Cyclosporine

Jean F Borel, Götz Baumann and Thomas Beveridge, Sandoz Pharma Ltd, Basel, Switzerland

Copyright © 1998 Elsevier Ltd. All Rights Reserved.

Cyclosporin (WHO)/cyclosporine (US Adopted Name Council)/cyclosporin A (British Approved Name) (CsA) is a cyclic undecapeptide containing a novel amino acid together with several N-methylated amino acids, which is produced as a major secondary metabolite of the fungus Tolypocladium inflatum Gams. This strain of fungi imperfecti was isolated from a soil sample collected in the Hardanger Vidda (Norway). Extracts of cultures exhibit a narrow spectrum of antifungal activities in vitro, but exert only limited effects in vivo. The striking selective immunosuppressive activity discovered in 1972, in an additional pharmacological screening, led to an extensive investigation of this fungal metabolite. CsA is the first immunosuppressive drug to selectively and reversibly act only on a limited population of lymphocytes and to be devoid of myelotoxicity. This explains its immediate impact in the field of transplantation. Today, however, its potential in the treatment of autoimmune diseases is steadily growing. CsA is also widely used as an experimental probe in basic research.

Pharmacology of CsA

The structure of CsA (Figure 1) was established by chemical degradation and X-ray analysis of an iodo-

MeLeu—Me Val -MeLeu

MeLeu—Me Val -MeLeu

Figure 1 Structure of CsA corresponding to the conformation observed in the crystal. (Reproduced with permission from Borel JF (ed) (1986) Ciclosporin. Progress in Allergy 38: 48. Published by Karger, Basel.)

derivative. Having a molecular weight of 1202, C62H| i |N, 1O12, CsA is a neutral, hydrophobic, cyclic peptide with 11 amino acid residues, all having the l-configuration of the natural amino acids, except for the D-alanine in position 8 and the achiral sarcosine in position 3. One amino acid had never been seen before. Located in position 1, it is abbreviated as MeBmt, for (4R)-4-[(£)-2-butenylj-4,N-dimethyl-l-threonine.

The molecular mechanism of action of CsA in the antigen-driven T cell activation process is shown in Figure 2. In contrast to previous immunosuppressive drugs (mostly cytostatic drugs), CsA exerts a specific-action on lymphocytes, but it does not influence the functions of phagocytes or hematopoietic stem cells. It is neither lymphocytotoxic nor mutagenic, and its action is reversible. Besides being accepted today as the first-line immunosuppressive drug for clinical use, CsA is also widely used as an experimental probe in basic research. Especially for immunolo-gists, it has become an important pharmacologic tool for defining in vitro the respective roles of cell interactions and mediators in lymphocyte activation, analyzing the cell regulatory mechanism of the genes, and studying the different steps involved generally in an immune response. Though in vitro several pathways can lead to the induction of gene transcription in lymphocytes, only pathways involving mobilization of intracellular calcium are CsA sensitive. There is compelling evidence that CsA acts by preventing the activation of resting lymphocytes at an early stage of the cell cycle (G0 to G, transition), thus inhibiting primarily the production and release of interleukin 2 and several other lymphokines by helper T cells.

CsA has been widely used in vivo to evaluate the contribution of various T cells not only in normal but also in pathological conditions. Since the drug can induce and maintain a remission in many models of autoimmunity as well as in clinical autoimmune diseases, it is evident that it also inhibits fully activated T cells, possibly by suppressing their continuous de novo restimulation. The in vivo mechanism of action is still being debated. However, most observations resulting from many different types of in vivo models suggest that CsA acts by suppressing lymphocyte activation within the allograft in transplantation or at the site of (chronic) inflammation in autoimmunity. Although necessary and often helpful, ex

