Human Gene Therapy

Jose Bruniard

Copyright 1996


A. The techniques of gene therapy.

I.- Techniques of gene transfer.

Genetic methods of transference: the viral vectors

II. Gene modulators.

III. A case study: Cystic Fibrosis

B. Public Policy Debate.

C. Importance of the topic. Opinion

D. References

A. The techniques of gene therapy.

A genetic disease is caused by a gene that does not work any more because it is defective, as a result of a mutation or deletion. Very few genetic disease can be cured, and only the symptoms are treatable. The mutant genes are not yet altered by the treatment.

The treatment prospect that has most gripped the public imagination is gene therapy. But attempts to treat familial hypercholesterolemia, cystic fibrosis and Duchenne's muscular dystrophy have each resulted in failures over the past year, apparently because patient's cells did not take up enough of the transplanted genes. The earliest treatment of adenosine deaminasa deficiency showed at best a modest effect. In a National Institute of Health (NIH) review, concluded that "clinical efficacy has not been definitively demonstrated at this time in any gene therapy protocol. "

So the great hope for patients with a dominant mutant gene is to replace the mutant gene in the cells of the body with a normal one. In cases where the mutant gene is recessive , the hope is to give patients extra copies of the normal gene.

The main idea being explored at present is to put the DNA coding for a normal gene into a gene of a vector based on a virus. Some viruses show tissue specificity, infecting only lung cells or brain cells, for example. These viruses need to be modified by genetic engineering to destroy their pathogenicity, without affecting their ability to infect their target cells.

The following is a summary of techniques used in gene therapy.

I - Techniques of gene transfer.

1. The gene gun.

This technique consists of bombarding cells with micro-projectiles made of gold or tungsten and coated with DNA. The particles are accelerated through an electric discharge, gas compression or using detonation. The gene gun was used with success in vitro and in situ. The transitory expression of the transferred genes (marker luciferase, beta-gal or CAT) has been observed. This technique is being fine tuned. The efficacy of transfer, measured by the expression, is very low. Only 0.5 to 10% of bombarded cells express the marker.

2. Injection of plasmid DNA in situ.

Circular plasmid DNA is injected into the skeletal muscle of living beings and the protein marker encoded by the DNA is detected in the muscle. The injected DNA is recovered under non integrated forms in the nucleus of cells proximal to the site of injection. This procedure has been repeated systematically and confirmed using different animals after the injection of marker DNA into the skeleton muscle , in rats , mouse, goldfish, cat and monkey. The marker (usually the enzyme luciferase) has been revealed after some weeks. Only negative results were obtained with the non-muscles tissue of the brain, liver, spleen, uterus, stomach, intestine, lung or kidney.

The human gene for muscular dystrophy also is expressed in the mouse. This animal can be a model for myopathy . However, the expression rate of the human gene for muscular dystrophy is fairly low (1% of myofibres) to reestablish the structure and muscular function of these animals. Nevertheless, recovery of function is possible by transfer to the embryo. It is not known if humans myofibres have the same capacity of the myofibres murines of take up DNA.

Because the injection of DNA into mouse muscle is one of the most efficient methods of gene therapy more than 10% of the myofibres express the marker luciferase. For these experiments the herpes retroviral vectors types were ineffective and adenoviral vectors were more effective in mature muscle and in muscle under regeneration. Injection of DNA is a simple method, that may replace the supply of intramuscular protein. In order to produce vaccines more economically, the production of the plasmid, which encodes the protein, has to be also industrialized.

3. Liposomes and polycations.

The liposomes are small, rounded vesicles, formed by a layer of lipids that are capable of transfering nucleic acids to the interior of cells. Its mode of penetration is not yet clearly defined, but it can be said that it is a simple adsorption followed by a fusion of the membrane.

Liposomes were the object of oncology clinical trials. Even though liposomes can be fused with the membrane of a target cell, the DNA can be directed towards the path of lysosome degradations. The first step is to avoid lysosome degradation. The second barrier is the cellular membrane. The transfected DNA should enter the cell correctly and be directed towards the nucleus. At the extracellular level, there are some problems utilizing liposomes. When administered intravenously, they are quickly phagocited by the macrophage. This problem of reliability in vivoexplains why only a few clinical treatments were succesful.

Liposomes were used to deliver DNA into a variety of cell types in a number of clinical trials. Current liposome-targeting developments have centered around the conjugation of liposomes with antibodies or ligand (much in the way that molecular conjugates have been developed). For example, liposomes combined with an antibody against a mouse major histocompatibility antigen delivered DNA more efficiently into targeted cells than liposome-DNA complexes alone. A peptide ligand (transferrin) delivered DNA in combination with liposomes to erythroid cells in bone marrow cells that express the transferrin receptor. Because the DNA must reach the nucleus intact, viral particles, which enhance entry and transit, were fused to liposomes.

Developments in this area might eventually lead to a "super-liposome " containing all of the above characteristics. These would target a cell via the appropriate specific antibody, fuse with the cell membrane and allow entry of DNA via viral protein interaction. Then efficient gene transfer to the nucleus could be achived using nuclear localization signals.

