Author Affiliations: Ronald O. Perelman Department of Dermatology, New York University School of Medicine, New York.
Intravenous immunoglobulin rapidly decreases serum levels of intercellular antibodies in patients with pemphigus vulgaris. However, little is known about the effects of this therapy on antibodies directed specifically against desmoglein 1 and desmoglein 3 and on the IgG subclasses of these antibodies. This study was conducted to study the effect of intravenous immunoglobulin therapy on serum levels of IgG1 and IgG4 antibodies against desmoglein 1 and desmoglein 3 in patients with pemphigus vulgaris.
Within 6 to 16 days after initiating a single cycle of intravenous immunoglobulin therapy in 9 patients, a significant decrease in serum levels of IgG4 and IgG1 antibodies against desmoglein 1 and desmoglein 3 occurred in 60% to 100% of the patients, depending on the antibody subclass and specificity. The median decrease in the antibody levels ranged from 34% to 80%. In addition, most patients (n = 6) showed clinical improvement. The decrease in IgG4 antidesmoglein 3 levels seemed to correlate with improvement in disease activity.
Intravenous immunoglobulin therapy rapidly lowers serum levels of IgG1 and IgG4 antidesmoglein 1 and desmoglein 3 antibodies. There seems to be a stronger association between the decrease in IgG4 antidesmoglein 3 levels and improvement in clinical activity than with changes in the other antibody levels, which suggests that IgG4 antibodies have a more important role in mediating pemphigus vulgaris.
Administration of intravenous IgG (IVIG) is a novel approach to treatment of pemphigus vulgaris (PV). This therapy seems to work by lowering serum levels of pemphigus IgG antibodies.1 However, little is known about the effect of IVIG on the levels of the subclasses of IgG antidesmoglein 1 (Dsg1) and antidesmoglein 3 (Dsg3), the specific antibodies that mediate the disease. This is relevant because disease activity correlates with the level and subclass of these antibodies.2- 4 We conducted a pilot study to determine the effect of IVIG therapy on serum levels of IgG1 and IgG4 antibodies against Dsg1 and Dsg3 in patients with PV.
The study included 9 patients with PV who met clinical, histologic, and immunofluorescence criteria and who received IVIG therapy. Enrollment requirements included active PV with new lesions at initiation of IVIG therapy despite administration of prednisone, 30 mg/d or more, for 2 weeks or longer or 40 mg/d or more for at least 5 days, or relative contraindications to high-dose steroid therapy; or rapidly progressive disease. The study was approved by the New York University Institutional Review Board, New York.
All patients received 1 cycle of therapy with human IVIG, 5% solution. Each cycle consisted of 400 mg/kg/d for 5 days. In all patients, the pre-IVIG dosage of prednisone and other adjuvant therapy was maintained.
Serum levels of IgG1 and IgG4 antibodies against Dsg-1 and Dsg-3 were measured at baseline before administration of IVIG therapy and 6 to 16 days after initiation of therapy using a commercially available enzyme-linked immunosorbent assay kit (Medical and Biological Laboratories Co, Inc, Nagano, Japan) as previously described.5,6 Assays were conducted in duplicate using 10 μL of serum diluted 1:100 and at higher dilutions if necessary to keep the readout within the linear range of the assay. Samples were incubated for 1 hour at 27°C on the assay plate, washed, and then incubated with 100 μL of either mouse antihuman IgG1 or IgG4 conjugated with horseradish peroxidase (Zymed Laboratories, Inc, San Francisco, California) diluted 1:1000. A standard curve, constructed with serial dilutions of a Dsg-positive serum sample, was run with each assay. A computer-generated best-fit log-log curve for each standard was generated and used to calculate the concentration of IgG1 or IgG4 in each serum sample tested. The results were expressed in units, with 1 U defined as the amount of anti-Dsg3 or anti-Dsg1 present in 100 μL of undiluted Dsg-positive standard serum. Desmoglein-negative control serum samples were included in each assay. Serum was considered positive for anti-Dsg antibodies if the antibody level was higher than 0.01 U/mL for IgG4 anti-Dsg3, higher than 0.5 U/mL for IgG4 anti-Dsg1, higher than 0.9 U/mL for IgG1 anti-Dsg3, and higher than 0.5 U/mL for IgG1 anti-Dsg1. These negative cutoff values correspond to an optical density (OD) reading of 0.5-OD unit when assaying for IgG1 antibodies and 0.3-OD unit when assaying for IgG4, as recommended by Hacker et al.5
Patients underwent a complete skin and oral mucous membrane examination at baseline before initiation of IVIG therapy and after completion of treatment. The time between these evaluations ranged from 6 to 16 days. Clinical response was graded as improved or not improved on the basis of the decrease in the number of new or existing lesions or a decrease in prednisone dosage.
The clinical characteristics of 9 patients who received 1 cycle of IVIG therapy are listed in Table 1. All had both mucosal and skin involvement. Six patients were receiving high doses of prednisone, 30 to 240 mg/d, for more than 2 weeks when IVIG therapy was initiated. One patient (patient 7) was given prednisone, 15 mg/d, for more than 2 months, and then prednisone, 40 mg/d, for 5 days before initiation of IVIG therapy. Three patients (patients 2, 3, and 5) were also receiving a combined therapy of azathioprine or cyclophosphamide before initiation of IVIG therapy, and these medications were continued with no change in dosage. In addition, 1 patient (patient 1) was given cyclophosphamide and 1 patient (patient 8) was given azathioprine concurrently with initiation of IVIG therapy.
