Impact of treatment with infliximab on anticyclic citrullinated peptide antibody and rheumatoid factor in patients with rheumatoid arthritis.
Methods: Sera from 33 RA patients receiving infliximab and disease modifying antirheumatic drugs were tested for anti-CCP antibody, IgA-, lgG- and IgM-RF using a commercially available semi-quantitative ELISA at baseline, 30 and 54 weeks after treatment.
Results: The serum levels of anti-CCP antibody and IgA-RF decreased significantly after 30 weeks (P = 0.002 and 0.024); however, the decrease was not significant at week 54 (P = 0.147 and 0.207). The decrease in IgG-RF level was not significant at 30 and 54 weeks (P = 0.059 and 0.097). IgM-RF levels, however decreased significantly at 30 and 54 weeks (P = 0.002 and 0.004). A strong correlation between anti-CCP and IgA-, IgG- and IgM-RF was observed at baseline ([r.sub.s] = 0.48, 0.43, 0.65, P = <0.05) and after infliximab treatment at 30 ([r.sub.s] = 0.45, 0.46, 0.62, P = <0.05) and 54([r.sub.s] = 0.49, 0.45, 0.60, P = <0.05) weeks.
Conclusion: Treatment with infliximab results in decreased anti-CCP antibody and IgA-RF early in the course of therapy that is not sustained. IgM-RF declines and remains decreased for at least 54 weeks. Investigations in larger cohorts of RA patients (especially early RA) with longer follow-up are needed to assess the impact of specific therapeutic interventions on anti-CCP antibody and RF levels and the relationship of their levels to disease activity.
Key Words: Rheumatoid arthritis, rheumatoid factor, anticyclic citrullinated peptide antibody, TNF-[alpha] inhibitors, infliximab
Rheumatoid arthritis (RA) is one of the most common systemic autoimmune diseases affecting approximately 1% of the world population. (1) It is characterized by synovial inflammation and subsequent pannus formation causing cartilage destruction and bone erosion, severe disability and increased mortality. The diagnosis of RA is based primarily on the clinical manifestations of the disease. Rheumatoid factor (RF) and more recently, anticyclic citrullinated peptide (anti-CCP) antibodies have been suggested to be useful diagnostic tools, especially in the early stages of the disease and may be detected in approximately half of the patients with RA before the onset of clinical disease. (2-5) In addition, anti-CCP antibodies are also detected in 34.5% of all patients with RA when all measured RF isotypes (IgG-RF, IgA-RF, and IgM-RF) were negative. (6) The presence of these antibodies may also be predictive of disease progression and radiological damage. (7-9) In particular, anti-CCP antibodies appear to be highly sensitive (approximately 80%) and specific (98%) for rheumatoid arthritis. (10-12) Although there is a considerable overlap between the presence of RF and anti-CCP antibodies, there is evidence that they belong to two separate autoantibody systems. Anti-CCP antibodies are associated with the shared epitope, whereas RF is not strongly associated with the presence of the shared epitope. (13-16) Moreover, extra-articular manifestations of RA (including subcutaneous nodules, vasculitis, leg ulcers, and neuropathy) are related to the presence of high titers of RF but not anti-CCP antibodies. (13,14) Several studies have detected that the level of IgM-RF decreases with the administration of effective disease-modifying therapies, especially methotrexate and parenteral gold, suggesting a link with disease activity. (17,18) In contrast, few studies have analyzed the variations in the titer of anti-CCP antibodies and RF during treatment with biologic agents such as TNF-[alpha] inhibitors. (19-22) The presence of citrullinated proteins in the inflamed joints and evidence of local production of anti-CCP antibodies suggests a pathogenic role. Given the proposed role of anti-CCP antibody in both the pathogenesis and the early diagnosis of RA, (2,3,23,24) it is important to study the relationship of their levels with therapeutic interventions and the resulting disease activity. The aim of this study was to analyze the effect of infliximab treatment on anti-CCP antibody, RF and acute phase reactant levels in patients with RA and to correlate these changes with clinical responses.
