Abstract

Inhibition of TNFa has become almost commonplace within the past few years. In spite of the popularity of TNFa inhibitors, we have an incomplete understanding of their modes of action. This review will discuss the well-known effects of TNFa and recent hypotheses regarding its actions. The best known effects of TNFa relate to its proinflammatory effects such as induction of adhesion molecules and stimulation of cytokine and chemokine production. Less well known is the fact that TNFa can also have immunosuppressive effects such as inhibition of T cell receptor signaling and the induction of immunosuppressive cytokine expression. Just as TNFa has a broad range of effects, inhibition of TNFa leads to numerous changes in immunologic function.

Clinical examples of TNFa over-production

TNFa over-expression has been documented in a number of inflammatory processes which led to the first successful attempts to block a cytokine therapeutically. A monoclonal antibody to TNFa or infliximab, was originally piloted in rheumatoid arthritis and Crohn’s disease. In both cases the therapeutic effect was dramatic with dose-dependent clinical and laboratory responses. The original rationale for the use of antibodies to TNFa in rheumatoid arthritis was the finding that the two predominant cytokines in synovial fluid are IL-1 and TNFa [1]. These two cytokines act both directly and indirectly. For example, IL-1 and chemokine production are driven largely by the TNFa. TNFa acts directly by promoting the release of metalloproteinases and leukotrienes which are responsible for tissue damage. In Crohn’s disease, TNFa is also elevated, with lamina propria T cells appearing to be largely responsible [2, 3]. This is supported by murine studies of TNFa over-expression. When over-expression is limited to monocytes and macrophages, the phenotype is limited to arthritis and dermatitis. The inflammatory bowel disease phenotype occurs with T cell production of TNFa {Kontoyiannis, 1999 #3479}{Kontoyiannis, 1999 #3479} [4].

While Crohn’s disease and rheumatoid arthritis were the first two disorders in which TNFa inhibition was tested therapeutically, there are now many other disorders in which TNFa inhibition has been tried and found effective (Table 1). These disorders are surprisingly diverse and the exact mechanism by which the TNFa inhibitor is acting is not always known.

While it would be tempting to administer TNFa inhibitors in any setting where TNFa over-expression is documented, caution is warranted as TNFa inhibition has been shown to worsen multiple sclerosis and can induce anti-dsDNA antibodies [5, 6]. In addition, the known risks of infection with intracellular organisms such as Salmonella, Listeria, Histoplasma, Mycobacteria, and Toxoplasma are becoming increasingly recognized. Individual reports of patients with viral infections, sepsis, thrombosis, heart failure, liver failure, and lymphoma also suggest that significant caution is warranted when using TNFa inhibitors. Understanding the mechanism of action of TNFa and the effect of inhibition can clarify some of the clinical effects that have been seen.

Mechanisms of action

Rheumatoid arthritis.

Inflammation is often thought of as an aberrant over-response of the innate immune system. Studies of murine models and in vitro organ culture reveal that inflammation is a complex process which may differ in different organs. An elaborate interaction between the innate and adaptive immune responses is required to sustain an inflammatory response. The best characterized example is the rheumatoid synovium. When actively inflamed, activation of T cells, B cells, macrophages, fibroblasts, endothelial cells and plasma cells can be identified. Pro-inflammatory cytokines are spontaneously produced by explant cultures [1]. Specifically, IL-1, TNFa, lymphotoxin, IL-6, GM-CSF, LIF, IL-12, IL-15, IL-18, and a range of chemokines have been identified. To counter this enormous load of pro-inflammatory cytokines, a number of anti-inflammatory mediators are up-regulated such as IL-10, IL-11, IL-1RA, and other soluble receptor antagonists [7]. The anti-inflammatory cytokines and receptors are insufficient to block the action of the pro-inflammatory cytokines and this leads to fatigue, fever, elevation of acute phase proteins, angiogenesis, bone marrow suppression, increase in adhesion molecules on endothelium, activation of macrophages, and induction of metalloproteinases and leukotrienes (Figure 1). All of these effects contribute to active rheumatoid arthritis. TNFa is a major mediator of both primary and secondary cytokine and chemokine effects. Understanding that TNFa is responsible for so many of the inflammatory pathways activated in rheumatoid arthritis was critical to the development of this important therapeutic strategy (Figure 2). Neutralization of TNFa in synovial organ cultures reduces IL-1, GM-CSF, IL-6, and IL-8 production. This effect is unidirectional because IL-1 inhibition diminishes IL-6 and IL-8 production but not TNFa production suggesting that TNFa is the predominant mediator of rheumatoid synovial inflammation [8].

