Clinical and Experimental Dermatology (1979) 4, 151.

Immunosuppressive drugs: how do they act?

A.M.DENMAN Clinical Research Centre, Division of Immunological Medicine, Watford Road, Harrow HAi 3UJ

Accepted for publication 31 Jw/y 1978

Cytotoxic drugs were first introduced into clinical medicine for the treatment of malignant lymphoproliferative disorders such as Hodgkin's disease and various forms of leukaemia. These drugs were used primarily for their ability to interfere with cell division since it was thought that the apparently more rapid growth of malignant cells would make these selectively vulnerable. There were relatively few diseases of the skin or other tissues which, on this basis, would then have been considered suitable for treatment with cytostatic drugs. The demonstration that cyclophosphamide, azathioprine and chlorambucil, amongst other drugs, are also immunosuppressive under certain experimental conditions coincided with an era of increased speculation that many chronic inflammatory diseases of unknown aetiology may have an immunological basis. Accordingly cytotoxic drugs have been used in a variety of such diseases. Although primarily the province of other disciplines, many of these are of interest to dermatologists because the skin is involved in the disease process. Diseases which have received particular attention are dermatomyositis, polyarteritis in its varying forms, systemic lupus erythematosus (S.L.E.), Behcet's disease and rheumatoid arthritis. There have indeed been numerous reports that cytotoxic drugs have induced worthwhile remissions of these diseases including their cutaneous manifestations. Nevertheless, few proper controlled trials have been reported and, of these, a relatively small number have shown clearly that the patients benefited. Ironically, there have also been increasing doubts that any amelioration of the disease that has been produced can be attributed to the immunosuppressive effects of these drugs. For the purposes of this paper we will side-step the clinical issues and accept that cytotoxic drugs do limit disease activity in these conditions. Instead we will consider three points related to the mechanisms by which these drugs might have this suppressive effect. Of these, the first concerns the issue of whether or not cytotoxic drugs are immunosuppressive. The second deals with the biological lessions which can be drawn from the observation that cyto* Symposium held in the Edward Lewis Theatre, Middlesex Hospital, London on 6 April 1978. 0307-6938/79/0600-0151 $02.00 ©1979 Blackwell Scientific Publications

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toxic drugs are undoubtedly effective in controlling some malignant lymphoproliferative disorders, at least temporarily. Finally, we will look at a more speculative possibility, namely that lymphoreticular cells, which are among the targets for cytotoxic drugs, harbour persistent virus infections and that the lysis of these cells exposes previously sequestered infectious agents to host cell defence mechanisms including neutralizing antibody and complement. Are cytotoxic drugs immunosuppressive ? The lymphopenia commonly induced by cytotoxic drugs in patients with connective tissue diseases suggests at first sight that these drugs are immtinosuppressive. However, even if this is the case, we are still forced to ask whether these forms of immune response that are affected are relevant to the pathogenesis of these diseases. We should recall that secondary antibody responses in experimental animals are highly resistant to cytotoxic drugs or even high dose X-irradiation and that in those diseases in which immunological mechanisms are most clearly of primary pathogenetic importance such as myasthenia gravis, auto-immune haemolytic anaemia and Goodpasture's syndrome, it is precisely this form of immune response which clinicians are attempting to control. Predictably, when immtme responses in patients receiving these drugs have been formally tested, it has been found that secondary antibody responses are little affected (Swanson & Schwartz, 1967; Denman et al., 1970). Moreover, even delayed hypersensitivity skin reactions and lymphocyte reactions in vitro are not invariably suppressed by cytotoxic drugs, and may even be increased, particularly if these reactions were depressed in the course of the disease before treatment (Fig. i). Similarly, antibody titres commonly rise steadily throughout a period of intensive treatment with cytotoxic drugs (Table i). Indeed, such observations have prompted speculation, albeit on slender evidence, that the therapeutic effects of cytotoxic drugs should more properly be attributed to 'immunoenhancement'. Despite these reservations it is an inescapable clinical fact that patients receiving these drugs are more vulnerable to a wide range of infectious agents. This susceptibility follow the demonstrated depletion of the many cell populations which mediate the host response to infection in a predominantly non-specific manner. These include granulocytes, K cells and cells of the monocyte-macrophage series. Such effects have commonly been termed 'antiinflammatory' but the distinction between these and 'immunosuppression' is largely semantic. However, specific immune responses continue to be generated despite temporary interference with the many forms of effector cell. Since conventional secondary antibody responses, both in theory and practice, are little affected by cytotoxic drugs, it is surprising that circulating auto-antibodies such as rheumatoid factor and antibody to nucleic acid antigens are suppressed in patients receiving cytotoxic drugs. This paradox suggests that the cells synthesizing auto-antibody are affected by cytotoxic drugs. The concept that auto-immune diseases are primarily benign lymphoproliferative disorders has been discussed for many years but has not so far received any experimental support. Malignant lymphoproliferative diseases have been successfully treated with cytotoxic drugs and the manner in which these drugs act in such diseases is clearly of interest to physicians attempting to adopt the methods of cancer chemotherapy to the treatment of other diseases.