Patients and Samples
Sera from 33 patients (28 females and 5 males, 18 whites, 14 blacks and one Hispanic, median age 57 yr, mean age 54.6 [+ or -] 12.1 yr, range 25-74 yr, mean disease duration 12.9 [+ or -] 8.3 year) meeting the American College of Rheumatology (ACR) (25) criteria for RA, who required therapy with multiple disease-modifying antirheumatic drugs (DMARDS) and infliximab, were selected for testing for anti-CCP antibody, IgA-, IgG- and IgM-RF. Sera from every RA patient treated with infliximab had been collected and stored at -70[degrees]C before each infliximab infusion until the time of the assays. We tested the baseline sera before initiation of infliximab treatment and at 30 weeks in 33 patients and at 54 weeks in 28 patients. Sera at 54 weeks on 5 patients were not available. In addition to infliximab, all patients received other DMARDS, including methotrexate, sulfasalazine, lefluno-mide, azathioprine, hydroxychloroquine, gold and nonsteroidal anti-inflammatory drugs (Table 1). Infliximab was administered initially at 3 mg/kg at baseline, then at 2 weeks, 6 weeks and subsequently every 8 weeks. Some patients required a dose adjustment between 3 and 10 mg/kg based on their clinical response and some required more frequent dosing at every six weeks. We also tested 44 patients with other rheumatic diseases (18 with SLE, 15 with scleroderma, 7 with polymyositis, and 4 with psoriatic arthritis) for anti-CCP antibody, IgA-, IgG-, and IgM-RF.
Clinical and Laboratory Assessments
All patients had history and physical examinations before each infliximab infusion. A complete blood count, electrolytes, blood urea nitrogen, creatinine, aspartate aminotransferase, alanine aminotransferase, albumin, erythrocyte sedimentation rate (ESR) and C-reactive protein (CRP) were measured before each infliximab treatment. The clinical and laboratory results were evaluated at baseline and at 30 and 54 weeks of infliximab treatment.
Anti-CCP and RF
Anti-CCP antibodies were detected using a commercially available second generation, semiquantitative enzyme-linked immunosorbent assay (ELISA) kit (QUANTA Lite CCP IgG ELISA of INOVA Diagnostics Inc., San Diego, CA, USA), following the manufacturer's instructions. The microtitre plates, precoated with highly purified synthetic cyclic citrullinated peptides (CCP) containing modified arginine residues, were incubated for 30 minutes at room temperature with patients' serum samples diluted at 1:101 by adding 5 [micro]L of sample to 500 [micro]L of horseradish peroxidase (HRP) sample diluent. Prediluted anti-CCP standards and positive and negative controls were added to each plate. After three washes with HRP wash concentrate (containing Tris-buffered saline and Tween 20), the plates were incubated for 30 minutes at room temperature with an HRP enzyme labeled antihuman IgG conjugate. After three more washes with HRP wash concentrate, the enzyme reaction was developed by adding tetramethylbenzene chromogen and incubating in the dark for 30 minutes at room temperature. The reaction was terminated with HRP stop solution (0.344M sulfuric acid). The absorbance of each well was read at 450 nm within one hour of adding the HRP stop solution. Anti-CCP antibody was considered positive when the absorbance value was higher than the cut off of the kit (<20 U/mL). The concentration of anti-CCP antibody was calculated from a dose-response curve based on standards; all sera were tested in triplicate and the results reflect the mean. IgA-, IgG- and IgM-RF were also detected by using semiquantitative ELISA kits (QUANTA Lite RF IgA, IgG and IgM ELISA of INOVA Diagnostics, Inc., CA, USA) using the same technique. RF was considered positive when the absorbance value was higher than the cut off of the kit (>6 U) (Fig. 1).
We analyzed our data using SAS 9.0 statistical software (SAS, Cary, NC) and performed Repeated Measures ANOVA Analysis to investigate the effect of infliximab on ESR, CRP, and anti-CCP antibodies, IgA-, IgG- and IgM-RF. Wilcoxon signed rank test was used for paired analysis. Correlations were sought using Spearman correlation coefficients ([r.sub.s]). A probability (p) value of <0.05 was considered significant.
