More prominent muscle involvement in patients with dermatomyositis with anti-Mi2 autoantibodies

Abstract

Objective

To define the clinical phenotype of dermatomyositis (DM) with anti-Mi2 autoantibodies.

Methods

In this longitudinal cohort study, the prevalence and severity of clinical features at disease onset and during follow-up in patients with anti-Mi2–positive DM were compared to patients with anti-Mi2–negative DM, antisynthetase syndrome (AS), and immune-mediated necrotizing myopathy (IMNM). Longitudinal anti-Mi2 autoantibody titers were assessed.

Results

A total of 58 patients with anti-Mi2–positive DM, 143 patients with anti-Mi2–negative DM, 162 patients with AS, and 170 patients with IMNM were included. Among patients with anti-Mi2–positive DM, muscle weakness was present in 60% at disease onset and occurred in 98% during longitudinal follow-up; fewer patients with anti-Mi2–negative DM developed weakness (85%; p = 0.008). Patients with anti-Mi2–positive DM were weaker and had higher creatine kinase (CK) levels than patients with anti-Mi2–negative DM or patients with AS. Muscle biopsies from patients with anti-Mi2–positive DM had prominent necrosis. Anti-Mi2 autoantibody levels correlated with CK levels and strength (p < 0.001). With treatment, most patients with anti-Mi2–positive DM had improved strength and CK levels; among 10 with multiple serum samples collected over 4 or more years, anti-Mi2 autoantibody titers declined in all and normalized in 3, 2 of whom stopped immunosuppressant treatment and never relapsed. Patients with anti-Mi2–positive DM had less calcinosis (9% vs 28%; p = 0.003), interstitial lung disease (5% vs 16%; p = 0.04), and fever (7% vs 21%; p = 0.02) than did patients with anti-Mi2–negative DM.

Conclusions

Patients with anti-Mi2–positive DM have more severe muscle disease than patients with anti-Mi2–negative DM or patients with AS. Anti-Mi2 autoantibody levels correlate with disease severity and may normalize in patients who enter remission.

The idiopathic inflammatory myopathies (IIM) are a heterogeneous family of diseases that affect skeletal muscle and, in some cases, the skin, lungs, or joints. At least 4 well-defined types of IIM are now widely recognized, including dermatomyositis (DM), the antisynthetase syndrome (AS), immune-mediated necrotizing myopathy (IMNM), and inclusion body myositis (IBM).¹

Myositis-specific autoantibodies (MSAs) are a common serologic feature of IIM and each MSA is closely associated with a particular type of IIM.^1,2 For example, autoantibodies recognizing one of the tRNA synthetases (e.g., Jo1, PL-7, or PL-12) are found in AS, a disease characterized by myositis, interstitial lung disease (ILD), arthritis, Raynaud phenomenon, and rash. In contrast, autoantibodies recognizing 3-hydroxy-3-methylglutarylcoenzyme A reductase (HMGCR) or signal recognition particle (SRP) are associated with IMNM, which is usually dominated by skeletal muscle involvement. Most patients with DM have an autoantibody recognizing Mi2, TIF1γ, NXP2, or MDA5; in addition to skin and muscle, DM may also affect other organ systems.

Within each type of IIM, individual MSAs define unique subtypes, each with its own clinical features, prognosis, and response to treatment. For instance, in DM, malignancy, cutaneous calcinosis, and rapidly progressive ILD are strongly associated with autoantibodies recognizing TIF1γ, NXP2, and MDA5, respectively.^1,2 Prior reports have characterized patients with anti-Mi2–positive DM as having mild muscle disease^3,4 along with typical DM skin manifestations, infrequent ILD, and low cancer risk.^3,5–7 However, these studies were limited by small numbers of patients,^5,6 a lack of longitudinal assessments,^3,5–7 or the absence of currently recognized comparison groups such as AS or IMNM.^3,5–7 In addition, the evolution of anti-Mi2 autoantibody levels over years of follow-up has not been described. Thus, the purpose of the current study was to define the phenotype of anti-Mi2–positive DM by comparing the prevalence and severity of the clinical features of these patients to those with anti-Mi2–negative DM, AS, and IMNM at disease onset and during the course of the disease. Furthermore, our study aimed to analyze the trend of anti-Mi2 autoantibody levels in patients with long follow-up times.

