John W. Baddley, MD, MSPH

Introduction

The number of patients receiving immunosuppressive agents has increased over the past decade and is expected to increase substantially in the near future. Therapeutic antibodies have become an important class of drugs, with annual sales that exceeded $27 billion in 2007[(A):1].  Important new therapeutic biologics include the TNF-a inhibitors (TNFi) infliximab, etanercept, adalimumab, certolizumab pegol and golimumab. These drugs are approved for the treatment of one or more of a number of diseases including rheumatoid arthritis, psoriatic arthritis, plaque psoriasis, Crohn’s disease, juvenile idiopathic arthritis and ankylosing spondylitis.  Although these drugs have revolutionized the treatment of inflammatory and rheumatologic disorders, there has also been an important safety concern:  the use TNFi is associated with an increased incidence of serious infections caused by a broad spectrum of pathogens, including viral, bacterial, mycobacterial, fungal, and protozoal organisms[2-7]. This review will summarize available data on TNFi and invasive fungal infections.
Biology of Tumor Necrosis Factor-α (TNF-α)

TNF-α, described in 1975, is a cytokine that induces responses in both innate and adaptive immunity.  It is expressed in T cells, macrophages, monocytes and Natural Killer (NK) cells.  TNF-α is synthesized initially as a 233 amino acid transmembrane precursor protein, and later cleaved by TNF-α converting enzymes to the 157 amino acid residue soluble TNF-α that is released into circulation and mediates its biological activities[8].

The biologic activity of TNF-α requires the aggregation of three TNF-monomers to form trimeric TNF-α, which then binds to two types of receptors: TNF-R1 (TNFRSF1A,  CD120a) or TNF-R2 (TNFRSF1B, CD120b) which are expressed on most nucleated cells  [8-10].  Soluble and transmembrane TNF-α bind to both receptors, but the biological activities of transmembrane TNF-α are mediated primarily through TNF-R2 [8].  The receptors exert multiple effects on the immune system, including 1) stimulation of inflammatory cytokines (IFN-g, IL-1, IL-6, IL-8);  2) upregulation of expression of endothelial adhesion molecules (ICAM-1, VCAM-1, E-selectin) and chemokines (MIP-1α, RANTES);  3) effective granuloma maintenance; and 4) and mediation of cytotoxic activity [8, (A):11, (A):12].

Currently Licensed TNF-α Inhibitors

TNF-α inhibitors are important advances in the treatment of a number of inflammatory conditions, including rheumatoid arthritis (RA), seronegative spondyloarthropathies and  inflammatory bowel disease [(A):13].   Five TNF-α inhibitors, infliximab, etanercept, adalimumab, certolizumab pegol and golimumab are FDA-approved for use in the United States (Table 1). The agents differ in structure, pharmacokinetic properties, tissue distribution and functional properties.

Infliximab (Remicade) was licensed in 1998 for the treatment of RA, psoriatic arthritis (PSA), ankylosing spondylitis (AS), Crohn disease and ulcerative colitis [(A):14].  It is a chimeric IgG1 monoclonal antibody with human constant and murine variable sequences. It binds and neutralizes both soluble and transmembrane TNF-α. Infliximab also fixes complement, inducing antibody-mediated lysis of cells expressing membrane-bound TNF [8].

Etanercept (Enbrel) was licensed for the treatment of RA, juvenile RA (JRA), PSA, AS and plaque psoriasis [(A):15-17].    Etanercept is not effective for Crohn’s, Wegener’s or sarcoidosis.  Etanercept is a human soluble TNF-α receptor fusion protein that contains the extracellular portion of two human TNF-R2 receptors linked to the Fc portion of human IgG1.  Etanercept binds soluble TNF-α and TNF-β (lymphotoxin), but has much less affinity to transmembrane TNF-α when compared with infliximab, adalimumab, or certolizumab [8].   In contrast to infliximab, etanercept does not fix complement, cause antibody-dependent cytotoxicity, or trigger T-cell or monocyte apoptosis [(A):10]. For these reasonr, theoretical risk of granulomatous infection with etanercept may be lower than with other TNFi [7, 9, (A):10].

Adalimumab (Humira) is licensed for the treatment of RA, AS and psoriatic arthritis [(A):18].  Adalimumab is a human IgG1 monoclonal antibody that binds both soluble and transmembrane TNF-α, but not TNF-β.  Similarly to infliximab, adalimumab also fixes complement, induces antibody-mediated lysis of cells and induces apoptosis of cells expressing membrane-bound TNF.

Certolizumab Pegol (Cimzia) was licensed for the treatment of Crohn’s disease and RA.  Certolizumab is a Fab’ fragment of an anti-TNF-α IgG1 monoclonal antibody and is lacking the Fc portion. The hinge region is covalently linked to two chains of polyethylene glycol, resulting in certolizumab pegol [(A):19].  Certolizumab binds and neutralizes both soluble and transmembrane TNF-α, but does not fix complement or induce apoptosis of T cells or monocytes due to the lack of the Fc portion.