At baseline, 27 of the 33 patients (81.8%) with RA were positive for anti-CCP antibodies, 28 (84.8%) for IgA-RF, 27 (81.8%) for IgG-RF and 29 (87.8%) for IgM-RF. In 25 cases, (75.7%) all 4 antibodies were detected. A strong correlation between anti-CCP and IgA-, IgG- and IgM-RF was observed at baseline ([r.sub.s] = 0.48, 0.43, 0.65, P = <0.05) and after infliximab treatment at 30 ([r.sub.s] = 0.45, 0.46, 0.62, P = <0.05) and 54 ([r.sub.s] = 0.49, 0.45, 0.60, P = <0.05) weeks. No clinical differences at baseline were observed among the patients according to whether they were positive or negative for anti-CCP or RF. The proportion of patients who were positive for anti-CCP and RF antibodies was similar at baseline and at weeks 30 and 54; anti-CCP antibodies (81.8% versus 78.7% versus 78.5%), IgA-RF (84.8% versus 84.8% versus 85.7%), IgG-RF (81.8% versus 84.8% versus 85.7%) and IgM-RF (87.8% versus 87.8% versus 89.2%) (Table 2). After infliximab treatment, one positive patient with anti-CCP antibody became negative at 30 weeks and remained negative at 54 weeks. No patient with a positive IgA-, IgG- or IgM-RF became negative during the study period. One patient who was negative for anti-CCP antibody and IgM-RF at 0 and 30 weeks became weakly positive for both at 54 weeks. One patient who was negative for IgA-RF before treatment became weakly positive at 54 weeks. One patient with negative IgG-RF at baseline and 30 weeks became positive at 54 weeks.
The serum levels of anti-CCP antibody and IgA-RF decreased significantly after 30 weeks (P = 0.002 and 0.024); however, the decrease was not sustained at week 54 (P = 0.147 and 0.207). The decrease in IgG-RF titers was not significant at 30 and 54 weeks (P = 0.059 and 0.097). IgM-RF titers, however, decreased significantly at 30 and 54 weeks (P = 0.002 and 0.004 respectively) (Table 3). Although a decrease in both ESR and CRP was observed after treatment, the differences were not statistically significant. The variations in ESR and CRP correlated with variations in tender and swollen joint counts ([r.sub.s] = 0.49 and 0.48, P = <0.05). At baseline, only IgM-RF correlated with swollen joint count; however, we did not find any correlation between variations in IgA-RF, IgG-RF, IgM-RF, anti-CCP antibody titers and variations in clinical and laboratory indices in response to infliximab treatment. Among 44 patients with other rheumatic diseases, 7 (15.9%) were positive for anti-CCP antibodies, (2 with SLE, 4 with scleroderma and I with polymyositis). Four patients (9%) were noted to be strongly positive (1 with SLE, 2 with scleroderma and I with polymyositis). Forty one of the 44 patients were positive for IgA-RF, 34 for IgG-RF and 30 for IgM-RF (Table 4).
Circulating antibodies are the main effector molecules of the humoral immune system and at present, represent the only practical indices for monitoring the immune response in certain autoimmune diseases. Pathogenic autoantibodies have been well documented in a number of different autoimmune diseases including autoimmune thrombocytopenia, myasthenia gravis, and systemic lupus erythematosus. Until recently, the only serologic test routinely used to confirm the diagnosis of RA was the determination of the presence of IgM-RF in the serum. RFs are antibodies directed against the constant region of immunoglobulins of the IgG class. RF can be of the IgM, IgG, IgA and IgE immunoglobulin classes. The presence of IgM-RF is a hallmark of RA, found in approximately 50 to 90% of these patients. (26,27) However, IgM-RF is nonspecific and is found in infections, malignancies, other auto-immune diseases, and some healthy individuals. (26,27) IgG-RF is less sensitive but more specific for RA than IgM-RF. (27)
It has been known for many years that antiperinuclear autoantibodies, also called antikeratin, are found in people with RA. (28,29) These antibodies recognize an epitope that contains the deaminated form of arginine called citrulline. (30) A synthetic circular peptide containing citrulline called CCP was found to be better at discriminating RA patients from other arthritis patients than either the perinuclear autoantibody test or the RF assays. (4,7,31-33) Schellekens et al, (5) Bas et al, (14) and Pinheiro et al (34) reported that anti-CCP antibodies may be present in over 70% of RA patients but in fewer than 5% of healthy individuals. Anti-CCP antibodies have been found in early stages of the disease, with a significantly greater prevalence in the sera of patients who subsequently develop more severe radiological damage. (2-9) This suggests that posttranslational modifications of arginine residues (citrullination of proteins) by the enzyme peptidyl arginine deaminase (PAD) are important early processes in the progression to disease. Increased expression