All patients had anti-Dsg1 or anti-Dsg3 antibodies at baseline (Table 2). Most had both IgG1 and IgG4 antibodies of both specificities. There was no correlation between the level of IgG1 and IgG4 antibodies to the same antigen in the same patient. The assays were reproducible, with similar OD values for the standard curves in assays repeated on 2 different days (observation by M.G.G., data not shown). The intra-assay coefficient of variation, calculated from multiple samples of the same serum tested on the same day, was less than 5, and the interassay variability, calculated from the same serum tested on 2 different days, was less than 4.
Within 6 to 16 days of initiating 1 cycle of IVIG therapy, IgG1 and IgG4 antibodies against both Dsg1 and Dsg3 levels decreased rapidly in most patients who had these antibodies at baseline. A decrease occurred in 60% to 100% of patients with preexisting antibodies, depending on the specificity and subclass of the antibody measured (Table 2). The magnitude of the decrease varied in individual patients (Table 2). On average, anti-Dsg1 IgG1 and IgG4 antibodies decreased by a median of 80% and 34%, respectively, and anti-Dsg3 IgG1 and IgG4 antibodies decreased by a median of 50% and 73%, respectively (Table 2). In a few patients, antibody levels did not change or the levels increased after IVIG therapy.
Six to 16 days after initiation of therapy, PV improved in two-thirds of the patients (Table 3). There was a decrease in the number of new or existing lesions in 5 of 9 patients (56%), and the dose of prednisone was reduced by 50% in a sixth patient.
There seemed to be a correlation between large decreases in IgG4 anti-Dsg3 antibodies and clinical improvement. The clinical response was improved in all 5 patients with a 70% or greater decrease in these antibodies but not in the 3 patients with a smaller decrease in these antibodies. It was difficult to draw an association between the decrease in levels of other classes or specificities of antibodies and clinical improvement.
One patient (patient 6) died of aspergillosis pneumonia 4 days after completing a second cycle of IVIG therapy administered 1 month after the first cycle of IVIG therapy. That patient was receiving a combination therapy of prednisone, 70 mg/d, and cyclophosphamide, 50 mg 3 times a day, when aspergillosis was diagnosed, and immediately previously had been receiving prednisone, 240 mg/d, for 10 days. There were no other adverse events associated with IVIG therapy.
The major finding of the present study is that IVIG therapy for PV causes a rapid decrease in serum levels of both IgG1 and IgG4 anti-Dsg1 and anti-Dsg3 antibodies in most patients. In this study of 9 patients with PV, IVIG therapy usually resulted in a very rapid decrease in both IgG1 and IgG4 antibodies against Dsg1 and Dsg3. Within 6 to 16 days of initiating 1 cycle of IVIG therapy, most patients demonstrated a decrease in both IgG1 and IgG4 anti-Dsg1 and anti-Dsg3 antibodies. The levels of these antibodies decreased by a median of 34% to 80% within this short time, depending on the class and specificity of the antibody. These findings confirm and extend our previous observations1,7 and those of others8,9 that IVIG therapy can cause a rapid decrease in the abnormal antibodies that cause PV. However, these previous studies did not provide information about the effect of IVIG therapy on the specific IgG subclasses directed against Dsg1 and Dsg3.
There seemed to be a correlation between the magnitude of the decrease in IgG4 anti-Dsg3 antibodies and clinical improvement. Improvement occurred in all patients with a greater than 70% decrease in serum levels of these antibodies but not in those with a smaller decrease. This observation must be interpreted cautiously because it is based on a few patients.
There was heterogeneity in the magnitude of the decrease in different IgG subclasses against the same antigen in the same patient. In some patients with PV, IgG1 antibodies decreased more than IgG4 antibodies, whereas the reverse occurred in other patients with PV. We suspect this heterogeneity reflects differences in the way different subclasses of IgG are affected by IVIG. Intravenous immunoglobulin seems to be effective in the treatment of PV by increasing the catabolism of all immunoglobulin molecules, which results in a specific decrease in only pathogenic antibodies, inasmuch as serum levels of normal antibodies are maintained by those present in the IVIG preparation.10 If this explanation is correct, the rate of catabolism of individual classes and subclasses of antibodies depends on their relative concentration in patient serum and in the IVIG preparation. Subclasses in which serum levels are increased to a relatively greater extent, either because they are present to a larger amount in the IVIG preparation or are particularly low in the patient, will be degraded more rapidly, resulting in a more rapid decrease in that IgG subclass.
A practical implication of our findings is that the effectiveness of IVIG preparations may vary depending on the relative level of different IgG subclasses they contain. Selecting preparations with high levels of IgG4 might be desirable because this subclass of pemphigus antibodies seems to be more pathogenic than the others and its levels are most critical to reduce.
Correspondence: Jean-Claude Bystryn, MD, Department of Dermatology, New York University School of Medicine, 550 First Ave, New York, NY 10016 (firstname.lastname@example.org).
Accepted for Publication: March 4, 2008.
Author Contributions: Dr Green had full access to all the data in the study and takes responsibility for the integrity of the data and the accuracy of the data analysis. Ms Goldberg had full access to all the data in the study and takes responsibility for the integrity of the data and the accuracy of the data analysis. Study concept and design: Bystryn and Green. Acquisition of data: Green. Analysis and interpretation of data: Bystryn and Green. Drafting of the manuscript: Green. Critical revision of the manuscript for important intellectual content: Bystryn and Green. Statistical analysis: Green. Obtained funding: Bystryn. Study supervision: Bystryn.
Financial Disclosure: None reported.
Funding/Support: This study was supported by grant 1R01FD-03343-01 from the US Food and Drug Administration.
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