Immunosuppressive drugs 10'-,

Control

Post

10'

10" Cell concentration/culture

Figure i. Increased responsiveness to PHA of blood lymphocytes from a patient receiving cytotoxic drugs. 28-year-old female patient with Behcet's disease receiving steroids, azathioprine and anti-lymphocyte globulin. Results pre- (•) and after one month's treatment (o). Same normal control (broken lines) studied on each occasion. PHA concentration 5 /jg/ml; cultxire volume 025 ml. Data by courtesy of Dr Stella Knight.

Table i. Failure of cytotoxic drugs to suppress an antibody response, 54-year-old female patient with dermatomyositis^ confirmed by muscle biopsy, E.M.G. and serum enzyme changes

Date (1977) January February March

Treatment

Coxsackie B4 antibody* (reciprocal titre)

Nil

Steroids; azathioprine antilymphocyte globulin Steroids; azathioprine antiiymphocyte globulin * Data of Dr Geraldine Cambridge.

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A.M.Denman Stem-cell 'Clonal proliferation'

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Active disease Platelets CML cells

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Figure 2. Clonal origin of 'malignant' and 'normal' blood cells in chronic myeloid leukaemia. Ph ^ Philadelphia chromosome; + present or - absent. Clonal or non-clonal origin determined by analysis of G-6-PD phenotype.

The action of cytotoxic drugs in malignant lymphoproliferative diseases Traditionally the ability of cytotoxic drugs to eliminate leukaemic cells and other abnormal lymphoid cells whilst allowing the normal bone marrow elements to recover has been attributed to the greater susceptibility of rapidly dividing malignant cells to chemotherapeutic agents. This view has been shaken by discoveries in two principal directions. Firstly, the proliferative capacity of malignant cells has been shown to be often very much lower than that of normal bone marrow cells. Secondly, studies in black female patients who are heterozygous for the X-linked enzyme, glucose-6-phosphate dehydrogenase (G-6-PD) have shown that many forms of acute and chronic leukaemia arise in a single, malignant stem cell, i.e. these are truly 'clonal' diseases (Fialkow, 1974). More surprisingly, when cytotoxic drug treatment induces a remission in diseases such as chronic myeloid leukaemia (CML), not only are the apparently normal granulocytes still derived from the malignant clone, but the red cells and other formed elements in the blood are, with the exception of T lymphocytes, also the progeny of this same progenitor cell (Fig. 2). Thus the failure of the bone marrow to produce the normal formed elements of the blood results not simply from the crowding out of normal stem cells by malignant cells but from the re-population of the marrow and blood by the poorly differentiated progeny of a single abnormal stem cell. Cytotoxic drugs induce remissions by allowing a more orderly differentiation of this clone and not by killing rapidly proliferating malignant cells of multifocal origin. The possibility that cytotoxic drugs efFect the differentiation of autoreactive lymphoid cells or of target cells for such lymphoid cells has received little consideration. However, it is a common observation that bone marrow function is often depressed in patients with autoimmune disorders such as systemic lupus erythematosus. As in malignant ljnnphoproliferative disease, clinical remission induced by cytotoxic drugs is frequently accompanied by a reversal of this depression suggesting that the normal sequence of stem cell differentiation and maturation has been restored.