of PAD isoenzyme 4 (PADI4) by neutrophils was seen in the presence of inflammation, and the expression appeared to correlate with the degree of inflammation in the murine models of RA. (35) A case-control association study has shown that polymorphisms of the gene that encode the PADI4 is associated with RA in Japanese populations. (36) A recent study found citrullinated proteins in the joints of patients with RA, leading to speculation that these proteins may trigger specific autoimmune responses in RA. (37) However, in another study, immunohistochemical staining of citrullinated proteins was observed in the lining layer, the sublining layer, and in extravascular fibrin deposits in inflamed synovial tissue from RA as well as non-RA patients. Thus, the presence of citrullinated proteins in the inflamed synovium may be related to inflammation rather than the specific disease process of RA. It is speculated that a low level of inflammation, secondary to trauma or infection that causes death of PAD-expressing cells, leads to activation of the PAD enzyme and citrullination of proteins in the joints. While this process is nonspecific, the presence of specific anti-CCP antibodies in RA patients is probably the result of an abnormal but disease-specific humoral response to citrullinated proteins. It appears that genetic factors such as HLA-DR4 and PAD gene polymorphisms are involved in the process that determines whether anticitrullinated protein antibodies are made. (38,39) Anti-CCP antibody production amplifies this low level of inflammation and may contribute to perpetuation of joint inflammation and thereby to chronicity and severity of RA. Reparon-Schuijt et al (40) identified anti-CCP antibody producing B cells within inflamed joints of patients with RA and Vossenaar et al (38) found that anti-CCP antibodies were enriched in synovial fluid compared with serum, indicating local production. As effector cells of the humoral immune response, B cells may contribute to both the initiation and perpetuation of the immune response that leads to chronic synovitis. Animal studies involving K/BxN transgenic mice have shown that antigen-specific antibodies (an-tiglucose-6-phosphate isomerase [GPI] antibody) may initiate inflammatory arthritis, and may link the adaptive and innate immune systems. The arthritogenic antibodies from this transgenic model induced the same pattern of arthritis when transferred to healthy mice. (41) The antigen-antibody complexes may stimulate the neutrophils, and mast cells through complement- and Fc gamma receptor-dependent pathways, ultimately resulting in arthritis. (42) It is also interesting that both IL-1 and TNF-[alpha] are prominently involved in the K/BxN model of arthritis; the cytokines centrally implicated in the pathogenesis of RA. (43) These results suggest a possible pathogenic role for autoantibodies which may be directly involved in the tissue damage in RA. Given the proposed role of anti-CCP antibodies in both the pathogenesis and the early diagnosis of RA, it is important to study the relationship of their levels and various therapeutic interventions. (2,3,23,40)
Only a few studies have analyzed the variations in the titer of anti-CCP antibodies and RF during treatment with TNF-[alpha] inhibitors with inconsistent results. (19-22) Caramaschi et al (19) measured the levels of IgM-RF, anti-CCP antibodies and CRP in 27 RA patients after 22 weeks of treatment with infliximab. They found that RF level and positivity, as well as CRP decreased significantly; however, the levels of anti-CCP antibodies remained stable. De Rycke et al (20) analyzed the effect of methotrexate and infliximab treatment on IgM-RF and anti-CCP antibodies in 62 RA patients. At baseline and at week 30, IgM-RF titers were reduced significantly during infliximab treatment (P < 0.001 and P = 0.038, respectively), whereas anti-CCP antibodies remained unchanged (P = 0.240). However, they described a subset of patients (23/38) who experienced a marked (>20%) decline in the anti-CCP antibody level. Alessandri et al (21) observed significant declines in the levels of both RF and anti-CCP antibodies in patients who had clinical improvement after six months of treatment with infliximab. They suggested that modification of serum anti-CCP appears to parallel the improvement in clinical and laboratory variables observed during treatment with infliximab. In another study, Bobbio-Pallavicini et al (22) tested serum samples from 30 RA patients treated with infliximab and methotrexate at baseline and after 30, 54 and 78 weeks. The median IgM-RF titer declined progressively and significantly from baseline to 30, 54 and 78 weeks. In contrast, the anti-CCP antibody titer decreased significantly at 30 weeks but returned to baseline thereafter.
