Correspondence
Treatment of Rheumatoid Arthritis with a Tumor Necrosis Factor Receptor–Fc Fusion Protein
N Engl J Med 1997; 337:1559-1561November 20, 1997
- Article
To the Editor:
Moreland et al. (July 17 issue)1 report that the administration of a recombinant human tumor necrosis factor receptor (p75)–Fc fusion protein (TNFR:Fc) to adults with refractory rheumatoid arthritis results in significantly more improvement in the inflammatory symptoms than placebo injections. This conclusion is highly dependent on the blinding of the clinical trial. The results of open and double-blind trials of anti-CD4 therapy make clear that the outcome variables used by Moreland et al.1 are highly susceptible to expectation bias.2 Although the design of the study included initial blinding, the question is whether such blinding was maintained for the three months of the study after 24 subcutaneous injections. Local reactions at the injection site and upper respiratory tract symptoms developed only in those receiving the two higher concentrations of the drug; no such reaction is mentioned for the placebo group. This raises the possibility that both the patients and the investigators could have become aware of the treatment assignments long before the 24th injection. Just as the benefit of this agent is said to be dose related, the frequency of the upper respiratory tract side effects is stated to be dose related.
The fact that unblinding can result in an amount of improvement similar to that attributed to the drug in this study is illustrated in prior studies. Using these same primary outcome measures, Moreland et al.3 reported that 43 percent of patients given anti-CD4 therapy in an open study had a decrease of 50 percent or more in the tender-and-swollen-joints count, but they found no significant efficacy as compared with placebo in the subsequent blinded study.4 In the present study, no assessment of residual blinding was made at the conclusion of the three months of treatment. Determining whether there is residual blinding is surprisingly simple. Gotzsche et al.,5 in a recent study of slow-acting antirheumatic drugs, questioned both subjects and investigators at the conclusion of the trial and provided evidence substantiating their belief that it was, indeed, a blinded study.
Given the very large number of antirheumatic biologic agents now in the therapeutic testing pipeline and the clear effect of unblinding when these outcome measures are used, it should be mandatory to assess whether there is residual blinding at the time of the last clinical measurement.
5 ReferencesWallace V. Epstein, M.D.
University of California, San Francisco, San Francisco, CA 94143-09201
Moreland LW ,Baumgartner SW ,Schiff MH , et al. Treatment of rheumatoid arthritis with a recombinant human tumor necrosis factor receptor (p75)-Fc fusion protein. N Engl J Med 1997;337:141-147
Full Text | Web of Science | Medline2
Epstein WV . Expectation bias in rheumatoid arthritis clinical trials: the anti-CD4 monoclonal antibody experience. Arthritis Rheum 1996;39:1773-1780
CrossRef | Web of Science | Medline3
Moreland LW ,Bucy RP ,Tilden A , et al. Use of a chimeric monoclonal anti-CD4 antibody in patients with refractory rheumatoid arthritis. Arthritis Rheum 1993;36:307-318
CrossRef | Web of Science | Medline4
Moreland LW ,Pratt PW ,Mayes MD , et al. Double-blind, placebo-controlled multicenter trial using chimeric monoclonal anti-CD4 antibody, cM-T412, in rheumatoid arthritis patients receiving concomitant methotrexate. Arthritis Rheum 1995;38:1581-1588
CrossRef | Web of Science | Medline5
Gotzsche PC ,Hansen M ,Stoltenberg M , et al. Randomized, placebo controlled trial of withdrawal of slow-acting antirheumatic drugs and of observer bias in rheumatoid arthritis. Scand J Rheumatol 1996;25:194-199
CrossRef | Web of Science | Medline
To the Editor:
Moreland et al. reported on the use of TNFR:Fc to treat patients with rheumatoid arthritis. The results were encouraging, but the patients had local reactions at the injection site. To explore the possibility that patients had become immunized to TNFR:Fc, tests were performed to determine whether antibodies to TNFR:Fc had developed. Although the results of those tests were reported to be negative, the assay had some technical limitations.
The authors used an enzyme immunoassay in which wells were coated with TNFR:Fc at a concentration of 63 ng per milliliter. This concentration of TNFR:Fc is probably suboptimal for the detection of antibodies. In later steps of the assay, the binding of patients' antibodies to the TNFR:Fc was detected with enzyme-conjugated goat antibodies to the F(ab')2 of human immunoglobulin. The sensitivity and specificity of this reagent for different immunoglobulin classes were not stated, but it is likely to detect IgG, IgM, and IgA. Many of the patients probably had serum rheumatoid factors (i.e., autoantibodies to the Fc part of IgG). Unless the Fc conformation in TNFR:Fc was substantially altered, rheumatoid factors would bind to the TNFR:Fc used to coat the wells. The authors scored the specimens as positive when optical-density values were at least four times as high as those of the corresponding pretreatment samples. The presence of rheumatoid factor or a decrease in the level of rheumatoid factor would mask the presence of antibodies to TNFR:Fc. Thus, the suboptimal coating of wells with TNFR:Fc and the presence of rheumatoid factors render the enzyme immunoassay potentially insensitive for the detection of antibodies to TNFR:Fc.