Juvenile chronic arthritis.

Although juvenile chronic arthritis (JCA) has many similarities with rheumatoid arthritis, the pathologic processes are distinct. TNFa over-expression is implicated specifically in all forms of JCA although each type of arthritis has different cytokine profiles, different pathology, and different natural history.

Systemic JCA has been extensively evaluated for cytokine abnormalities. IL-6, IL-18, IL-8, monocyte chemoattractant protein-1, and migration inhibitory factor have all been found to be markedly elevated in the serum of patients [9-12]. This elevation of IL-6 may in part mediate the angiogenesis and growth failure seen in systemic JCA [13, 14]. TNFa is elevated in synovial fluid, and serum levels are extremely high in patients with macrophage activation syndrome. TNFa inhibitors have been rarely successful in patients with systemic JCA; however, the response is often poor. There is currently a trial using an antibody to IL-6 which is showing early promise [15].

The synovium in pauciarticular and polyarticular JCA can appear quite similar histopathologically; therefore, it is not surprising that the cytokines found in the synovial fluid are similar. TNFa, IL-1, IL-12, monocyte chemoattractant protein-1, interferon-γ, IL-18 and IL-15 are overproduced locally [10, 16, 17]. One significant difference between pauciarticular and polyarticular synovial cytokine expression is that pauciarticular JCA is associated with increased IL-4 expression while polyarticular synovial samples are not [18]. Thus, the secretion of TNFa in the synovium is a commonality exploited therapeutically. The mechanism of action in JCA may, in fact, be quite similar to that described for rheumatoid arthritis.

Systemic effects.

TNFa has numerous systemic effects. High levels of circulating TNFa are known to impair T cell responses and treatment with TNFa inhibitors has been shown to restore T cell function [19]. Aberrant cytokine production by T cells also normalizes after treatment [20]. Furthermore, a recent study in mice demonstrated that the production of the CD4/CD25 T cells in the thymus is impaired in the presence of elevated TNFa and is improved by administration of anti- TNFa antibodies [21]. These CD4/CD25 T cells have been shown to be critically important for the prevention of organ-specific autoimmunity in mice and appear to have a similar role in humans [22]. Other systemic effects have been noted during treatment with TNFa inhibitors. Endothelium-dependent vasodilation improved in patients treated with TNFa inhibitors and endothelial cell expression of adhesion molecules diminished [23]. Macrophage activation was diminished and less nitric oxide was produced after treatment [24, 25]. Bone marrow suppression was reversed and angiogenesis impaired. Thus, TNFa inhibition acts not only in the joint space to suppress inflammation but globally resets the immune system in a favorable way and suppresses the systemic symptoms associated with chronic inflammation such as fatigue and fever.

Mechanisms of adverse events

The features described above all contribute to therapeutic efficacy. The adverse effects associated with its use can also be understood by examining the mechanisms of TNFa action. The risk of infection with intracellular organisms is clearly increased in patients treated with TNFa inhibitors and this risk is derived from the role TNFa plays in the activation of intracellular killing in macrophages. Interferon-g, TNFa and various chemokines participate in granuloma formation which serves to contain and suppress intracellular organism growth. TNFa is also required for intracellular killing of pathogens. Figure 3 demonstrates the cytokine network that governs intracellular killing. It may be seen that TNFa is essential to the process. Thus, it is no surprise that clinical inhibition of TNFa promotes susceptibility to intracellular pathogens. It is less clear whether inhibition of TNFa leads to a greater susceptibility to infection in general. Patients who are unable to respond to interferon-g due to an inherited mutation, have a slightly increased risk of infection with Herpes viruses which may be due to the effect of interferon-g on Th1 cell maturation [26]. Relatively few severe viral infections have been reported in patients receiving TNFa inhibitors and the expectation would be that any increase in susceptibility to viral infections would be modest and more likely to be seen in children when the first exposures to Herpes family viruses occur.

