17 minutes reading time (3391 words)

    IMMUNOSUPPRESSION IN TRANSPLANTATION

    Michael J.B. Moore
    University of Canterbury
    Private Bag 4800
    Christchurch, New Zealand

    INTRODUCTION
    The kidney transplant has become both a routine and common practice over the latter half of the 20th century. The procedure has a high success rate reflecting the low risk involved in this operation relative to other more complex transplants such as those of the heart, liver, and lung. A stumbling block of transplant surgery today is that the demand for operations exceeds the supply of organs. In the case of the kidney, the organ supply has the potential to meet the demand as most people are born with two kidneys, yet they can live happily with one. In addition, diseases afflicting the kidney are not fatal, and hence the kidney presents itself as the ideal transplant organ. A typical transplant scenario may involve kidney injury to an otherwise healthy individual, for instance in an automobile accident. An indefinite hospitalisation period may then follow when the patient's kidney function is replaced temporarily by an extracorporeal machine until a suitable kidney is found for transplant. The donated organ may come from a relative, for example a parent or sibling, or a cadaver, and once surgically implanted into the patient natural kidney function can resume(1).

    The success of a transplant procedure has typically already been determined before the operation has begun. The major factor limiting the success of a transplant is the compatibility of the donor organ and the recipient. The immune system of the body identifies the transplanted organ as foreign matter and responds in what can be a violent reaction to eliminate it from the body. It is this phenomenon, known as rejection, that forms the major obstacle to a successful transplant procedure. Two methods have been used to prevent rejection. Tissue typing is a way of pretesting the compatibility between donor and recipient before surgery. If the biological make up of the donor and recipient are similar, rejection of the graft is less likely to occur, promoting transplant success. The science of tissue typing is not as advanced as the science of immunosuppression and hence the latter is the more popular means of controlling rejection(2,3). This review is concerned with advances in immunosuppression and the effects they have had on the field of transplantation.

    Described as the "gift of life" transplantation is the replacement of diseased body parts to prolong life. The grafting of skin to treat burns patients has its origins in the 19th century. Primarily due to advances in the field of immunosuppression, success has been obtained with heart, liver, and lung transplants, though these procedures are less advanced than that for the kidney. Combined heart/lung transplants are also possible, as is the replacement of heart valves. Breathing complications arising from the mass of the liver and the problems with organ supply, have led to the development of partial-lobe organ transplants where the function of a complete liver is replaced by a portion. (Brain transplant surgery is still very much a futuristic goal. Despite success achieved with the grafting, of nerve tissue in the treatment of Parkinson's disease, the brain still remains too complex, earning itself the title of being the 'Final frontier' of transplant surgery(4,5).

    HISTORY OF TRANSPLANTATION
    The idea of transplantation has been with the medical community and society for a long time. The transplantation story has its beginnings in 700 B.C. The use of skin grafts in nasal reconstruction was undertaken in ancient Hindu surgeons, as recorded in the Sanskrit. Historical details remain sketchy until 300 AD, when Cosmos and Damien, the Patron Saints of Medicine, performed what has become known as the "fourth century miracle". The two physicians replaced the diseased leg of a parishioner with that of an Ethiopian Moor. The story continues in 1542 when Tagloozi performed similar cosmetic surgery to that reported in 700 8.C., reconstructing noses with skin still partially attached to the forearm. He would then sever the skin from its origin when it obtained adequate growth and blood supply(4).

    Modern history contains the majority of work in transplantation. John Hunter, the father of experimental surgery, undertook the first animal autograft (tissue taken from one part of the body and then placed on a different part of that same body) in 1728 when he implanted the claw of a rooster into its comb. The early 19th century saw Brown and Sequard perform the first interspecies transplant; now the rooster's comb sported a rat's tail. Late in the same century, several transplant methods arose that are clinically useful today. Thierock, in 1886, devised the split-thickness graft used today in the treatment of burns patients, and the corneal graft was developed by Zurin(4).

    At the turn of the century, mainly due to the efforts of Guthrie and Carrel, surgical techniques had progressed so that organs could be moved and their blood supply maintained. In 1902, Emerich Ullman used magnesium tubes to connect an allograft (a graft performed between two members of the same species) kidney to the neck vessels of a dog with successful function. This amazing feat caught the attention of the media. Fuelled by the consequent publicity, new antibiotic therapies and antiseptic techniques, surgeons experimented with animal-human transplants all of which failed, throwing the field of transplantation into disrepute(4).

    At this point in history the main concern of surgeons was the clotting of grafts. Advances in the surgical disciplines contributing to the field of transplantation had minimised this problem, but little had been achieved in combating the sparsely mentioned rejection phenomenon. Attention therefore turned to the field of immunology in order to find a solution to the rejection problem that could arrest transplantation from a state of "clinical moratorium'.