Methods

Study population and autoantibody testing

All patients enrolled in the Johns Hopkins Myositis Center Longitudinal Cohort Study between 2002 and 2018 were included in the current study if they were positive for autoantibodies recognizing Mi2, NXP2, TIF1γ, MDA5, Jo1, PL-7, PL-12, SRP, or HMGCR by at least 2 immunologic techniques from among the following: ELISA, in vitro transcription and translation immunoprecipitation, line blotting (EUROLINE myositis profile), or immunoprecipitation from S35-labeled HeLa cell lysates.^8,9 Patients were included in the DM group if they had autoantibodies recognizing Mi2, NXP2, TIF1γ, or MDA5. Alternatively, patients were classified as having AS if they had autoantibodies against Jo-1, PL-7, or PL-12. Finally, patients were included in the IMNM group if they tested positive for anti-SRP or anti-HMGCR autoantibodies.

Strength was evaluated by the examining physician using the Medical Research Council scale. This scale was transformed to the Kendall 0–10 scale for analysis purposes as previously described.¹⁰ Although more than 10 different physicians measured patient strength, serial strength measurements for each patient were made by the same physician. For the purposes of analyses, right- and left-side measurements for arm abduction and hip flexion strength were combined and the average was used for calculations (possible range 0–10). Serum creatine kinase (CK) levels were included for the longitudinal analysis if obtained within a period of 6 weeks before or after strength testing. Skin manifestations (i.e., heliotrope rash or Gottron sign), weakness, symptoms of esophageal involvement, antisynthetase syndrome–associated clinical features (e.g., mechanic’s hands, Raynaud phenomenon, arthritis, fever), and other clinical features were documented both retrospectively at the onset of the disease (by asking patients about features present at the onset of disease) and prospectively at each visit. ILD was defined through a multidisciplinary approach as recommended by the American Thoracic Society.¹¹ All available muscle biopsies were interpreted at the Johns Hopkins Neuromuscular Pathology Laboratory by pathologists blinded to autoantibody status. The pathologists systematically reported on the presence or absence of perifascicular atrophy, perivascular inflammation, primary inflammation (i.e., the invasion of non-necrotic fibers by mononuclear cells), and necrotizing myopathy (i.e., prominent myofiber necrosis in the absence of perifascicular atrophy or primary inflammation).

Anti-Mi2 autoantibody titers

For the quantitative anti-Mi2 autoantibody ELISA, 96-well ELISA plates were coated overnight at 4°C with 100 ng of Mi2b protein (ab124864; Abcam, Cambridge, UK) diluted in phosphate-buffered saline (PBS). Replicate wells were incubated with PBS alone. After washing the plates, human serum samples, diluted 1:400 in PBS with 0.05% Tween (PBS-T), were added to wells (1 hour, 37°C). After washing, horseradish peroxidase–labeled goat anti-human antibody (Jackson ImmunoResearch [West Grove, PA] 109-036-088; 1:10,000) was added to each well (30 minutes, 37°C). Color development was performed using SureBlue peroxidase reagent (KPL) and absorbances at 450 nm were determined. For each sample, the background absorbance from the PBS-coated wells was subtracted from that of the corresponding Mi2-coated well. Test sample absorbances were normalized using linear regression to a range of serial dilutions from an arbitrary positive anti-Mi2 patient, a reference serum included in every ELISA. Sera from 49 healthy control participants enrolled at the NIH Clinical Center were tested using this ELISA, revealing a mean absorbance of 0.08 with an SD of 0.03. The cutoff for a negative anti-Mi2 autoantibody titer was subsequently defined as the mean absorbance value plus 3 SDs of the healthy control participants (i.e., 0.17).