Cellular penetration can be improved by optimizing lipid formulation of the liposomes. The DNA can be complexed over the exterior surface of positively charged liposomes. There are certain poly-cationic complexes that are capable of complexing DNA at its surface. The additional ligand (viral proteins, transferrine, asiaglycoproteines) could ensure a cleavage of the cellular complex.

DNA can be complexed indirectly with a ligand via polycations such as polylysine. Various molecular conjugate vectors have been described, including asialoglycoprotein ligand conjugates to hepatocytes and IgA-ligand conjugates which target respiratory epithelial cells.

DNA coated to an external cationic support is exposed to degradation by nuclease. Another limiting factor to the technique is the immune response directed toward the antagonist component of the complex. This is especially important in cases of repeated treatment. Another major problem associated with liposome vehicles is they are internalized into endosomes. This limits the efficacy of gene transfer. Attempts to circumvent this problem have centered on the inclusion of adenovirus (or portions of adenovirus) in the conjugated vectors because they possess endosomolytic activity.

Genetics methods of transference: the viral vectors

Virus is one of the tools used to transfers genes into human cells. Virus naturally infect cells in vivo and in vitro very efficiently. Because the viral genome sequences such as herpes viruses are known, they can be manipulated. The deletion of a large portion of its genetic material prevents the virus from replicating itself. These deleted sequences can be then replaced by the gene to be transferred. These cells will produce viral particles but they can not replicate.

Current vectors are mainly derived from Adenovirus (Ad), herpes virus (HSV), or retrovirus (mainly from Mo-MuLV, Molony murine leukemia virus). The use of these vectors has security and reliability problems. The following are the advantages and disadvantages of each type.

4. Adenovirus (Ad)

Adenovirus vectors were developed from serotypes of the human adenovirus. They are capable of infecting almost all types of human cellular tissue and carrying large insertions (up to 8 kb). The adenovirals vectors can infect quiescent cells, or cells undergoing slow division. These include muscles endothelial, skeletal, cardiac, and the pulmonary epithelium cells. For adenovirals vectors, DNA is not integrated in the genome of the host cell. This is a positive point because the treatment is innocuous without the risk of insertional mutations. But at the same time is negative point because the DNA persists during the life of the cell. This requires a recurrent treatment for the permanent correction of the genetic defect.

5. Herpes virus (HSV)

Herpes simplex virus of type 1 (HSV-1) naturally infects human nerves cells. The genome of the virus is integrated into the cells of the neuron and is expressed, without replication or modification. It is possible to insert up to 50 kb of DNA. Because Herpes virus infects nerve cells it is a logical candidate for gene therapy of neurological diseases.

6.1. Retroviral Vectors

The development of retroviral vectors that target specific cell types enables novel delivery strategies and possibly reduces the infection of non-target cells. The interaction of the retroviral envelope (encoded by the env gene) with a specific cell surface receptor determines the potential for a retrovirus to infect target cells. Thus, any modification to the env gene might alter the host-range specificity.

Certain viruses, amphotropes, are capable of infecting cells of different animals species. Whereas other types, ecotrophos, are only able to infect the cells of the species that they naturally inhabit. It is possible to obtain retroviral vectors from both types of viruses.

Retroviral vectors are today the most used vectors in human gene therapy. There are several advantages of retroviral vectors. They infect an important cellular types; a amphotrope vector may be tested on diverse animals and humans cells and the efficacy of infection allows the oblation of rare cell such as hematopoietics (1 cell out of 10 000). The hematopoietic cells are very important because they persist throughout the life on the organism. Therefore, the integration of sequences in its genome is a guarantee it will persist.

It is a unique advantage of retroviral vectors that the genetic material is integrated and replicated in the genome of the host cell. One inconvenience is that the integration of the viral genome only occurs during cell division. This limits the therapy to tissues in which cells are dividing. The integration of the viral genome into the cell creates an insertional mutation. Up to now this insertional mutation appears silent, but it is possible that a mutation might alter the regulation of a gene which controls cellular division leading to a cancerous state.

The retroviral size genome is about 10 kb. After deleting viral genes during vector construction, it is impossible to insert up to 6 to 8 kb of exogenous genetic material. The majority of the retroviral vectors now available are based on the Molony murine leukemia virus (Mo-MuLV). gag, pol and env genes are removed and replaced by the gene(s) to be transferred. The sequences necessary for RNA encapsulated and integratation (LTR, Psi and PBS) are conserved. The DNA is transfected into a cell that provides gag, pol and env functions in trans.

6.2. Different types of retroviral vectors.

The retroviral vectors can be classified according to the number of coding sequences (genomic fragment of cDNA, presence or non presence of a selection gene) and the transcriptional signals utilized (Molony or heterologous).

6.2.1. Simple.

The first vectors constructed were simple Molony provirus, in which all the gene replaced by the gene to be transferred.