    REJECTION
    The phenomenon of rejection has been observed over several hundred years. In 1898, Winston Churchill donated some skin to an injured soldier, only to find that the skin having adhered for a couple of days, began to slough and peel off. In 1923, Williamson noted the microscopic changes in such rejected allografts and attributed them to a biological incompatibility between donor and recipient. A year later Holman developed the idea of acquired immunity describing the features of rejection. When human transplant studies were reinitiated in the 1930s, it was noted that operations were more successful if the recipient and donor were genetically similar. In 1945, Hufnagel and Landsteiner successfully attached a kidney to the arm of a woman who was comatose from renal failure. The transplanted kidney never developed any measurable function, surviving for only 48 hours, but long enough for the patient's own kidneys to recover (4).

    The advent of extracorporeal kidney dialysis encouraged a number of attempts at thigh kidney transplants in the following years. In the best case a patient was able to leave the hospital for three months; however, as yet there was no long term survival. 1953 saw the first living related donor transplant. A young man in Paris had fallen from a roof and severely injured a kidney, which turned out to be the only one he possessed. A week after the accident, surgeons transplanted a kidney into the young man from his mother. The immediate results were striking. The kidney began to form urine and functioned well for three weeks improving the patient's condition. But on the 22nd day, blood appeared in the urine and soon the kidney stopped functioning. The transplanted kidney had been rejected (1,4).

    The culmination of experience in treating critically ill uremic patients and observing that the immune response (rejection) was not so violent in genetically similar patients led to the first successful organ transplant with long term survival. On December 23, 1954, J.E. Murray transplanted a kidney from a young man to his identical-twin brother, who was severely ill from kidney failure and high blood pressure. With natural kidney function having resumed, the patient's health improved remarkably, and after removal of the diseased native kidneys a full recovery was made. This landmark operation was evidence that long-term transplant success could be obtained if immune factors were controlled. Kidney transplantation had become a life-saving procedure. This however was only true if a patient with serious kidney disease had an identical twin who showed no sign of kidney pathology, was otherwise healthy, and was willing to have a kidney surgically removed. It was to be the advent of immunosuppression that made transplantation accessible to all individuals (1,4).

    IMMUNOSUPPRESSION
    In 1958 Robert S. Schwartz and William Dameschek carried out an experiment that was to prove vital to the success of immunosuppression in transplantation(6). At this time many laboratories made use of antimetabolites in the treatment of cancer. An example of such an antimetabolite is 6-mercaptopurine (1) whose chemical structure is similar to adenine (2) (Figure 1). Adenine is an essential structural element of DNA, which in turn is essential for cell growth. insertion of 6-metacaptopurine into DNA instead of adenine, by the body's biosynthetic machinery would render the cell incapable of division. This should only occur in proliferating cells, hence having a much more toxic effect on an actively growing cancer than on normal body cells.

    At this time, it was believed that the immune response involved the employment of rapidly growing cell populations suggesting that it may be susceptible to anticancer therapies. This hypothesis was supported by experimental results; when an appropriate dosage of 6-mercaptopurine was injected with an immunizing dose of a foreign antigen (bovine serum albumin) into rabbits, no antibody (or a greatly reduced amount) of the antigen was produced (6).

    The experiment created much interest and shortly thereafter, in association with Roy Caine, an English surgeon interested in experimental transplantation, Schwartz and Dameschek demonstrated the use of 6-mercaptopurine in prolonging the survival of skin allografts. Clinical trials followed and two years later, in 1962, azathioprine (Immuran), a derivative of 6-mercaptopurine, became the first commercial immunosuppressant to be used in renal transplant.

    Other chemicals capable of suppressing the immune response, and therefore of dealing with critical "rejection episodes", were also under clinical development at this time. These included methotrexate, (a folic acid antagonist that inhibits the synthesis of nucleic acid), cyclophosphamide, (a N-mustard derivative that attacks DNA by alkylation and crosslinking), corticosteroids (anti-inflammatories), and antilymphocyte globulin (ALG). With the availability of these and azathioprine, a substantial proportion of kidneys taken from accident victims and live donors could be used in transplantations with satisfactory acceptance and prolonged function. Within five years, greater than 50% of the operations were successful in the sense of providing a substantial extension of life without invalidism (2,8).