Golimumab (Simponi) was approved in 2009 for the treatment of RA, PsA and AS [(A):20, (A):21].  Golimumab is a human IgG1 monoclonal antibody that binds both soluble and transmembrane TNF-α, but does not fix complement or induce apoptosis of T cells or monocytes.

TNF-α inhibitors and Invasive Fungal Infection

Despite the clinical benefits associated with TNFi, their use is associated with an increased incidence of serious infections caused by a broad spectrum of pathogens [3, 5-7, (A):22, (A):23].  However, the attributable risk for infections has been difficult to characterize for several reasons: First, patients with inflammatory or autoimmune disorders often have impaired immune responses to infection. Second, many patients treated with TNF-α inhibitors have more severe disease, and therefore may be more likely to develop infection. Third, patients on TNFi are often receiving other immunosuppressive medications (corticosteroids, methotrexate), making it difficult to ascertain risk of infection from a particular drug.  Recently, there have been increasing reports of the relationship of TNF-α inhibitors and invasive fungal infection [4, 6, 11, 12, (A):24, (A):25]; however, the true incidence is unknown.  A recent prospective study, the French RATIO registry, identified non-tuberculous opportunistic infections associated with TNF-α therapy [6]. Of 45 cases identified, 10 (22%) were caused by fungal organisms. A summary of fungal infections associated with TNFi follows.

Histoplasmosis

TNF-α is an important part of the protective immune response to both primary and secondary histoplasmosis.  In animal models, TNF-α is released after a pulmonary challenge with H. capsulatum cells [(A):26]; and TNF-a neutralized mice cannot control infection [(A):27, (A):28]. Thus, there is a theoretical risk for histoplamsosis in patients receiving TNFi.

Histoplasmosis is the most common fungal infection in patients receiving TNF-α inhibitors [4, 24].   To date, approximately 100 cases have been reported in the literature and over 250 have been evaluated by the FDA [4, 29-33]. The incidence of histoplasmosis in patients receiving TNFi has been estimated by Wallis and others with use of data from the FDA Adverse Event reporting System (AERS) [7]. Wallis and colleagues collected data from cases reported to AERS from January, 1998 through September, 2002 and identified 40 cases of histoplasmosis.  The estimated rate of histoplasmosis, per 100,000 patients treated was 18.78 in patients treated with infliximab and 2.65 in patients treated with etanercept [7].

Soon after the release of the first TNFi in cinical practice, several case reports or series were published [30, 31, 33].  In summary, most patients had received infliximab, and few etanercept.  Pneumonia and/or progressive disseminated histoplasmosis (PDH) were common.  In 2008, the FDA reviewed 240 cases of histoplasmosis in patients treated with infliximab (207 cases), adalimumab (16 cases), or etanercept (17 cases). Most cases arose from endemic areas, and delay of diagnosis was not uncommon, occurring at least 21 patients. Of those 21 patients, 12 (57%) died.

The largest cases series was reported recently by Hage and colleagues from the Indiana University Medical Campus [4].  They identified 19 patients diagnosed with histoplasmosis while receiving TNF-α inhibitors from 2000-2009.  Seventeen (89%) patients presented with PDH; pulmonary involvement was present in 15 (79%) patients.  Thirteen cases occurred in patients on infliximab, 3 on etanercept and in 3 receiving adalimumab. Duration of treatment with TNF-α inhibitors ranged from <1 month to 3 years.  Fifteen (79%) patients required hospitalization, and all 19 patients recovered from infection. Of note, after discontinuing TNF-α therapy, clinical deterioration was observed in 8(42%) patients, perhaps related to immune reconstitution inflammatory syndrome (IRIS).

It is unclear whether new Histoplasma infection or reactivated infection occurs in patients receiving TNFi [4, 12]. Despite the large number of people in endemic areas exposed previously to H. capsulatum, the incidence is less than expected in patients on TNFi, suggesting development of new infection. In contrast, cases have occurred outside endemic areas, and time to development of histoplasmsosis after administration of TNFis may be brief (<2 weeks), suggesting reactivation.

For histoplamsosis and other opportunistic fungal infections, recommendations support the discontinuation of TNFi in patients with suspected or diagnosed invasive fungal infection.  However, duration of treatment and timing of re-institution of TNFis is unclear.  For patients with histoplasmosis, data on restarting TNFi are available for few cases; but, this has been done safely [4]. Routine screening for histoplasmosis with Histoplasma antigen or antibody testing in patients beginning TNFi is not currently recommended; however, it is important to obtain a thorough exposure and travel history, radiographic studies  and remain aware of symptoms and signs associated with infection [4].

Coccidioidomycosis

TNF-α plays an important role in the control of infection by Coccidioides immits. TNF-α is produced when human or rat cells are exposed to C. immitis spherules or arthroconidia and incubation of human monocytes with recombinant TNF-α results in increased fungicidal activity [34-36]. Thus, TNF-α inhibition may be expected to result in the increased incidence of coccidioidomycosis.