6.2.2. The retroviral vectors with LTR hybrid.

The promoter of Molony LTR is not active in differentiated cells like the hematopoietics. These cells persist throughout the life on the organism and can be very important to obtain a definitive genetic correction in treated tissue. One approach to overcome this limitation is the recombination of the LTR of the vector. The U3 region of the viral LTR has sequences of amplification (enhancers), which activates the transcription in the differentiated cells . In the cells starting to differentiate or undifferentiated, like the hematopoietic cells or the cell of carcinoma embryonery, these sequences can play a role of repression of the transcription. The most impressive results have come from a retroviral vector containing a muscle creatine kinase (MCK) enhancer element in the U3 region of the viral LTR.

The MCK enhancer is able to exert an active negative effect on transcription from the retroviral LTR, so that it is only in cells induced to undergo muscle differentiation where there is any expression of the transgene. The ability to restrict expression to appropriate circumstances will be a crucial factor in ensuring safety in gene therapy. The second option to improve the vector expression is the insertion of an internal promoter which ensure the transcription of the transgene in the cleavaged cell.

6.2.3. Internal promoter and Self-inactivating,( SIN).

The internal promoters cloned in a vector in the 5' end of the Molony are today heterologous. They can be of viral origin (SV40) or cellular origin (cells of the type TK of the thymidine kinase of HSV-1). They can be combined with an amplifier heterologous sequence and adapted to the transcription in the tissue of interest. The DNA plasmid of the vector transfected in the cells of encapsidation, have one deletion in the LTR DNA. This example presents a sequence of deletion and a sequence of amplification (called AMPLI).

The RNA is transcripted after the internal promoter is extended to the region R in 3' (at the site of polyadenylation AAUAAA). The vectors auto-inactivants are frequently used in genetic therapy because they present the advantage of a minimum risk to active proto-oncogenes, because the amplifiers disappeared after the integration of the vector in the genome of the treated cell .

6.2.4. Double-copy.(DC)

Like the retroviral vector auto-inactivants, the double-copy vectors are recombinations of LTR 3' of the DNA viral plasmid. The modification introduced in region U3 does not grealy affect the useful viral functions. It is a question of the insertion of the transgene, its promoter, and probably of sequences cis-regulated. Since this coding insertion is replicated in the target cells, we call them double copy vectors. The principal advantage is that the "internal" promoter is located outside the transcription viral unit after infection and integration.

II - Gene Modulators.

1. Antisense.

All viral infections, including AIDS, can be described as acquired genetic diseases. Viruses are packages of genetic material that insert themselves into DNA, the double-stranded chain of genes inside the nucleus of every cell, and transform the cell into a factory for producing new copies of the virus. Each sequence on the DNA provides the blueprint for the production of a specific protein. Proteins are complex structures which form the building blocks of all life.

Human DNA contains all the necessary information to produce the proteins that compose our bodies. However, once infected by the virus, the DNA also programs the cell to manufacture the component proteins of HIT. In order to produce proteins, the DNA must transmit its information to a messenger RNA (mRNA) molecule, also known as the sense strand. The mRNA uses this information to organize the building and assembly of proteins into the finished product, usually essential cellular components but, in the case of infected cells, new copies of HIT.

An anti-sense drug is the exact opposite (i.e. the mirror image) of a specific "sense" strand. Antisense drugs attach to mRNA, and thereby block the production of particular proteins at the genetic level. Antisense is an attractive option because each mRNA molecule produces large numbers of each protein. Antisense technology, by attacking a single mRNA strand which is responsible for producing large quantities of single protein, may be a more efficient way of eliminating large quantities of unwanted proteins at once. Since an antisense drug only binds with its exact opposite, it should be extremely specific, producing minimal toxicities.

Since the genes of HIT have been extensively studied by molecular biologists, researchers can design antisense compounds aimed at specific mRNA molecules that produce proteins essential to HIT's survival. Although very few clinical trials have actually begun, and clinical efficacy is a long way from certain, the pharmaceutical industry has devoted significant resources to this burgeoning field. Its proponents believe antisense will radically transform medicine and open up the possibility for new treatments for AIDS and other viral diseases.

2. Triple Helix DNA.

An innovative twist on the antisense idea is the "Triple Helix" technology being developed by several companies. Whereas naturally-occurring DNA consists of two interlocking strands in the famous double helix structure, scientists have developed the means to attach a third strand of DNA to the double helix, effectively blocking transcription. They are now working to synthesize small strands of DNA which are targeted against specific sequences of the HIT genome. In the same way that RNA-targeted therapeutics are more efficient than drugs which bind with proteins, Triple Helix promises to be more efficient than conventional antisense. The compound would bind to the DNA itself, rather that to the thousands of mRNA transcripts. Thus less drug would be needed, greater efficacy would be achieved, and transcription of the unwanted gene would cease completely. However, this technology is at an earlier stage of development, and many biochemical obstacles remain.