    THE PROBLEMS OF IMMUNOSUPPRESSION
    It soon became evident, that although a dosage of azathioprine by itself or in combination with other general immunosuppressants could be reduced, it could not be safely discontinued. A suitable maintenance dose having been found, the patient was advised to continue at that level indefinitely, as with the broad spectrum antibiotics now required for resistance to infection (the immune system now being depressed). Transplant patients became afflicted with tumours that could be ascribed to this indefinite maintenance therapy. Increased incidence of skin and cervical cancer was also noted. Statistical comparison showed that immunosuppressive patients were 350 times more susceptible to cancer than their normal counterparts. Other signs of immune depression also observed included the appearance of chronic cold sores, warts and solar keratoses. Given the severity of these side effects induced by general immunosuppressants such as azathioprine specific more efficient immunosuppressive agents were desperately required. This demand was met by nature (8).

    NEW IMMUNUSUPPRESSANTS FROM NATURE
    Cyclosporin
    "Unique", "superior", "wonderful": these were the words used to describe the natural product cyclosporin (see figure 2), which has "revolutionized the field of organ transplantation" through its remarkable activity as an immunosuppressant. The fungal metabolite was isolated from Tolypocladium inflatum gams in 1972 by J.F. Borel at the Sandoz Pharmaceuticals Corporation (8). Coupled with the evolution of surgical and organ procurement techniques, its use led to what the medical community termed a "boom" or "explosion" in the range, number, and combinations of tissues and solid organs that were transplanted from the early 1980s through 1990 in efforts to ameliorate or cure genetic disorders, birth defects, diabetes, and cancer. Cyclosporin has also been accredited with the movement of heart, liver, and lung transplants from experimental operations to accepted modes of treatment. The initial and long-term survival of transplants has improved, immune risk factors involved with transplantation have been eliminated and hospitalization costs reduced (9).

     However since receiving clinical approval, numerous toxic side effects of cyclosporin have been progressively cited. Gastrointestinal cancer, hypertension and renal dysfunction have all been observed in patients treated with cyclosporin. These effects, characterized as "unforeseen" and "surprising" by a shocked medical community, were largely attributed either to the continuing search for the optimal dose of the drug or to complications caused by the "lethal diseases" from which the patients receiving it suffered. The desperate optimistic wish of surgeons to be able to save the organs, therefore the lives of their patients had instilled-a lingering belief that cyclosporin provided an utopian, magic-bullet kind of therapy. In the face of evidence linking patient death to cyclosporin use it was obvious that more potent, selectively effective, and less toxic immunosuppressants were required. Such agents were again to be provided by nature.

    FK506
    Despite predictions of gloom, 1984 heralded the discovery of a new immunosuppressive agent FK506 (see figure 3), which was later described as a "miracle" drug in the same vein cyclosporin had been a decade earlier. It was isolated from Streptomyces tsukubaensis by the Fujisawa pharmaceutical Company in a soil sample screening program (10). Two prominent transplant surgeons, Roy Caine in Cambridge, England and Thomas Staizl of the University of Pittsburgh, U.S.A. obtained samples of FK506 and began in vitro and animal testing to assess its safety and efficacy. Here the two groups came to opposite conclusions.

    The British group judged that FK506 was too toxic to scientifically and ethically warrant human tests. The American group found the drug's toxicity varied from species to species, but in terms of graft survival was 500 times more effective than cyclosporin and with less side effects. Based on their results, the Pittsburgh group decided to initiate clinical testing despite criticism from a medical community now anxious to fully determine toxicity parameters before drug scheduling: a lesson learnt from cyclosporin. In the first human trial the drug was administered as a last resort to ten "gravely ill" patients undergoing rejection of retransplanted organs despite conventional immunosuppression. Of the ten patients, only one died. Seven of the liver grafts were "salvaged", and two were successfully replaced(5).

    From this original trial, the use of FK506 has expanded to include all types of tissue and solid-organ graft, with higher survival and lower mortality rates than those observed with cyclosporin and with less side effects. If thorough clinical testing reinforces early predictions of both its potency and safety, FK506 will usher in a new era of transplant surgery. The drug offers a solution to the acute organ shortages of today by reducing the number of retransplantation operations undertaken and leading to a resurgence of research on animal-human transplants. FK506 also presents itself as a therapeutic agent in the treatment of organ "self destruction" thought to be a major factor in autoimmune disease and graft rejection (11).

    NATURAL PRODUCTS AS MECHANISTIC PROBES
    Cyclosporin and FK506 are only two examples of natural products with immunosuppressive activity. Rapamycin (see figure 4), (isolated from an Easter Island fungus) (12) has biological activity similar to that of FK506 and like the latter has attracted considerable clinical interest. Among the natural products pool there remain other biologically active so-called "orphan" ligands whose modes of action are unknown. Since many of these natural ligands can readily penetrate the plasma membrane of cells they can be used to probe the very same processes they modulate. Hence, in addition to the important medical applications inherent with the discovery of cyclosporine, FK506, and rapamycin, important advances in the understanding of cell biology have been made.