The risk of bacterial infections is almost certainly increased in patients receiving TNFa inhibitors, although clinical data has been difficult to collect. TNFa is released from neutrophils and macrophages upon first encounter with bacterial pathogens. Recognition of pathogens via toll-like receptors leads to TNFa expression along with other proinflammatory cytokines [27]. Early expression of cytokines and chemokines is important to upregulate adhesion molecules to bring other responding cells to the site [28]. TNFa expression also leads to activation of macrophages which improves phagocytosis and antigen presentation. Thus, early TNFa expression is important for innate responses to infection and to improve interactions between macrophages and T cells.

There have been 170 cases of lymphoma occurring in patients receiving TNFa inhibitors as of March 2003 (http://www.fda.gov/ohrms/dockets/ac/cder03.html#Arthritis). Most cases were non-Hodgkins lymphoma. Several of these cases regressed when the patients were taken off their TNFa inhibitor. An interesting feature is that most lymphomas developed very soon after initiation of therapy [29]. At this point, causality has not been demonstrated because patients with autoimmune diseases have an increased risk of malignancy over the general population. Improved epidemiologic analysis should advance our understanding of the relationship of TNFa inhibition and lymphoma development. Nevertheless, there are theoretical reasons to believe the relationship may be real. Both T cells and natural killer cells are important in the surveillance for malignancies [30]. Figure 3 demonstrates the role of TNFa in natural killer cell activation. It is possible that TNFa inhibition leads to impaired natural killer cell function which in turn, allows cells already undergoing malignant transformation to escape detection and proliferate. This is a topic of great importance because patients are being maintained for longer and longer periods on TNFa inhibitors.

Summary

TNFa is well known for its proinflammatory effects and inhibition has unquestionably been an enormously beneficial strategy. Less well known are the immunosuppressive effects of TNFa and the deleterious consequences of TNFa inhibition.

Figure Legends

Figure 1. The roles of TNFa in inflammation and immunosuppression. TNFa, like most members of the TNF family of proteins, has seemingly contradictory roles. It plays important roles in driving inflammation but also has immunosuppressive effects such as bone marrow suppression, induction of apoptosis and inhibition of dendritic cell function.

Figure 2. TNFa stimulates the production of many cytokines. Many of the inflammatory effects of TNFa can be understood by examining the cytokines and chemokines induced in the presence of TNFa.

Figure 3. The role of TNFa in the defense against intracellular organisms and malignancy. TNFa is part of a circuit of cytokines that form the basis for innate responses to infection. Through increasing uptake and killing of microbes, TNFa augments innate defenses. Through increasing co-stimulatory molecule expression and expression of major histocompatibility proteins, TNFa also stimulates the adaptive immune response. Its actions on natural killer cells may augment killing of transformed target cells and the defense against viral infections.

Table 1

Autoimmune disorders treated with TNFa inhibitors

Disorder	Type of study*	References
Rheumatoid arthritis	PC	[31-33]
Crohn’s disease	PC	[34, 35]
Spondyloarthropathy	PC, OL	[36, 37]
Juvenile rheumatoid arthritis	PC	[38]
Psoriasis (skin and arthritis)	PC	[39, 40]
Wegener’s granulomatosis	OL	[41]
Adult onset Still’s disease	OL	[42-44]
Behcet’s	OL	[45]
AA amyloidosis	OL	[46]
Pyoderma gangrenosum	OL	[47, 48]
Chronic inflammatory demyelinating polyneuropathy	OL	[49]
Polymyalgia rheumatica	OL	[50]
Uveitis	OL	[51]
Chronic ITP	OL	[52]
Reactive arthritis	OL	[53]
TNF receptor associated periodic syndrome	OL	[54]
Ulcerative colitis	OL	[55, 56]
Sjogren’s syndrome	OL	[57]
SAPHO syndrome	OL	[58]
Sarcoidosis	OL	[59]

*OL=Open label, PC= placebo controlled

Figure 1

IL-6

IL-8

IL-1

GM-CSF

VEGF

Angiogenesis

Chemokines

Adhesion molecule expression

Figure 2

Figure 3

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