3. Ribozymes.

Another promising variation on antisense is a class of compounds called Ribozymes. These are naturally-occurring RNA molecules which function as catalytic molecular scissors, chopping up RNA strands at selected sites. Ribozymes can be synthesized and targeted against specific RNA sequences, just like antisense compounds. However, since ribozymes can effectively destroy many targets, according to its proponents, lower levels of drug might be needed and therapy could be more thorough.

IV - A case study: Cystic Fibrosis.

Cystic fibrosis is a fatal lung disease marked by excessive thick mucus in the airways. The malfunctioning cells, located deep in the lungs, are particularly concentrated in the alveoli, the tiny balloon-like sacs at the end of each air passage. It is due to a faulty gene for transferring salts across cell walls which causes mucous buildup in the tissues, particularly the lungs.

Cystic fibrosis, one of the most common fatal genetic diseases among Caucasians in the United States, afflicts 30,000 Americans. Infections lead to early death, usually by age 30. There is no effective treatment. In general, cystic fibrosis lung disease is a failure of airway defense against bacterial infection. The goal of gene therapy in young children is to prevent the generation of infectious lung diseases, where the realistic goal in cystic fibrosis patients with established lung diseases is to prevent a further loss of lung function.

Dr. James Wilson, Director of the Institute for Human Gene Therapy at the University of Pennsylvania, has initiated a human gene therapy trial for cystic fibrosis. Gene therapy for cystic fibrosis must involve delivering healthy genes directly to defective cells in the lungs. "Because of the location of the target cells, we had to take a more pharmacologic approach to gene delivery and think of the gene as a drug," says Dr. Wilson. He and other researchers have developed methods for delivering therapeutic genes into air passages by using an adenovirus, a tiny virus that causes colds.

The adenovirus is an ideal delivery vehicle because it naturally infects the cells that line the airways. The virus is rendered harmless and a normal copy of the gene that is defective in patients with cystic fibrosis is inserted into it. Solutions containing the modified viruses are instilled into the patients' lungs. So far none of the patients have exhibited complications related to the clinical procedures. "We are now evaluating biological samples for evidence of gene transfer," Dr. Wilson says.

The future of gene therapy depends on the continued availability of key resources and technologies. Especially critical are those resources that aid large animal per-clinical studies and facilities for production of clinical grade recombinant viruses. "The ultimate challenge, however, is to prove the principle of gene therapy in patients, and the GCRC has become a key element in this field," says Dr. Wilson. "It is important to remember that no patient has yet been cured as a result of gene therapy."

In another CF gene therapy study at Stanford University Medical Center, a 20-year-old Santa Rosa woman will be the first patient in a unique study of gene replacement therapy for cystic fibrosis. Healthy cystic fibrosis (CF) genes will be sprayed into one of her maxillary sinuses. The other sinus will receive a different dose a month or two later. The new genetic material should replace faulty CF genes that cause thick mucous buildup that can lead to infections and death. The study is designed specifically to overcome obstacles that have led to failure in other CF gene therapy trials with other vectors. It is the first time genetic replacement has been attempted in the sinuses of CF patients.

Gene therapy for CF has been largely ineffective to date. Earlier attempts at CF gene replacement have been disappointing: not enough healthy genes have been transferred to change the disease, or the patient's immune system has attacked and destroyed cells containing the replacement genes. The Stanford team hopes to succeed by using a different site for the therapy, and a different vector to import the new genes.

The Stanford team chose the maxillary sinuses (below the eye) for this study for safety and convenience: The surface is the same tissue as the lungs, but the site is small and easily accessible. The treatment can be contained, and two different doses can be studied very precisely in one patient. A vector is needed to import the healthy genetic material into the cells.

Prior trials have used an adenovirus, which is one of the causes of the common cold.

The problem was that the viral material caused inflammation in the respiratory tissues before enough genetic material was transferred. The new AAV vector from Targeted Genetics is a parvovirus which alone is not known to cause disease or irritation. It has been disabled so it can't reproduce itself. Since AAV may not activate the immune system, the sinuses may absorb more healthy genes and the genes may remain active longer.

Only 5 to 10 percent of the patient's CF genes need to be replaced, because the cells all connect electrically. One working cell electrically affects many others. Despite the difficulties encountered thus far, the road map for achieving CF gene therapy and testing relevant strategies has become more clear. The observation that CFTR exhibits multiple functions suggests that the most parsimonious route to control this disease remains gene therapy.

B. Public Policy Debate.

1. Introduction

Among bio ethicists, the anti-technological agenda has focused on abuses and social dangers in medical research and practice, and our alleged need to accept death and technological limits. The response of bioethicists and those to the left of the political spectrum to genetic engineering has been particularly fevered, driven by accusations of eugenics and the defilement of sacred boundaries. Rifkin and his Foundation on Economic Trends have led the fight against the release of genetically engineered organisms and the funding of genetics research.