In the present study, we detected a significant decrease in the titer of anti-CCP antibodies, IgA-RF and IgM-RF in the sera of patients after 30 weeks of treatment with infliximab. This trend continued at 54 weeks, but while the decrease remained significant for IgM-RF titers, it was not statistically significant for anti-CCP antibodies and IgA-RF. This is the first study analyzing the effect of infliximab treatment on anti-CCP antibodies and all the isotypes of RF. The relatively high prevalence of patients who were positive for anti-CCP and IgA-, IgG- and IgM-RF reflects the probable selection of patients with more aggressive disease, and were candidates for anti-TNF-[alpha] therapy, indicating an association between these autoantibodies and a more severe disease course.
Two recent studies analyzed the variation in anti-CCP antibody titers in response to treatments other than infliximab. (24-44) Cambridge et al (24) reported a significant decrease in anti-CCP antibodies, IgG-, IgA- and IgM-class RF and CRP in RA patients who responded to B-lymphocyte depletion therapy obtained by rituximab administration with or without cyclophosphamide and/or prednisone. The fall of anti-CCP antibody levels was significantly greater than that of total serum IgG. The decrease was observed rapidly after treatment; the median time taken for serum levels of anti-CCP antibodies to fall by 50% of pretreatment levels occurred within 5 weeks. Nonresponding patients did not present any significant variation in either anti-CCP antibodies or RF levels. The B-lymphocyte count took as long as 21 months to reach baseline. The time for return of synovitis, after B-lymphocyte return was often long and unpredictable (range 0-17 mo); however, was closely correlated with rises in the level of at least one autoantibody. Increased autoantibody levels were rarely observed in the absence of clinical change. In another study, Mikuls et al (44) reported a decline in the serologic levels of circulating autoantibodies among RA patients, the majority of whom received active therapy with DMARDs. Although autoantibody levels only rarely normalized with treatment, one-half of patients experienced a 25% or greater reduction and one-third had at least a 50% reduction in the level of anti-CCP antibody and RF. Although there was no association of treatment response with changes in anti-CCP levels, RA patients with disease duration of <1 year were substantially more likely than those with more established disease to experience a significant decrease in the anti-CCP antibody level. Conversely, treatment response (not disease duration) was the primary determinant of treatment-related declines in the RF level.
The majority of the patients in our study and the other studies examining the effect of infliximab treatment on anti-CCP antibody level had chronic severe RA. The success of clinical interventions in lowering anti-CCP antibody levels appears to depend on the patient's response to therapy and the disease duration. (21,24,44) Lower levels of anti-CCP antibodies at baseline also seemed to predict a better clinical response. (21) The association of shorter disease duration with greater declines in anti-CCP antibody levels is highly consistent with the growing body of evidence showing improved clinical outcomes with earlier disease intervention in RA. (45,46) It is possible that the early disease period represents an important therapeutic window in RA.
This and other studies suggest that in addition to being an important diagnostic tool, serial evaluation of these antibodies could be useful in monitoring the clinical course of the disease. It is unknown whether decrements in the levels of circulating autoantibodies translate directly into improvements in long-term outcomes in RA. It is possible that anti-CCP antibody level declines may predict improvement only over longer periods of follow-up or improvements in radiographic or other outcome measures including morbidity and mortality. Investigations in larger cohorts of RA patients (especially early RA) with longer follow-up are needed to assess the impact of specific therapeutic interventions on anti-CCP antibody and RF levels and the relationship to disease activity.
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M. Mubashir Ahmed, MD, Eisha Mubashir, MD, Robert E. Wolf, MD, Samina Hayat, MD, Vicky Hall, PhD, Runhua Shi, MD, PhD, and Seth Mark Berney, MD
From the Center of Excellence for Arthritis and Rheumatology and the Section of Rheumatology, the Division of Hematology/Oncology, the Department of Medicine, and Overton Brooks Veterans Administration Hospital, Louisiana State University Health Sciences Center, Shreveport, LA.
Reprint requests to Mohammed Mubashir Ahmed, MD, Assistant Professor of Medicine, Center of Excellence for Arthritis and Rheumatology, LSU-HSC-S, 1501 Kings Highway, Shreveport, LA 71130. Email email@example.com
Accepted June 14, 2006.
RELATED ARTICLE: Key Points
* Treatment with infliximab results in decreased anti-cyclic citrullinated peptide (anti-CCP) antibodies and IgA-RF early in the course of therapy that is not sustained.