    Signal transduction is the process by which an extracellular molecule can influence an intracellular event. Understanding of these processes has been limited by difficulties experienced in identifying the elements and sequence involved in propagating the signal through the cytoplasm of the cell. Hence this cytoplasmic signal propagation has been referred to as the "black box" of signal transduction. The three natural products cyclosporin, FK506, and rapamycin inhibit intracellular signal transduction pathways that regulate early events in the cell cycle (the universal program instructing cells to divide) of T-cells. Structural studies of these natural products and variants have provided new insights into the molecular architecture involved in signal transduction. For example, a new family of intracellular receptors called immunophilins, believed to have an active role in protein folding and ceil synthesis, have been discovered. Mechanisms involved in cell cycle regulation thought to be impenetrable by contemporary cellular techniques have also been able to be elucidated (10).

    THE FUTURE
    What of future trends in organ transplantation? Advances will be moulded by forces affecting the field today. Numerous educational, social, technological, and economic factors combine to give rise to the great problem of organ supply. Examples include more stringent vehicle safety laws, organ procurement expenses, increased public awareness and advances in medical chemotherapy. The problem of fewer organ donors has been compounded by an increased number in patients who have been accepted as organ recipients. The dwindling donor/recipient ratio has spawned both innovations in surgical procedures for utilizing organs and developments in alternative means of organ replacement.

    Common place today are so-called "domino" transplants. The hearts of patients with pulmonary disease who undergo heart-lung transplant are implanted into individuals with cardiac disease. "Reduced size" and "split-liver" transplants (dividing the liver and implanting its lobes into two recipients) have also been developed. The possibility of using organs from different species has also been reexplored. October 26, 1984 heralded the transplant of a baboon heart into a human infant. "Baby Fae" however died twenty days later. Attempts have been made to replace organs with man-made substitutes. On December 2, 1982 Barney Clark became the first recipient of the Jarvik-7 total artificial heart, an experimental device intended to function as a permanent replacement for the human heart. The death of "second recipient "Bionic Bill' Schroeder in August 1986, 620 days after implantation signalled a temporary end to Jarvik heart experiments and has placed a quasimoratorium on such total artificial devices (5).

    It is the field of immunology however that may provide the breakthrough advance to these experimental techniques as it has done so in the past. Whereas the immunosuppressive properties of cyclosporin initiated the mentioned 'transplant boom' of the 1980s, FK506 has set the stage for radical advances in the field of organ transplantation. In addition to FK506 there exist numerous other natural products with similar therapeutic value. It has been surmised that the family of immunophilins play a role in signal transduction pathways within the brain and hence may provide chemotherapeutic targets for as yet untreatable diseases associated with this complex organ. Success with the treatment of patients suffering from Parkinson's disease has already shown that the study of natural products such as cyclosporin and FK506 and their cellular counterparts may provide a route to the 'final frontiers' of brain transplant surgery and chemotherapy.

    REFERENCES

    1. Merrill J.P., "The Transplantation of the Kidney" from "immunology: readings from Scientific American", Sir F.M. Burnet, W.H. Freeman and Company" San Francisco, 187-93 (1975)
    2. Roitt I.V., "Essential lmmunology',3rd ed., Blackwell Scientific Publications, London, 225-248 (1977)
    3. Longmore D., Ross-Macdonald M., Ed., "Spare Part Surgery: the surgical practice of the future", Aldus Books Ltd, London,11-27 (1968)
    4. Demmy T.L. and Magovern G.A. Jr., "History of Transplantation and Future Trends" from "New Harvest: transplanting body parts and reaping the benefit", Ed., Keyes C.D., Co-Ed., Wiest W.E. Humana Press Inc., Clifton, New Jersey, 78-89 (1991)
    5. Fox R.C., Swazey J.P., "Spare Parts: organ replacement in American society", Oxford University Press, New York, (1992).
    6. Scharwtz R.S. and Dameschek W., Nature,183, 1682 (1959)
    7. Sir F.M. Burnett "Immunodeficiency: Investigations since 1957" from "Immunology: readings from Scientific American", Sir F.M. Burnet, W.H. Freeman and Company, San Francisco, 174-6 (1975)
    8. Borel J.F., Immunology,31,631 (1976)
    9. Oates J.O., Wood A.J.J., Kahan B.D. N. Engl. J. Med.,321 1725 (1989)
    10. Tanaka H., Kuroda A., Marusawa H. et al.. J. Am. Chem. Soc.,109, 5031 (1987)
    11. Starzl T.E. and Fung J.J., J. Am. Med. Assoc.,263. 2686 (1990)
    12. Vezina C., KudelskiA. and Seghal S.N., J. Antibiotics.'.721,28 (1975)
    13. Schreiber S.L., Chem. Eng. News,70 (43). 22 (1992).
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