Standard protocol approvals, registrations, and patient consents

This study was approved by the Johns Hopkins and NIH Institutional Review Boards; written informed consent was obtained from each participant.

Statistical analysis

Dichotomous variables were expressed as percentages and absolute frequencies and continuous features were reported as means and SDs. Pairwise comparisons for categorical variables between groups were made using χ² test or Fisher exact test, as appropriate. Student t test was used to compare continuous variables among groups and paired t test was used to compare the level of weakness of different muscle groups. CK, a highly positively skewed variable, was expressed as median, first, and third quartile for descriptive purposes, and was transformed through a base-10 logarithm for regression analysis.

The standardized mortality incidence rate (SMR) and standardized cancer incidence rate (SCR) were calculated as previously described.⁹

To account for differing numbers of visits per patient, the evolution of the CK levels and muscle strength were studied using multilevel linear regression models with random slopes and random intercepts. The mean of hip flexor and arm abductor strength (range 0–10) was used as the strength outcome for regression analysis.

Locally weighted regression was applied to graphically analyze the evolution of the strength, CK levels, and pulmonary function tests. Kaplan-Meier curves and Cox regression were used to study the evolution of each of the clinical features over time.

The influence of nonmodifiable risk factors (sex, race, duration of disease, and age at the onset of the first symptom), the corticosteroid dose, and the administration of IV immunoglobulins, rituximab, methotrexate, azathioprine, and mycophenolate were used as adjusting covariates. Other treatments administered to fewer than 10% of the cohort were not included in the analysis.

All statistical analyses were performed using Stata/MP 14.1. A 2-sided p value of 0.05 or less was considered statistically significant with no adjustment for multiple comparisons.

Data availability

All data relevant to the study are either included in the article or will be shared at the request of other investigators.

Results

Epidemiologic features of patients with anti-Mi2 DM

From among 2,475 patients enrolled in the Johns Hopkins Myositis Center longitudinal cohort (which includes patients with IBM and other muscle diseases), 533 patients (22%) were included in this study. Among these, 58 (11%) had anti-Mi2–positive DM, 143 (27%) had anti-Mi2–negative DM, 162 (30%) had AS, and 170 (32%) had IMNM. The general features of anti-Mi-2–positive patients and the comparator groups are detailed in table 1. Compared to patients with anti-Mi2–negative DM, patients with anti-Mi2–positive DM were less likely to be female (62% vs 78%; p = 0.02) or white (64% vs 79%; p = 0.02). The prevalence of coexisting anti-Ro52 autoantibodies was markedly less in patients with anti-Mi2–positive DM than in patients with AS (17% vs 81%, p < 0.001). The longitudinal analysis included information from 5,013 patient visits and 492 (10%) of these were from patients with anti-Mi2–positive DM. The rates of mortality and cancer were not significantly different between the different myositis groups. Furthermore, patients with anti-Mi2–positive DM did not have higher mortality or cancer rates than a control reference population (SMR 0.9, 95% confidence interval [CI] 0.1–3.2; SCR 2.4, 95% CI 0.8–5.5). The 2 patients with anti-Mi2–positive DM who died during the study follow-up did so at an advanced age from causes unrelated to the myositis.

Table 1.

General features of anti-Mi2–positive patients

Muscle involvement

At disease onset, weakness was present in 60% of patients with anti-Mi2–positive DM and in 46% of patients with anti-Mi2–negative DM (p = 0.07). Compared to those with IMNM, fewer patients with anti-Mi2–positive DM had weakness at disease onset (60% vs 88%; p < 0.001) (table 2). During the follow-up period, weakness occurred more commonly in patients with anti-Mi2–positive DM than in patients with DM without this autoantibody (98% vs 85%; p = 0.008) (table 3).

Table 2.