Normal IgG molecules have a functional antigenic valence of 1 for interaction with rheumatoid factors, since the second binding site on the Fc domain is inaccessible owing to the flexibility of the hinge region.1,2 In contrast, free Fc fragments have a valence of 2,1 a finding confirmed by crystallography.3 The valence of the TNFR:Fc for interaction with rheumatoid factors has not been described, but it would depend on the flexibility of the fusion protein.
Interaction of rheumatoid factors or other antibodies with the TNFR:Fc molecules could contribute to local reactions at the injection sites, and the clearance of the drug could also be influenced by these antibodies. Interaction of rheumatoid factor with this and other Fc fusion molecules could be clinically important when fusion proteins are used to treat patients with rheumatoid arthritis.
3 ReferencesMart Mannik, M.D.
Mark Wener, M.D.
University of Washington, Seattle, WA 98195-64281
Nardella FA ,Teller DC ,Mannik M . Studies on the antigenic determinants in the self-association of IgG rheumatoid factor. J Exp Med 1981;154:112-125
CrossRef | Web of Science | Medline2
Stone MJ ,Metzger H . Bonding properties of a Waldenström macroglobulin antibody. J Biol Chem 1968;243:5977-5984
Web of Science | Medline3
Sohi MK ,Corper AL ,Wan T , et al. Crystallization of a complex between the Fab fragment of a human immunoglobulin M (IgM) rheumatoid factor (RF-AN) and the Fc fragment of human IgG4. Immunology 1996;88:636-641
CrossRef | Web of Science | Medline
Author/Editor Response
The authors reply:
To the Editor: We agree with Dr. Epstein that blinding is critical in evaluating new therapies. Efficacy was demonstrated in this study not only by clinical responses but also by objective laboratory measures of disease activity (the erythrocyte sedimentation rate, C-reactive protein level, platelet count, and hemoglobin level) that are not susceptible to expectation bias. These objective measures were not included in the anti-CD4 trials.1,2 Two events (upper respiratory tract symptoms and injection-site reactions) were more frequent in TNFR:Fc-treatment groups and potentially could have “unblinded” the trial. However, several observations suggest that this possibility is unlikely: upper respiratory tract symptoms and injection-site reactions were also reported in the placebo group, no strict dose–response relation was evident with respect to the upper respiratory tract symptoms, and although injection-site reactions did appear to be dose-related, improvements in disease activity were of similar magnitude in patients with and those without injection-site reactions. To address these issues directly, separate blinded joint assessors not involved in the clinical care of patients were included in a subsequent phase 3 study, which replicated the phase 2 results.
The potential technical limitations of the antibody enzyme-linked immunosorbent assay (ELISA) for TNFR:Fc addressed by Drs. Mannik and Wener were considered during the development of the assay. The coating concentration of TNFR:Fc was selected empirically as the concentration that generated the highest optical-density signals in heterologous immune serum, while simultaneously exhibiting low background optical densities. The antibody ELISA successfully detects antibodies in the serum of TNFR:Fc-treated monkeys and therefore should be capable of detecting similar antibodies in the samples from patients.
Rheumatoid-factor–positive samples from TNFR:Fc-treated patients have been tested untreated and after absorption with insolubilized aggregated IgG before assay. The optical densities were slightly reduced by absorption (0.050 optical-density unit). However, the relative optical-density ratios of serum samples obtained before and after TNFR:Fc treatment were unchanged. In addition, an IgG fraction of pooled seropositive monkey serum specific for tumor necrosis factor receptor was affinity-purified and mixed with serum obtained from a patient with rheumatoid arthritis. These monkey antibodies were readily detected by the ELISA, even in the presence of the serum from the patient with rheumatoid arthritis. Finally, samples from trials of patients with other diseases, in which rheumatoid factor is not present, have also been negative when tested. These observations suggest that TNFR:Fc is not frequently immunogenic in humans.
2 ReferencesLarry W. Moreland, M.D.
University of Alabama at Birmingham, Birmingham, AL 35294-7201Michael B. Widmer, Ph.D.
Consuelo M. Blosch, M.D.
Immunex Corporation, Seattle, WA 981011
Moreland LW ,Bucy RP ,Tilden A , et al. Use of a chimeric monoclonal anti-CD4 antibody in patients with refractory rheumatoid arthritis. Arthritis Rheum 1993;36:307-318
CrossRef | Web of Science | Medline2
Moreland LW ,Pratt PW ,Mayes MD , et al. Double-blind, placebo-controlled multicenter trial using chimeric monoclonal anti-CD4 antibody, cM-T412, in rheumatoid arthritis patients receiving concomitant methotrexate. Arthritis Rheum 1995;38:1581-1588
CrossRef | Web of Science | Medline
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H. E. Seymour, A. Worsley, J. M. Smith, S. H. L. Thomas. (2001) Anti-TNF agents for rheumatoid arthritis. British Journal of Clinical Pharmacology 51:3, 201-208
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A LATRONICO, D SEGALOFF. (1999) Naturally Occurring Mutations of the Luteinizing-Hormone Receptor: Lessons Learned about Reproductive Physiology and G Protein–Coupled Receptors. The American Journal of Human Genetics 65:4, 949-958
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