The incidence of coccidiodomycosis in patients receiving TNFi is estimated to be 5.58 per 100,00 person treated with infliximab and 0.88 per 100,000 persons treated with etanercept [7, 24].  To date, at least 30 cases have been described, most with infliximab exposure [2, 37-39]. An early series by Bergstrom and colleagues described 13 cases among patients receiving TNF-α inhibitors from May 1998 through February 2003 in areas endemic for coccidioidomycosis. One case received etanercept and the others received infliximab. The interval between TNFi and infection ranged from 1 to 96 weeks (mean, 27 weeks). All patients had pneumonia on presentation, with 4 (30.7%) having disseminated disease.  Two patients had previous coccidioidomycosis and were thought to have reactivation disease. Five patients were hospitalized and 2 (15.4%) of 13 died. The authors, by reviewing a cohort of inflammatory arthritis at one center, determined the incidence of cocidioidomycosis in patients receiving TNFi to be 1%. Moreover, the risk of infliximab in development of symptomatic coccidioidomycosis when compared to other agents was greater (RR 5.23, 95% CI 1.54-17.71; p<0.01). In a logistic regression model, the use of infliximab was associated with development of symptomatic coccidioidomycosis after adjusting for methotrexate use, prednisone use and age [2].

Aspergillosis

TNF-α is important in host defense against A. fumigatus infections and TNF blockade may lead to increased incidence of infection.  Increased levels of TNF-a are found in mice infected with Aspergillus fumigatus conidiaand TNF-α enhances leukocyte killing of A. fumigatus [25, 40].TNF/lymphotoxin double knockout mice are more susceptible to pulmonary IA, whereas mice exposed to TNF blockers are more susceptible to aspergillosis, resulting in increased fungal burden and mortality [25]. A recent report describes the importance of TNF-R1 polymorphisms and risk of IA [41].

The estimated incidence of aspergillosis is approximately 7 cases/100,000 persons treated [7, 24].  To date, at least 75 cases of aspergillosis have been reported in patients receiving TNFi, mostly with use of infliximab [11, 42-44]. A recent summary of cases was published by Tsiodras and colleagues, who reviewed published reports from January 1966 through June 1, 2007 to determine the association of fungal infections with TNF-α blockade[11]  They identified 64 cases of aspergillosis, mostly invasive pulmonary disease.  The most common TNFi used was infliximab in 48 cases (75%), followed by etanercept in 14 (22%) and adalimumab in 3 cases (3%).  Of 24 cases reviewed with data available, 18 (75%) had received TNFi for GVHD; 4(17%) for RA and 2(8%) for inflammatory bowel disease.  The median number of infusions to diagnosis was 3 (interquartile range, 2-6).  Overall mortality was 82% among patients with GVHD; 3 of 4 cases with RA survived.  Three additional cases were described as part of the prospective French RATIO registry, which identified 45 non-tuberculous opportunistic infections associated with TNF therapy [6]. Two cases were due to A. fumigatus, and sites of infection included sinus, colon and lung.

Candidiasis

TNF-α also plays an important role in host defense against Candida infections [25]. For example, TNF-α is released in response to Candida infection and enhances phagocytosis and killing by polymorphonuclear leukocytes [45].  In addition, when mice with disrupted TNF-α genes are challenged intravenously with C. albicans, survival is decreased when compared to wild type mice [46].

The estimated incidence of Candida infection ranges from 5-20 cases/100,000 persons treated [7, 24].  Over 60 cases have been reported and many were reviewed recently by Tsiodras and colleagues [(A):11].  Of 64 cases identified, 54 (84%) were associated with infliximab and 9 (14%) with etanercept use. The most common underlying disease was GVHD.  Few data on site of infection were available, but esophagitis and bloodstream or endovascular infection were most common.

Cryptococcosis

At least 35 cases of cryptococcosis in patients receiving TNF-α therapy have been published, with an estimated rate of 5-8 cases per 100,000 patients treated [6, (A):11, 24, (A):47-53]. In cases reviewed by Tsiodras and colleagues, most (61%) received infliximab and had underlying RA or inflammatory bowel disease.  Types of infection included pneumonia, meningitis, arthritis, skin disease, or fungemia. Patients typically responded well to antifungal therapy.

Conclusions

The use TNFi is associated with an increased incidence of fungal infections, although these infections appear to be uncommon.  As use of TNFi increases, we can also expect to see an increase in opportunistic fungal infections.  It is important to remain vigilant to allow early diagnosis of these infections.  Several important issues remain, including choice of appropriate screening methods; use of prophylaxis for high-risk patients; length of therapy and use of chronic suppressive therapy. Finally, optimal timing of re-initiation of TNFi has not been defined.  Several studies are underway that will address these important issues.