2. Modification of the somatic versus germ-line cells.

Many writers on these technologies draw distinctions between "negative" and "positive"

genetic modification and the modification of the somatic versus germ-line cells. Negative genetic modification has been defined as the correction of a genetic disease, while positive modification has been defined as the attempt to enhance human ability beyond its normal limits. The somatic-germ-line distinction has been made to address the alleged ethical difference in modifying only one's own body, versus modifying one's progeny as well.

As to the modification of one's own genes versus future progeny, the argument is made that current generations would be violating the self-determination of future generations by doing so. The first response is that our choice of breeding partners already "determines" the biology of future generations.

3. Ethical Points for A Defense of gene therapy.

In the case of genetic engineering gene-technologies can, and probably will, give people

longer, healthier lives with more choices and greater happiness. In fact, these technologies offer the possibility that we will be able to experience utilities greater and more intense than those on our current mental pallet. Genetic technology will bring advances in pharmaceuticals and the therapeutic treatment of disease, ameliorating many illnesses and forms of suffering.

Most utilitarians accept the general rule that liberal societies, which allow maximum self-determination, will maximize social utility. Self-determining people should be allowed the privacy to do what they want to with their bodies, except when they are not competent, or their actions will cause great harm to others. Individuals should not be forced to modify their own or their children's genetic code. This has been in the formulation of the Preliminary Draft of a Universal Declaration on the Human Genome and Human Rights [UNESCO International Bioethics Committee, 1995].

Genetic technology does promise to create a more equal society in a very basic way: by

eliminating congenital sources of illness and disability that create the most intractable forms of inequality in society. The general principle is that genetic technology promises to make it possible to give all citizens the physical and cognitive abilities for equal participation.

4- Public perception

Investigations of public acceptance of biotechnology present a disturbing view. Of respondents in one survey, 39% "familiar with the term" said biotechnology may have some benefits but only in limited areas and another 24% felt dangers outweighed any potential benefits. Concern has been voiced that the non-scientifically-trained public receives its information about biotechnology only through the popular press, from books, and from movies like Jurassic Park There is additional concern that such immensely popular but negative depictions could start a backlash against acceptance of biotechnology.

The "framing" of news stories on biotechnology, therefore, has considerable potential for structuring the terms, and therefore the outcome, of public debate on the wisdom and worth of further biotechnological development. News coverage constructs our reality of developments in science just as it does our reality of political events or social trends. While scientific testimony played an important role in each decision-making process, these debates were ultimately resolved in the public arena, in the courts, through congressional action, or by a decision of a public-agency head.

Ironically, despite the efforts on the part of the Asilomar scientists to protect the values of

self-determination, freedom of inquiry, and peer review, the judgements on the outcome of that meeting would eventually be issued by non-scientists. There is a paradox in trying to argue that genetic engineering is novel, different, and exciting, while at the same time insisting that it carries no new risks.

The public's attitude towards biotechnology is probably one of apprehension. The only way to overcome this apprehension is by being open and not giving even the slightest appearance of trying to hide anything. By constructing the public as ignorant when that public may in its own idiom be expressing legitimate concern or dissent, scientific institutions inadvertently encourage yet more public ambivalence or alienation.

The general public may be aware of biotechnology, but if it does not understand its complex scientific issues then it is clearly the responsibility of the scientist to keep the public informed. Ultimately it must be the public that makes decisions about the use of biotechnology and evaluates its social, legal and economic repercussions. The public should be viewed as a "partner" and a level of trust needs to be created. Developing this style will be a major challenge for business leaders as well as university scientists and government regulators.

A 1987 survey of Americans by the U.S. Office of Technology Assessment found that support for genetic engineering ranged from 84% approval for genetic modifications to "Stop children from inheriting a usually fatal genetic disease," to 44% support for "positive" genetic modification to "Improve the intelligence level and physical characteristics that children would inherit."

In a 1993 survey, more than 50% of the respondents in India and Thailand supported the use of gene therapy for the purposes of physical, intellectual or moral enhancement. A 1994 Gallup poll in the UK reports 20% of people accepting enhancement gene therapy.

5- Arguments Against Genetic Technology

There are at least two kinds of criticisms of genetic technology, fundamentalist and non-fundamentalist. Few of these concerns about genetic technology raise new questions for medical ethics. The same questions have been raised by previous medical research and therapy and those challenges have been met without bans on those technologies.

Some non-fundamentalist critics believe that, cumulatively, the risks posed by new genetic

technologies are great enough to warrant postponing genetic research for some indefinite period of study and preparation. With these concerns it will be argued that, with adequate technology assessment and anticipatory regulation, there will be adequate time to regulate genetic technology as we proceed that none of the risks, individually or cumulatively, outweigh the potential benefits.

Medicine Makes People Sick

One extreme position was elaborated by Ivan Illich: medicine itself makes us sick and should be done away with. A variant on this argument is that genetic screening will eventually determine that all of us are "at risk," making everyone see themselves as sick.

More troubling, genetic diagnosis might create a two-tier social system, divided between those with relatively clean genes and those with genetic disease. In other words, genetic diagnosis will make us all genetically diseased. This would be even more problematic if the genetic diagnosis was for a disease which was not yet curable.