Figure 2 General overview of T cell signaling pathways following ligation of the T cell receptor (TCR) or the IL-2 receptor (IL-2R). Mode of action of CsA. In the presence of costimulatory signals provided by multiple cell surface receptor-ligand interactions (e.g. CD28/B7-1, 2; LFA-1/ICAM; CD2/LFA-3; CD45/CD22; IL-1R/IL-1) antigen (AG) presented by an antigen-presenting cell (APC) in the context of major histocompatibility complex (MHC) proteins induces a T cell differentiation process that results ultimately in lymphokine secretion (G0 to G, transition of cell cycle) and proliferation (G, to S) of the antigen-specific T cell. Signal transduction involves a number of biochemical events, including phosphorylation and dephosphorylation (P) of tyrosine, threonine or serine residues of intracellular signaling proteins, leading to gene transcription of 'early' genes like IL-2. CsA exerts its immunosuppressive effects downstream from the very early membrane-associated events, such as activation of phospholipase Cy (PLC-y) and recruitment of tyrosine kinases (e.g. LCK, FYN, CSK, ZAP-70) and SHC to the 'immunoreceptor tyrosine-based activation motifs' (ITAM) in the t and J chains of the T cell receptor (TCR)/CD3 complex. It is hypothesized that cyclophilin-bound CsA inhibits the calcium-dependent (Ca21) phosphatase activity of calcineurin (CN-A, -B and Cam, calmodulin) as a crucial step in the activation (*) and nuclear translocation of cytoplasmic transcription factor subunits like NFATc/p and/or NKkB required for early' gene transcription. IL-2R-mediated signaling processes involving activation of JAK kinases, STATs (signal transducers and activators of transcription). RAFTs (rapamycin and FKBP target) and p70 S6 kinase (p70S6K) are not affected.

Figure 2 General overview of T cell signaling pathways following ligation of the T cell receptor (TCR) or the IL-2 receptor (IL-2R). Mode of action of CsA. In the presence of costimulatory signals provided by multiple cell surface receptor-ligand interactions (e.g. CD28/B7-1, 2; LFA-1/ICAM; CD2/LFA-3; CD45/CD22; IL-1R/IL-1) antigen (AG) presented by an antigen-presenting cell (APC) in the context of major histocompatibility complex (MHC) proteins induces a T cell differentiation process that results ultimately in lymphokine secretion (G0 to G, transition of cell cycle) and proliferation (G, to S) of the antigen-specific T cell. Signal transduction involves a number of biochemical events, including phosphorylation and dephosphorylation (P) of tyrosine, threonine or serine residues of intracellular signaling proteins, leading to gene transcription of 'early' genes like IL-2. CsA exerts its immunosuppressive effects downstream from the very early membrane-associated events, such as activation of phospholipase Cy (PLC-y) and recruitment of tyrosine kinases (e.g. LCK, FYN, CSK, ZAP-70) and SHC to the 'immunoreceptor tyrosine-based activation motifs' (ITAM) in the t and J chains of the T cell receptor (TCR)/CD3 complex. It is hypothesized that cyclophilin-bound CsA inhibits the calcium-dependent (Ca21) phosphatase activity of calcineurin (CN-A, -B and Cam, calmodulin) as a crucial step in the activation (*) and nuclear translocation of cytoplasmic transcription factor subunits like NFATc/p and/or NKkB required for early' gene transcription. IL-2R-mediated signaling processes involving activation of JAK kinases, STATs (signal transducers and activators of transcription). RAFTs (rapamycin and FKBP target) and p70 S6 kinase (p70S6K) are not affected.

vivo results may be misleading, because the experimental reactions are occurring outside the organism in which CsA is present in biologically effective levels.

CsA in transplantation

CsA was first registered in 1983 (trade name Sandimmune*) for use in organ transplantation. Recently, a new microemulsion formulation of CsA (trade name Sandimmun Neoral® or Neoral®) which shows more consistent oral absorption, is bile independent, and has dose linearity, has become available. This has the advantage that its absorption is not affected by food intake and it shows reduced variability between as well as within patients. Neoral* has been shown to be safe and well tolerated.

