be isolated, modified so that they are no longer recognized by the targetís immune system and returned to the parental microbe to produce essentially a new strain which will not be recognized by the immune defenses of a vaccinated target.

4. Transfer of toxic gene products from one infectious bioweapon to an alternative agent.

A specific antibiotic, vaccine or other strategy might be developed against an infectious, toxin-producing bioweapon making that weapon ineffective. Using methods adapted from gene therapy the gene coding for the toxin could be identified, isolated and inserted into a new microorganism (for example a different bacteria or a virus) thus delivering the same toxin with a different vector.

5. Transfer of a non-microbial toxin gene into a microbe.

The gene for a non-microbial protein toxin (such as a snake, fish or spider venom) could be inserted into the genome of an infectious agent (such as a bacteria or a virus) so that the toxin would be produced within the targetís cells. Multiple toxin genes could also be inserted into the same vector to increase toxic potential.

6. Changing the tropism of an infectious bioweapon.

Many infectious agents infect specific cells within the human body by binding to proteins on the surface of the target cells. This binding to specific target cells is mediated by specific proteins of the surfaces of the viruses or bacteria. By exchanging the genes which code for these microbial proteins, the normal target tissues of the weapon could be changed so that a new organ can be targeted. For example a virus that normally infects the liver and needs to enter a persons blood stream to be effective could be altered to target lung tissue so that it could be administered by inhalation.

7. Development of novel infectious agents.

In order to create more effective viruses to transfer therapeutic genes, new versions of viruses have been developed from which many or most of the viral genes have been removed and then replaced by the gene or genes to be carried. While in many cases this has been done to remove genes coding for virulent proteins, similar manipulations could be performed to enhance the virulence of a virus. For example, genes coding for multiple toxins could be inserted into viruses. Another example is that a disease causing virus such as the AIDS virus could be made more virulent by the addition of a toxin or by changing the viral surface proteins so that the virus is resistant to vaccines.

8. Transfer of genes without microorganisms.

Because of potential hazards and inefficiencies involved with the use of microorganisms as vectors to transfer genes in gene therapy several strategies have been developed in which DNA genes can be put into a patientís cells directly. These technologies include the injection of gold particles coated with DNA into a personís skin, direct injection or inhalation of naked DNA or DNA complexed to lipids. While these strategies are not likely to be applicable to large scale bioweapons, they might be effective as local weapons. One could envision the transfer of a toxin producing gene into a target or even the introduction of a gene that might cause cancer in a target several months or years after the attack.

9. Regulated expression of toxic genes.

In certain gene therapy applications it is advantageous to be able to turn genes which have been delivered to a patient on or off a specified times specific genes on or off by the administration of a drug. Such systems have already been developed and are being employed in models of gene therapy. These could be used as part of a controlled or clandestine bioweapons strategy where targets could be infected with a virus (for example) carrying a toxic gene. The gene would lie dormant inside the targetís cells until the signal, such as an common antibiotic tetracycline was ingested. This would then activate the gene and produce a lethal response.

Due to the high degree of plasticity of biologically-based weapons, the use of gene transfer concepts seems more applicable to offensive strategies. They may, however also be used defensively. One example is given below.

1. Vaccine Development

Vaccines are among the potentially most effective protective mechanisms against microbiologic microorganisms and toxins. However, concerns over the safety of vaccines based on the actual organisms remains. One way around this would be to use vaccines based on the genes or proteins of the microorganisms. Gene therapy experiments have demonstrated the feasibility of vaccinating people with very small quantities of DNA which code for microorganism proteins. The DNA is taken up by the personís cells, the protein coded for by the gene is made and the body then develops a potent immune response to the foreign protein thus offering protection against the intact microorganism even though only a small piece of DNA has been administered to the person. Similarly, small pieces of protein derived from a microorganism can be administered to elicit an immune response. These DNA fragments can be produced efficiently and inexpensively and administered safely to subjects.