Clinical features of anti-Mi2–positive patients at the onset of the disease

Table 3.

Cumulative clinical features of anti-Mi2–positive patients

At the initial visit, patients with anti-Mi2–positive DM had weaker neck flexors, arm abductors, elbow extensors, hip flexors, hip extensors, and knee flexors than those with anti-Mi2–negative DM or AS (all p < 0.05, table 4). However, patients with anti-Mi2–positive DM had stronger hip flexors than did patients with IMNM at their first visit. During the course of follow-up, patients with anti-Mi2–positive DM had significantly weaker mean hip flexor and mean arm abduction strength than patients with anti-Mi2–negative DM or patients with AS (all p < 0.01, table 5). During this time, patients in the anti-Mi2–positive DM group had substantially stronger mean hip flexors than those in the IMNM group (p < 0.001). However, the mean arm abduction strength of the patients with anti-Mi2–positive DM was remarkably similar to those with IMNM. Also, muscle enzymes were significantly higher in anti-Mi2–positive than in anti-Mi2–negative DM (p < 0.001) (table 5). Distal weakness was minimal in anti-Mi2–positive DM and control groups (table 4).

Table 4.

Pattern of weakness at the first visit of anti-Mi2–positive patients

Table 5.

Disease activity

Multilevel regression analysis confirmed that patients with anti-Mi-2–positive DM were weaker than patients with anti-Mi2–negative DM or patients with AS (all p < 0.004) and stronger than patients with IMNM (p = 0.003) independent of the time from onset, age, sex, race, or treatments at any given time during follow-up. This analysis also demonstrated that, compared to anti-Mi2–positive DM, CK levels were lower in anti-Mi2–negative DM (p < 0.001), higher in IMNM (p < 0.001), and similar in AS (p = 0.6). The regression analysis also showed that in anti-Mi2–positive DM, CK levels were inversely associated (p < 0.001) with strength, independent of the aforementioned variables.

Kaplan-Meier curves and Cox regression confirmed that most patients with anti-Mi2–positive DM developed weakness within the first 2 years of disease and that this occurred faster than in anti-Mi2–negative DM or AS (all p < 0.001) independent of the age at onset, race, or sex.

Of those patients with anti-Mi2–positive DM who underwent thigh muscle MRI (n = 23), muscle edema, muscle atrophy, fatty replacement, and fascial edema were present in 78%, 26%, 39%, and 43% of patients, respectively (table 6). Compared to patients with anti-Mi2–positive DM, more patients with IMNM had atrophy (68% vs 26%; p < 0.001) and fatty replacement (87% vs 39%; p < 0.001).

Table 6.

Muscle biopsy and thigh MRI results

Most patients with anti-Mi2–positive DM with weakness at the first visit regained full strength within the first year of treatment (figure) using a combination of corticosteroids along with methotrexate, mycophenolate, or azathioprine. Substantial flares of weakness (defined either as increasing CK levels or worsening weakness) after treatment introduction were exceptional in our cohort.

Figure. Evolution of strength and creatine kinase (CK) levels in anti-Mi2–positive patients.

(A–C) In these graphs, the dots represent individual data points for strength and CK levels; the lines are the output of locally weighted regression analyses using these data to graphically analyze the evolution of the strength and CK over time.

Overall, anti-Mi2 autoantibody titers showed a weak but significant association with increased CK levels (b = 0.34, p < 0.001) and decreased strength (b = −1.13, p < 0.001). Ten patients had 5 or more serum samples available during the course of disease and autoantibody titers declined over time in each patient. Indeed, antibody titers normalized in 3 (30%) of these patients. Importantly, 3 (30%) of these patients with decreasing autoantibody titers (including 2 of the 3 who reached the autoantibody normality threshold) could stop treatment and never relapsed. Moreover, the strength and CK levels mirrored the evolution of the autoantibody titers in most of the patients.