CsA is used for the prevention of acute rejection of kidney, liver, heart, heart-lung, lung and pancreas allografts. It may also be used to treat rejection episodes in patients who have already received other immunosuppressants. In bone marrow transplantation, CsA is used to prevent rejection of the transplanted bone marrow as well as for prevention and treatment of graft-versus-host disease (GVHD). In transplantation CsA has long become established in all drug regimens and is today the drug of first choice in immunosuppression. However, it is seldom used alone but generally in combination with other immunosuppressive agents. These drug combinations have the advantage of exploiting additive and synergistic drug effects while concomitantly minimizing the adverse reactions.

From the very beginning the steroid-sparing potential of CsA has been a main consideration in all clinical situations. In transplantation a state of true tolerance has not been achieved (with the exception of bone marrow transplants) and, therefore, as with other immunosuppressants, lifelong therapy is necessary. Although acute rejection can now be effectively controlled, there remains, in the long-term, the problem of chronic rejection which is considered as one of the leading causes of late allograft failure.

The kidney is the most frequently transplanted organ. Before the CsA era the overall one-year graft survival rate was about 60%, depending on the center. Since the introduction of CsA it has now risen to 80-90%. In kidney recipients followed for up to 10 years renal function tended to worsen during the first year, but remained stable thereafter with no progressive renal damage. The use of CsA in liver transplantation had the effect of revitalizing the field and soon this procedure was accepted as a treatment option for end-stage liver disease and no longer considered experimental. The 5-year survival of patients, which was about 20% before the use of CsA, increased to 60%. With CsA the number of heart transplantations increased rapidly and currently a 5-year survival rate of approximately 70% is obtained. Heart-lung and lung transplantation was never successful without CsA. The one-year survival of the former has been about 60-65% with CsA. Transplantation of the pancreas simultaneously with a kidney offers a successful treatment for patients with type I diabetes mellitus and end-stage renal failure. A one-year graft survival of about 80% can be achieved at the best centers. In bone marrow transplantation the use of CsA has resulted in a reduction in the severity of graft-versus-host disease. Its use is also associated with prompt and sustained engraftment, quicker hematological recovery and a shorter hospital stay.

CsA in autoimmunity

Since 1987 CsA has also been registered for the treatment of several autoimmune disorders. The use of CsA in this field was based on a solid pharmacological rationale and was also a logical consequence of its superior efficacy in organ transplantation. By their very nature the autoimmune diseases demand a different evaluation of the risk-benefit ratio inherent in all forms of therapy. Indications subjected to investigation were selected on the basis of the experimental results in animal models of autoimmunity and on the medical need. The efficacy of CsA has been proven in several such diseases with clinical benefit being demonstrated in some and remaining uncertain in others as shown in Table 1.

CsA is efficacious in many autoimmune diseases, both for induction and maintenance of remission. Common features of the clinical effects of CsA are relatively quick onset of effect and also the occurrence of relapses when treatment is stopped. The autoimmune reaction seems temporarily 'frozen', without correction of the intrinsic autoimmune defect. Nevertheless, it is evident that many patients who do not respond satisfactorily to conventional therapy, in terms of efficacy or toxicity, may profit from CsA. However, in therapy-resistant end-stage patients who present with substantial pathology which is irreversible, the full potential cannot be realized. The use of the drug may thus eventually be extended to less advanced disease stages.