Sacred Limits of the Natural Order

Rifkin has joined forces with religious leaders to assert another fundamentalist tenet, that genetic engineering transgresses sacred limits beyond which we should not "play God".

There are no natural limits in our taking control of our biology or ecology. There is no "natural" way to have a baby or die. It may be that this idea of a divinely ordained biological order is distinctly Judeo-Christian-Islamic and not shared by religions and cultures which believe in different cosmogonies.

In a 1993 survey of attitudes towards genetic therapy in the Asia-Pacific region, Macer reports that there was overwhelming support for genetic therapy to cure disease, and that almost no respondents were concerned that the therapy violated the natural order or God's plan.

Technology serve ruling interests

Some argue that the powerful always shape and apply technologies to further their domination of the less powerful. While this is probably true, the conclusion is that all technology should be abandoned.

The Genome is Too Complicated to Engineer

A fundamentalist conviction is that the genome is too complicated to engineer, and therefore there are certain to be unpleasant, unintended consequences. This argument is directly parallel to the deep ecologists' conclusion that human management of the complex global eco-system is impossible, and that our only hope is to leave the planet alone to its own self-organization.

The genome and eco-system are both very complicated and the ability to do more than

correct local defects in either may be many decades away. But eventually we will have the capacity to write genetic code and re-engineer eco-systems and to computer-model the structural consequences of our interventions on future bodies and plants. Our understanding of the genome and ability to predict consequences must be very robust before we allow human applications or the release of animal applications.

While Elias and Annas object to "positive" germ-line therapy, they propose sensible preconditions on the application of gene-engineering: (a) that there should be considerable prior experience with human somatic cell gene therapy, which has clearly established its safety and efficacy; and (b) that there should be reasonable scientific evidence using appropriate animal models that germ-line gene therapy will cure or prevent the disease in question and not cause any harm, and (c) all applications should be approved by the NIH's Working Group on Gene Therapy and local Institutional Review Boards, with prior public discussion.

The formulation of the Preliminary Draft of a Universal Declaration on the Human Genome and Human Rights [UNESCO International Bioethics Committee, 1995] states that: No intervention affecting an individual's genome may be undertaken, whether for scientific, therapeutic or diagnostic purposes, without rigorous and prior assessment of risks and benefits. Undoubtedly, genetic design will undergo extensive experimentation in the design of animals before any human experimentation begins.

The problem with animal research is that it might produce species that are dangerous if released into the eco-system. Release of gene-engineered creatures should be done very cautiously, and it may be that we should have a moratoria on the release of genetically engineered plants and animals until we have adequate oversight. Genengineered micro-organisms are a much greater risk than genengineered humans, since humans don't breed rapidly, are completely vulnerable for years of childhood, are large and visible.

These are many of the same issues raised by drugs and medical devices today. With or without genetically design products we are moving to a new phase of technological assessment of medical products, balancing the demands for demonstrated efficacy and safety with demands for rapid release of useful therapies.

Fascist Applications

Another concern expressed by many critics of genetic technology is the direct consequences of the re-emergence of fascist, racist and authoritarian regimes, and their use of gene engineering to produce compliant, genetically uniform subjects. The first point to make about fascist uses of eugenic ideology or technology is that nothing a democratic society does to forbid itself genetic technology will have any impact on future or contemporary fascist regimes.

The Geneticization of Life

A more diffuse "cultural" concern about genetic technology is that people will begin to see genetics as more central and influential in life than they should. Some people believe that eugenics and genetic determinism are being fueled by contemporary genetic technology and research, at the expense of attempts to ameliorate social ills. Other critics see genetic technology as contributing to the reification of the genetic ties between people, at the expense of valuing their social relationships.

Both of these concerns have some legitimacy. Undoubtedly the public will invest genetics with more importance in the production of disease, intelligence and other characteristics than will be warranted by a more balanced scientific perspective. Our ability to control genetics will help to clarify the appropriate weight to give to genetics in cultural and social affairs. As the nature-nurture relationship becomes clarified, people will not be any less likely. In fact, they will probably, be more likely, to fix the nurture side of their problems.

Genetic Discrimination and Confidentiality

Many opponents of genetic investigation are concerned that increasing genetic knowledge will lead to discrimination against the "genetically diseased and disabled." It is certainly true that employers are already attempting to discover the genetic risks of their employees, and deny employment or health insurance on the basis of this risk profile. A bill guaranteeing the confidentiality of genetic information has been introduced in the U.S. Congress. Regardless of the progress, some form of confidentiality is certain to be guaranteed by the turn of the millennium. In addition, the Americans with Disabilities Act and similar legislation in the U.S. will clearly be mustered to defend workers from genetic discrimination.

The U.S. Human Genome Project's Task Force on Genetic Information and Insurance has recommended that genetic screening be accompanied by universal access to insurance and that genetic screening not be used to deny insurance. Keeping genetic information confidential from insurers and other non-medical personnel in the health care system is tricky, since records will show any special screening or treatment that genetic risks call for.