Muscle biopsies were available for review from 27 patients with anti-Mi2–positive DM, 32 patients with anti-Mi2–negative DM, 26 patients with AS, and 81 patients with IMNM. There were no significant differences in the prevalence of perifascicular atrophy or perivascular inflammation between patients with anti-Mi2–positive DM and those with anti-Mi2–negative DM or AS (table 6). As expected, perifascicular atrophy was not present in muscle biopsies from patients with IMNM. Primary inflammation (i.e., the invasion of non-necrotic fibers by mononuclear cells), although only present in a minority of cases, was more common in anti-Mi2–positive than in anti-Mi2–negative DM (19% vs 0%; p = 0.02). Myofiber necrosis was the most prominent histologic feature in many anti-Mi2–positive DM muscle biopsies. Nonetheless, the histologic diagnosis of necrotizing myopathy was relatively uncommon in those with anti-Mi2–positive DM (19%), anti-Mi2–negative DM (6%), or AS (23%) compared to those with IMNM (78%; p < 0.001).

Extramuscular manifestations

At disease onset, the prevalence of extramuscular manifestations affecting the skin, lungs, esophagus, and joints was similar between patients with DM with and without anti-Mi2 autoantibodies (table 2). However, over the course of the disease, compared to anti-Mi2–negative patients, patients with anti-Mi2–positive DM had less calcinosis (9% vs 28%; p = 0.003), ILD (5% vs 16%; p = 0.04), and fever (7% vs 21%; p = 0.02). Similarly, compared to patients with AS, patients with anti-Mi2–positive DM had more frequent DM-specific rashes (i.e., heliotrope or Gottron sign/papules) (93% vs 57%; p < 0.001) and dysphagia (53% vs 37%; p = 0.03) but less frequent mechanic's hands (21% vs 56%; p < 0.001), ILD (5% vs 80%; p < 0.001), arthritis (21% vs 58%; p < 0.001), and fever (7% vs 22%; p = 0.009).

Discussion

In this study, we have shown that patients with anti-Mi2–positive DM are more likely to have weakness at disease onset and during the course of disease than patients with anti-Mi2–negative DM. Furthermore, patients with DM with anti-Mi2 autoantibodies have higher maximum CK levels and more severe proximal muscle weakness than patients with DM without these autoantibodies. Taken together, the data presented here suggest that patients with anti-Mi2–positive DM have more severe muscle disease than patients with anti-Mi2–negative DM. This study also shows that patients with anti-Mi2–positive DM have more severe muscle disease than patients with AS. In fact, the severity of muscle disease in the upper extremities of anti-Mi2–positive DM was comparable to that of IMNM. These findings contradict prior reports concluding that patients with anti-Mi2–positive DM have relatively mild myositis.^3,4 Although the reasons for this discrepancy are not immediately apparent, it should be noted that the conclusions of the prior reports were not supported by quantitative data such as the comparative analyses of muscle strength that were performed in the current study.

In contrast to the frequent and severe muscle disease seen in anti-Mi2–positive DM, this study shows that extramuscular manifestations are generally less common in this group than in anti-Mi2–negative DM or AS. Specifically, calcinosis, ILD, and fever were less common than in anti-Mi2–negative DM, whereas ILD, arthritis, and fever were less common than in AS. As expected, many extramuscular manifestations were more common in anti-Mi2–positive DM than in IMNM, which predominantly affects skeletal muscle.

Unlike other DM myositis-specific autoantibodies like anti-TIF1γ¹² and, to a lesser extent, anti-NXP2,¹³ where the association with cancer has been confirmed by several groups, the association between anti-Mi2 autoantibodies and cancer is still a matter of debate.^3,6,7 In our study we could not find statistical evidence that anti-Mi2 autoantibodies are associated with cancer. However, the standardized cancer rate showed a trend towards a positive association with cancer (95% CI 0.8–5.5). Based on this, the safest recommendation for anti-Mi2 at this moment is to continue being screened for cancer within 3 years of the onset of myositis symptoms.

