Molecular Therapy

Realistic forms of gene therapy that are likely to be applied in the near future are those that affect diseases of aging rather than aging processes. Some of the potential future therapies are listed in Table 1, but this is not a comprehensive list. Because these therapies have the aim of restoring normal health by correcting an abnormal condition, and do not involve germ-line manipulations, they do not have the ethical problems associated with attempting to change maximal life span. Some of these therapies are close to being used in human subjects, having been successfully demonstrated in experimental animals. The idea of using gene therapy is especially attractive for chronic diseases in which current forms of medication must be administered frequently, often several times per day; compliance with such regimens is a problem, especially for older people. There is an obvious advantage to replacing drug therapy with a form of molecular therapy if the effects are equivalent. Moreover, for many diseases of older individuals, present treatments are inadequate or nonexistent, making the development of new therapies very desirable.

Since the beginning of gene therapy as an idea, different delivery methods have been developed in parallel, without any one of them eclipsing the others, and the existence of multiple strategies for gene delivery is likely to persist in the foreseeable future. Different genes may require different delivery methods for optimal effects. Several recombinant (genetically engineered) viruses can be used as vectors (i.e., carriers of genes). Adenovirus-mediated gene Table 1 Proposed gene and cell therapies applicable to age-related diseases. SOURCE: Author delivery has been used successfully in young individuals, and has the advantage of very efficient delivery to nondividing cells, particularly the liver. However, earlier generations of adenovirus vectors were highly immunogenic, leading to dangerous patient reactions. These problems appear to have been solved in later generations of these vectors, but all adenoviruses exert a therapeutic effect only over a short time. Longer-term effects have been achieved with vectors based on the adeno-associated virus (AAV) and the herpes simplex virus, and with lentivirus vectors based on components of HIV (human immunodeficiency virus). These vectors can infect nondividing cells, and their genetic material becomes stably integrated into the infected cell's DNA. Vectors based on other retroviruses, such as mouse leukemia virus, can be used only to infect dividing cells, and so have not been particularly successful in in vivo applications, but they can be very useful in ex vivo cell modification. Gene delivery can also be accomplished without the use of viruses. Nonviral DNA delivery suffered until recently from a lack of efficiency, making it impractical, but trials of newer versions of liposomes have been very promising. Nonviral vectors could be just as efficient as viruses and could obviate the problems associated with viruses—not only side effects but also public acceptance of the therapy, especially for the treatment of diseases that are not immediately life-threatening.

An inherent problem with the use of viral and nonviral vectors is the difficulty of ensuring that the gene is delivered to the appropriate number of the patient's cells. The number of cells infected, and hence the amount of product delivered, is hard to control. A second major problem is maintaining long-term gene product delivery, so as to avoid the necessity for repeated administration of the vector. Another concern is that because gene therapy either intentionally modifies cells of the body (introduces genes into the host genome) or has the potential for accidental permanent genetic modification, expression of endogenous genes in the modified cells could be altered, potentially causing the cells to become cancerous. A concern that is of more importance in young patients is that germ-line cells will be unintentionally modified. More generally, it may be undesirable to create the potential for continued production of the gene product in cases where only temporary delivery is required. These concerns are less important in critically ill patients, of course, but must be satisfactorily addressed before these forms of gene therapy are widely adopted.

Cell-based delivery, or ex vivo gene therapy, uses genetically modified cells as the gene product delivery system. In this method, possible unintended alterations in gene expression can be tested before the cells are used. There is no potential for accidental germ-line modification. However, two disadvantages to cell-based therapy are that introduction of the cells into the patient requires a surgical procedure, and that cells must be protected from immune rejection. The cell transplantation procedure may require only minor surgery, however, and in some sites in the body the cells could be removed after their task is completed. In other sites into which they might be transplanted, such as the brain, implantation would be intended to be permanent. Immune rejection is the most severe problem, especially if cells are derived from nonhuman animals (e.g., cows or pigs). Although the problems of rejection of xenotransplants are substantial, great progress has been made in understanding the host response and modulating it so as to improve long-term graft acceptance. Immune rejection could also be avoided by encapsulation, so that cells are physically protected from host immune cells. Most promising is the prospect of genetically modifying cells so that they are "invisible" to the host immune system. Alternatively, the patient's own cells could be used in ex vivo gene therapy, but such customized cell therapy would be much more expensive than using "off-the-shelf" cell lines, and the time involved in preparing the cells would prevent this method from being used in diseases where treatment is needed urgently.

The cell types that could be used in cell-based therapies are those that can act as vehicles for a variety of gene products (such as myoblasts and keratinocytes) or those intended to directly replace or restore damaged tissues (such as neurons and chondrocytes). In the latter case various forms of stem cells could be used. Recent advances in stem cell biology, such as the isolation and characterization of human embryonic stem cells, neural stem cells, and mesenchymal stem cells, have brought therapy based on these cell types closer to reality.

Because of the large expansion of the cell population needed in culture, any cell type used in cell therapy must avoid the shortening of telomeres that limits the proliferative potential of somatic cells. Many forms of stem cells are normally telomerase positive, but cells that are not telomerase positive will require genetic modification to prevent telomere shortening. This could be done by introduction of the telomerase reverse transcriptase gene, thereby producing "telomerized" cells. A concern about the use of telomerized cells is that they might have a propensity to undergo neoplastic conversion, but experiments on transplantation of telomerized cells in experimental animals have shown that they produce normal tissue. Another method that has been proposed is to take advantage of the fact that nuclear transfer, the passage of a nucleus and its progeny through the environment of the fertilized egg and early embryo, can restore telomere length when this process is used on cells with short telomeres (senescent cells).

It is important to realize that the same goal (restoration of healthy tissue and prevention of further damage) can potentially be achieved by two forms of cell therapy. In the first, gene products are delivered from transplanted cells and act to affect the behavior of host cells; in the second, the transplanted cells themselves replace the function of the host tissue. Both strategies would be applicable to the treatment and prevention of age-related diseases (see Table 1).

A full treatment of the possible uses of stem cells in human medicine is beyond the scope of this entry. Also not covered are other important topics within the fields of tissue engineering and regenerative medicine, such as the concept of growing or constructing entire organs in vitro, as a source of organs for transplantation, or the production of transgenic "humanized" animals (principally pigs) as a source of organs. In addition, some important molecular therapies are not covered, such as the use of gene therapy in cancer treatment, and possible therapies that use stimulation of the immune system to provide a protective response against the accumulation of damaged molecules, such as β-amyloid in Alzheimer's disease.

Since the beginning of gene therapy as a concept, it has been repeatedly predicted that the use of gene and cell therapy will have a major impact on human medicine, and the fact that no forms of these therapies are yet routine could be viewed as a failure of this technology. However, it must be remembered that most of the great advances in medicine did not find a place in everyday clinical practice for many years after their discovery. More than fifteen years passed between the discovery of penicillin and its routine use in treatment of infectious diseases. There is every reason to believe that gene and cell therapies will play significant roles in treatment of chronic diseases in the elderly, but therapies that aim to alter human aging and change maximal life span are unlikely in the foreseeable future.