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Virus replicalion

-2 150 PHA response 100 50

0

12 Time (months)

Figure 3. Restoration of normal virus growth in lymphocytes of a patient with dermatomyositis. Patient with dermatomyositis treated with steroids, azathioprine and antilymphoeyte globulin from |. Virus replication: Negative figs (below the broken Une): herpes simplex virus fails to grow. Positive figs: virus growth (Denman et al., 1976). PHA response: normal range hatched: supranormal response falls below normal range.

Lympho-reticular cells and persistent virus infection The viral aetiology of chronic infiammatory and auto-immune diseases has been repeatedly debated and explored but with meagre results. Only with respect to hepatitis B virus in some forms of polyarteritis nodosa has tangible evidence of virus infection been forthcoming. For a variety of reasons, lympho-reticular cells are plausible sites of virus persistence. In an indirect attempt to obtain evidence for this idea, we are examining the ability of viruses which invariably grow in normal lymphocytes to replicate similarly in lymphocytes isolated from the blood and tissues of patients with connective tissue disorders. Herpes simplex virus commonly fails to grow in lymphocytes from abnormal donors, a finding which may reflect interference by defective virus particles in usually permissive lymphocytes although this is by no means the only explanation to be entertained (Denman et al., 1976). One point that has emerged from these studies is that therapeutic regimes which include cytotoxic drugs generally reverse this non-permissiveness of lymphocytes irrespective of their effect on the in vitro responsiveness of the same lymphocytes in conventional immunological terms. Thus, lymphocytes from patients who are receiving cytotoxic drugs support the growth of herpes simplex virus despite a range of immunological responsiveness which varies from increased to severely depressed (Fig. 3). If the regenerating lymphocyte populations which replenish the peripheral blood pool no longer carry interfering virus particles, we can speculate that these cells would support the growth of indicator viruses in a normal fashion. Moreover, the lysis of cells which provided a privileged site for virus persistence inaccessible to the

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Figure 4. Cytotoxie drugs in the treatment of S.L.E. Patient with generalized vasculitis, renal disease, pulmonary oedema, staphylococcal septicaemia, neutropenia and the serological features of S.L.E. treated with antibiotics and the drug regime indicated (AZA = azathioprine, 150 mg daily).

patient's anti-viral immune mechanisms could allow the released virus to be neutralized by, for example, antibody in combination with complement and by other forms of host defence. Conclusions Despite the dearth of proper clinical trials there is a widespread impression that cytotoxic drugs in combination with steroids control many inflammatory diseases of unknown aetiology, including skin manifestations, better than steroids do alone. The clinical course summarized in Fig. 4 illustrates the kind of remission which can be achieved by such drug combinations. However, imaginative, carefully controlled studies are needed not simply to establish the therapeutic efficacy of cytotoxic agents but also to analyse their mode of action. Slavish adherence to the original notion that immunosuppression must explain any benefits of these drugs in immunological disorders has not helped matters because it has diverted attention from the central issue, namely our failure to understand how these drugs act even when their therapeutic benefit is undeniable. References A.M., PELTON, B.K., APPLEFORD, D . & KINSLEY, M . (1976) Virus infections of lymphoreticular cells and auto-immune diseases. Transplantation Review, 31, 79-115. DENMAN, E.J., DENMAN, A.M., GREENWOOD, B.M., GALL, D . & HEArH, R.B. (1970) Eailure of cytotoxic drugs to suppress immune responses of patients with rheumatoid arthritis. Annals of the Rheumatic Diseases, 29, 220-231. FIALKOW, P.J. (1974) The origin and development of human tumors studies with cell markers. Nezv DENMAN,

England Journal of Medicine, 291, 26-35.

M.A. & SCHWARTZ, R.S. (1967) Immunosuppressive therapy. The relation between clinical response and immunologic competence. New England Journal of Medicine, 277, 163-170.

SWANSON,

Immunosuppressive drugs: how do they act?

Clinical and Experimental Dermatology (1979) 4, 151. Immunosuppressive drugs: how do they act? A.M.DENMAN Clinical Research Centre, Division of Immu...
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