Unregulated, the use of genetic risk information could greatly strengthen the ability of insurers to exclude the illness-prone from their risk pools, or charge them premiums equivalent to the costs of their potential treatments. Again however, popular insurance reform legislation before the U.S. Congress will ban "risk-rating" and excluding clients with "pre-existing conditions." These two reforms will likely reduce the number of insurance companies in the country by half or more, and make genetic discrimination in health insurance a more or less moot point.

6- Ethical, legal and social implication (ELSI)

In October 1988, James D. Watson announced that a fixed portion of budget of the Human

Genome Project at the National Institutes of Health (NIH) would be set aside for studies on the impact of genetics research on society. That decision has steered $40 million of genome project funds into studies regarding the "ethical, legal, and social implications" (ELS I) of genetics research. The ELSI program now gets a 5% slice of the budget of the National Center for Human Genome Research (NCHGR) at NIH, and 3% of the Office of Health and Environmental Research in the Department of Energy (DOE), NIH's partner in the Human Genome Project.

Leading geneticists, ethicists, and genome program officials contend that ELSI has made important contributions to ethics research and has accomplished the acceptance of a set of principles to guard against genetic discrimination, echoed in a bill protecting workers against loss of insurance that passed Congress. Others maintain that its biggest accomplishment has been to fill the library shelves with bioethics texts or subsidy to the vacuous pronunciamentos of self-styled "ethicists" and that it is a "Welfare program" for ethicists, who only talked but didn't change the world.

Today, NCHGR's ELSI spends more than two-thirds of its budget on extramural grants andcontracts. From its inception in 1990 through 1995 it has funded more than 125 projects, resulting in the publication of more than 150 articles and books. They cover a wide range, including a public television series on genetics, a book by Leroy Hood and Daniel Kevles (The Code of Codes), educational materials, a study of patents and genetics, research on how to educate clinicians, and scores of studies of genetic testing. DOE has used its ELSI money to develop a high school genetics curriculum, fund a model genetic privacy law, and hold seminars on genetics for judges.

The ELSI program shifted its focus in 1992, moving away from a concentration on technical problem such as the genetic discrimination issue, ensuring quality in DNA testing labs, educating doctors in the use of genetic data, and guiding researchers on obtaining informed consent. The existence of genetic data banks would pose "genuine problems" to the individual and ELSI would lead to new guarantees of privacy. It is important to educate the public about their own genetic risks "so that they can make choices." The great ethical failure is having knowledge and not using it. ELSI staff nudged the Equal Employment Opportunities Commission into ruling that a person who tests positive for a disease gene may be viewed under the law as having a disability and therefore be protected against discrimination by employers. (Elliot Marshall, 1996).

7- International Regulation on Gene Therapy

The UNESCO International Bioethics Committee is drafting general guidelines and an international declaration on the human genome and human genetics, which they hope will be approved by the United Nations General Assembly in 1998. The committee considered three major themes: genetic screening, population genetics, and gene therapy. The report on gene therapy has some interesting features but the central theme can be summarized; somatic cell gene therapy is encouraged for any disease whereas germ-line gene therapy, not to be illegal but should not be done.

The conclusions are more liberal than some national guidelines and the Council o f Europe

Bioethics Convention. There is a large debate over whether national or international guidelines are appropriate. UNESCO intends countries to implement more specific national laws, if they wish, in addition to a basic international framework. The call for international approaches is based on several arguments, including shared biological heritage, and the precedents for international law to protect common interests of humanity. Those calling for national guidelines argue that each culture should make their own standards because of national autonomy and because people in each country have different attitudes.

There is support in all countries that have been surveyed in the world for gene therapy,

and genetic screening. Similar results exist for the USA from an Office of Technology survey in 1987 and a March of Dimes survey in 1992. The diversity of views of people in countries around the world is generally similar within each country, which has been called universal bioethics. We need to recognize that people in all countries are mixed in their opinions, this diversity is universal. The types of reasons given are generally similar. This data supports the concept of international guidelines.(Darryl R. J . Macer, 1994.)

In the following table there is a summary of the position of different countries 5 and institutions over gene therapy.
COUNTRY ORGANIZATION Somatic Cell Gene Therapy Germ-live Gene Therapy
USA American Medical Association Yes Yes
USA Catholic Health Association Yes Yes
USA Commission for the Study of Ethical Problems in Medicine and Biomedical and Behavioral Research Yes Not yet
World World Council of Churches Yes No
World World Medical Association Yes
World Council for International Organization of Medical Sciences Yes Not yet
World UNESCO International Bioethics Committee Yes Not be illegal
Europe Council of Europe Parliamentary Assembly Yes No
Europe European Medical Research Councils No
Australia National Health and Medical Research Council Yes No
Canada Medical Research Council Yes Only on animals
France National Ethics Advisory Committee Yes No
G. Britain Committee on the Ethics of Gene Therapy Yes Not yet
Netherlands Health Council Committee on Gene Testing and Gene Therapy Yes Voluntary No
Sweden Ministry of Health and Social Affairs Yes No
Germany Enqute Commission Yes No

This table was summarized from Scope Note 24, Mary Carrington Coutts, National Reference Center for Bioethics Literature.