Prior studies have shown that serum levels of some MSAs, including anti-SRP¹⁴ and Jo-1,¹⁵ are closely associated with disease activity and may normalize during periods of remission. Levels of other MSAs, such as anti-HMGCR, may have an association with disease activity but remain present at high levels even when the disease appears to be relatively quiescent.^16,17 In the current study, we demonstrate that anti-Mi2 autoantibody levels are associated with muscle enzyme levels and strength, confirming the findings of a prior report.¹⁸ We also show that in 30% of patients followed for 4 or more years, anti-Mi2 autoantibody levels not only decline, but even normalize as the disease becomes less active. Although a prior study demonstrated a median drop in anti-Mi2 levels of 38.1% 44 weeks after receiving B-cell depletion therapy, normalization of autoantibody levels was not reported.¹⁸ The comparatively short follow-up period of this study may have precluded recording more substantial declines. Indeed, patients in the current study only had normalization of anti-Mi2 titers after 4–10 years of treatment.

Our observations regarding longitudinal anti-Mi2 autoantibody levels have several implications. First, since anti-Mi2 autoantibody levels declined or normalized even in the majority of patients who did not receive rituximab, declines in autoantibody levels are most likely not exclusively due to B-cell depletion. Also, because anti-Mi2 autoantibodies can become negative over time, it may be important to perform MSA testing early in the course of DM to avoid false-negative results. Moreover, taken together with those of a prior study,¹⁸ our results suggest that anti-Mi2 autoantibody levels may be a clinically useful biomarker of disease activity. However, future studies will be needed to determine whether it is safe to completely withdraw immunosuppressive medications from patients whose anti-Mi2 autoantibody levels have normalized.

The strengths of our study include the relatively long duration of follow-up of anti-Mi2–positive DM and control groups at the Johns Hopkins Myositis Center, enabling the study of clinical trajectory and long-term outcomes. Furthermore, the large size of this cohort has allowed for the robust comparison of multiple control groups including large numbers of patients with AS and patients with IMNM. However, due to the long-term existence of our cohort, many recently developed outcome measures were not collected for many patients. These include the Cutaneous Dermatomyositis Disease Area and Severity Index, patient and physician global assessments, and patient-reported outcome measures. Further work is warranted to incorporate these measures longitudinally for better comparison across cohorts.

Despite its limitations, this study shows that patients with anti-Mi2–positive DM have more severe muscle disease than patients with anti-Mi2–negative DM or patients with AS. This finding supports the growing body of evidence that DM is a heterogeneous disease and that MSAs such as anti-Mi2 can be used to classify patients with DM into more homogeneous subgroups.

Glossary

AS

antisynthetase syndrome

CI

confidence interval

CK

creatine kinase

DM

dermatomyositis

HMGCR

3-hydroxy-3-methylglutarylcoenzyme A reductase

IBM

inclusion body myositis

IIM

idiopathic inflammatory myopathies

ILD

interstitial lung disease

IMNM

immune-mediated necrotizing myopathy

MSA

myositis-specific autoantibodies

PBS

phosphate-buffered saline

SCR

standardized cancer incidence rate

SMR

standardized mortality incidence rate

SRP

signal recognition particle

Appendix. Authors

Study funding

This research was supported by the Intramural Research Program of the National Institute of Arthritis and Musculoskeletal and Skin Diseases of the NIH. S.K.D., L.C.-S., and the Myositis Research Database are supported by the Huayi and Siuling Zhang Discovery Fund. The authors thank Dr. Peter Buck for support. C.M. and J.A. are supported by the Jerome L. Greene Foundation. I.P.-F.’s research was supported by a Fellowship from the Myositis Association. E.T. is supported by the Jerome L. Greene Foundation and the Rheumatology Research Foundation Science Development Award.

Dislcosure

The authors report no disclosures relevant to the manuscript. Go to Neurology.org/N for full disclosures.