* The decrease in IgG-RF level was not significant at 30 and 54 weeks.
* IgM-RF declines and remains decreased for at least 54 weeks.
* Investigations in larger cohorts of rheumatoid arthritis (RA) patients (especially early RA) with longer follow-up are needed to assess the impact of specific therapeutic interventions on anti-CCP antibody and RF levels and the relationship of their levels to disease activity.
Table 1. Demographic and baseline clinical characteristics of 33 patients with rheumatoid arthritis Characteristic Value Age 54.6 [+ or -] 12.1 Female/male 28/5 Caucasian 18 African American 14 Hispanic 1 Disease duration (years) 12.9 [+ or -] 8.3 Number of tender joints 18.06 [+ or -] 11.38 Number of swollen joints 18.42 [+ or -] 7.68 ESR 37.34 [+ or -] 25.26 CRP (mg/dL) 2.38 [+ or -] 1.76 DMARDs Previous Current Methotrexate 32 27 Hydroxychloroquine 7 1 Sulfasalazine 15 14 Leflunomide 23 15 Gold 12 2 Azathioprine 5 2 Minocycline 1 0 Cyclosporin A 1 0 Etanercept 3 0 Where applicable, values are expressed as mean [+ or -] SD. Anti-CCP. anticyclic citrullinated peptide antibody; RF. rheumatoid factor; ESR, erythrocyte sedimentation rate; CRP, C-reactive protein; DMARDs, disease modifying antirheumatic drugs. Table 2. Percentages of rheumatoid arthritis patients with autoantibodies at baseline and after treatment. At baseline Week 30 Week 54 Autoantibody (n = 33) (n = 33) (n = 28) Anti-CCP antibody positivity 81.8% 78.7% 78.5% IgA-RF positivity 84.8% 84.8% 85.7% IgG-RF positivity 81.8% 84.8% 85.7% IgM-RF positivity 87.8% 87.8% 89.2% anti-CCP, anticyclic citrullinated peptide antibody; RF, rheumatoid factor. Table 3. Changes in clinical characteristics of patients with rheumatoid arthritis from baseline and at different times after infliximab treatment Variable Week 0 Week 30 Week 54 Tender joint count 18.06 (11.38) 8.31 (9.92) 9.89 (9.78) Swollen joint count 18.42 (7.68) 10.63 (6.47) 10.57 (7.2) ESR (mm/h) 37.34 (25.26) 34.88 (27.42) 34.75 (25.55) CRP (mg/dL) 2.38 (1.76) 1.81 (1.52) 1.78 (0.96) Anti-CCP (IU) 172.23 (119.63) 163.38 (113.99) 153.77 (121.62) IgA-RF (IU) 60.97 (49.38) 52.82 (47.89) 53.2 (54.26) IgG-RF (IU) 54.24 (42.14) 49.15 (41.01) 44.46 (39.83) IgM-RF (IU) 85.27 (43.65) 76.79 (43.16) 70.32 (43.69) P Value Variable Week 0 vs. 30 Week 0 vs. 54 Tender joint count 0.001* 0.003* Swollen joint count 0.001* 0.001* ESR (mm/h) 0.129 0.475 CRP (mg/dL) 0.265 0.222 Anti-CCP (IU) 0.002* 0.147 IgA-RF (IU) 0.024* 0.207 IgG-RF (IU) 0.059 0.097 IgM-RF (IU) 0.002* 0.004* Values are mean (SD). *P < 0.05 ESR, erythrocyte sedimentation rate: CRP, C-reactive protein; anti-CCP, anticyclic citrullinated peptide antibody; RF, rheumatoid factor. Table 4. Anticyclic citrullinated peptide antibody, IgA-, IgG- and IgM-RF in 44 patients with other rheumatic diseases. Disease No. anti-CCP IgA-RF IgG-RF IgM-RF Psoriatic arthritis 4 0 (0%) 3 (75%) 4 (100%) 2 (50%) SLE 18 2 (11%) 17 (94%) 9 (50%) 7 (39%) Scleroderma 15 4 (27%) 15 (100%) 14 (93%) 14 (93%) Polymyositis 7 1 (14%) 6 (86%) 7 (100%) 6 (86%) No., number of patients; anti-CCP, anticyclic citrullinated peptide antibody; RF, rheumatoid factor; SLE, systemic lupus erythematosus.