8- Germline Interventions and Eugenics

Some people believe that any attempt at germline therapy is wrong since it requires imposing the risk of harm on future generations, either by causing unanticipated side-effects in unborn infants or by introducing dangerous genes into the gene pool.

The other major reason for not undertaking germline interventions is that hereditary information which is of value, not for the individual but for the species, may be lost. If lethal or disabling genes are removed from a certain individual's gametes it may be that benefits conferred on the population when these genes recombine with other, non-lethal genes will be lost.

Other argument is about eugenics. There are examples in our own time of governments and private organizations avidly and unashamedly pursuing eugenic goals. The government of Singapore in 1980 instituted a police of providing financial incentives to 'smart' people to have more babies. The California-based Repository for Germinal Choice, has assigned itself the mission of seeking out and storing gametes from men selected for their scientific, athletic or entrepreneurial acumen.

There is no slope that leads inexorably from therapeutic germline interventions intended to benefit future persons to the creation of eugenically-driven, genocidal social policies. Nazi eugenic policies were not aimed at benefitting individuals. Public health not individual therapy was the driving force behind the Nazi medicalization of eugenics.

Some genetic diseases are so miserable and awful that at least some genetic interventions with the germline seem justifiable. It is at best cruel to argue that some people must bear the burden of genetic disease in order to allow benefits to accrue to the group or species. At best, genetic diversity is an argument for creating a gamete bank to preserve diversity. It is hard to see why an unborn child has any obligation to preserve the genetic diversity of the species at the price of grave harm or certain death. The danger inherent in such stances is that they will result in important benefits being delayed or lost for persons who have impairments or diseases that might be amenable to germline engineering.

C- Importance of the topic. Opinion.

The debate about human gene therapy is controversial because it touches many sensitive,

personal, ethical, and philosophical issues for humankind. The principal ethical issue created by modern biology is human embryo research. Here the problem lies in the lack of consensus on a basic human question: When does a fertilized human egg becomes a human being?. For some religious groups life starts from the fertilized egg, so abortion and embryo research is absolutely unacceptable. Others believe that a person is not only the genes, cells or embryo for this matter, but something much more complex, with culture, education, personal belief and feelings which are all much more important than the biological makeup of the body.

The misuse of genetic screening:

Any new technological development might be potentially misused. The problem is not with technology, is with the use we, the human, do with the tools to shape our society.

Our modern world is filled with cases of discrimination based on sex, color or origin, those who are used to discriminate are not waiting for the development of biotechnology, they just do it. We need to improve our society with more democratic and tolerant values, this has to be done at the same time with development of science and technology. A politician can not help very much to solve a DNA sequence, in the same way that science and technology can no t solve our political and social problems.

Insurance companies, like any other company, wants to make as much money as possible. But if they start to refuse insurance based on genetic screening, there is the risk for them to, that other companies, that understand that we are all "defective" in some sense, will be willing to take the risk and the customer from the others who are reluctant. Specific laws and regulation might be necessary, as a framework to solve disputes, but it has a relative importance.

The concern about releasing genetically engineered microbes, plants and animals into the environment has a rational foundation, but we are already releasing to the environment pollution, new non- biodegradable materials, new cultivars developed by conventional means that may carry genes conditioning susceptibility to diseases (i.e. corn with T cytoplasm and the toxin of Helmintosporium maydis race T). There is always risk with new or conventional technologies, but also there is a considerable amount of knowledge in population genetics, plant pathology, epidemiology and ecology that tell us that there is a low probability for something new to develop as a superweed or animal, and at the same time to have the fitness necessary to become prevalent. In the worst case scenario there is always the means to control them.

The modification of genes in the human germ line:

In this case fears are based on historical and current events of eugenics intents. But, once again, there was no gene therapy available when Hitler dreamed about a superhuman. At this point we need to said that it is not yet possible, mainly for technical reasons, to transform human germ cells genetically. But this has not prevented people to imagine scenarios of genes for IQ and make up of people accordingly to father desire.

Even if in the future were possible to use gene therapy for such banal things like change hair or eye color, is not going to be possible to improve IQ, for the simple reason that it is not on the genes, as has been discussed by Lewontin, 1996. The argument that a father can not decide the genetic makeup of their sons, if it is taken seriously, we should ask ourselves, if the the society should prevent the father to educate their sons on the name of self determination, which was the argument of Plato who believed the state should educate all children. The parents transmission of education and ethical values along with fears and misconceptions is by far much more important in the development of a personality than the genetic makeup.

The perspective to ameliorate the suffering of those people with genetic disorders worth by far the risk and fears that people may have today. With continuous education the perception of new technologies by the public will improve.


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