Table 1 Effect of CsA on autoimmune diseases in humans

Efficacy proven by controlled studies Established benefit in severely affected patients resistant to conventional therapy Uveitis (autoimmune, Behget disease) Psoriasis

Idiopathic nephrotic syndrome Rheumatoid arthritis Unclear benefit Severe aplastic anemia Crohn disease Atopic dermatitis Asthma

Primary biliary cirrhosis Myasthenia gravis Insulin-dependent diabetes mellitus Efficacy suggested by uncontrolled studies Systemic lupus erythematosus Dermatomyosltis/polymyositis Ulcerative colitis Pyoderma gangrenosum

Membranous nephropathy and other autoimmune glomerulonephrltides Efficacy questionable Multiple sclerosis Endocrine ophthalmopathy Sjögren syndrome Sarcoidosis Pemphigus Polychondritis

Adverse events

One of the most important issues is safety on long-term treatment. The adverse events recorded are similar in all indications. Apart from the general risks of immunosuppression (opportunistic infection, malignancy) the most important side-effects of CsA are nephrotoxicity and hypertension. Others include anorexia, nausea, vomiting, paresthesia, hypertrichosis, gingival hyperplasia and tremor.

Although infections do occur with CsA, they appear to be more amenable to treatment. The occurrence of lymphomas or lymphoproliferative disorders is now recognized to be the result of over-immunosuppression. Experience gained so far indicates that the incidence of cancers following the use of CsA is no higher than with other types of immunosuppressive therapy. Today the nephrotoxicity of CsA is considered to be manageable. This is achieved, in transplantation, by dosage adjustments based on the monitoring of CsA blood concentrations. Added to that is the increased understanding of the nephrological findings in kidney biopsies from patients exposed to the drug. In autoimmune diseases CsA is kept at the lowest effective dose, i.e. exceeding 5 mg kg"1 day-1, and this is decreased whenever serum creatinine increases by more than 30% over the individual baseline value. In addition, regular monitoring of blood pressure is recommended. Hypertension, if it occurs, is responsive to the standard therapy. One must also be aware of pharmacokinetic interactions which may result in either an increase (e.g. ketoconazole) or decrease (phenytoin) of the CsA blood concentration.

See also: Anti-inflammatory (nonsteroidal) drugs; Antilymphocyte serum; Autoimmune diseases; Cytokines; Effector lymphocytes; Glucocorticoids; Graft rejection; Graft-versus-host reaction; Immunosuppression; Lymphoma; Monoclonal antibodies (mAbs); Opportunistic infections; T lymphocytes; Tolerance, peripheral; Transplantation.

Further reading

Borel JF, Baumann G, Chapman I et al (1996) In vivo pharmacological effects of ciclosporin and some analogues. Advances in Pharmacology 35: 115-246. Borel JF, Di-Padova F, Mason J et al (1989) Pharmacology of cyclosporine. Pharmacology Review 41: 239-434. Borel JF, Feutren G, Baumann G and Hiestand P (1994) Cyclosporin and some new immunosuppressive drugs in the treatment of autoimmune diseases. In: Lydyard PM and Brostoff J (eds) Autoimmune Disease. Aetiopathogenesis, Diagnosis and Treatment, pp 169-217, Oxford: Blackwell Science. Borel JF, Kis ZL and Beveridge T (1995) The history of the discovery and development of cyclosporine (Sandimmune®). In: Merluzzi VJ and Adams J (eds) The Search for Anti-inflammatory Drugs, pp 27-63, Boston: Birkhàuser. Bram RJ, Hung DT, Martin PK, Schreiber SL and Crabtree GR (1993) Identification of the immunophilins capable of mediating inhibition of signal transduction by cyclosporin A and FK506: roles of calcineurin binding and cellular location. Molecular and Cellular Biology 13: 4760-4769.

Ke H, Myarose D, Belshaw PJ et al (1994) Crystal structures of cyclophilin A complexed with cyclosporin A and N-methyl-4-[(E)-2-butenyl]-4,4-dimethylthreonine cyclosporin A. Structure 2: 33—44. Thomson AW (ed) (1989) Cyclosporin. Mode of Action and Clinical Applications. Dordrecht: Kluwer Academic.

Thomson AW and Starzl TE (eds) (1994) Immunosuppressive Drugs. Developments in Anti-rejection Therapy. London: Edward Arnold.

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