Frequent coexistence of anti-topoisomerase I and anti-U1RNP autoantibodies in African American patients associated with mild skin involvement: a retrospective clinical study

Autoantibodies to topoisomerase I (topo I, also known as Scl-70) is an established serologic marker of scleroderma (systemic sclerosis, SSc) and associated with diffuse scleroderma and severe interstitial lung disease (ILD) [1‐3]. It is highly specific for SSc when tested with standard double immunodiffusion [4, 5]; however, several studies using enzyme-linked immunosorbent assay (ELISA) reported high prevalence of anti-topo I in systemic lupus erythematosus (SLE) [6‐9], causing confusion and controversies [10, 11]. SSc could start from the Raynaud's phenomenon (RP), preceding the onset of SSc for many years, ILD, arthritis, and others [12]. Because autoantibodies are usually produced before typical clinical manifestations, it would not be a surprise to find anti-topo I in undifferentiated connective tissue disease (UCTD), undiagnosed patients [5], or even in certain patients with SLE who are going to develop SSc later [13]. The clinical association of anti-topo I was reevaluated based on radioimmunoprecipitation screening of sera from a cohort of unselected population in a rheumatology clinic that includes undiagnosed patients and patients with a wide variety of diagnoses in addition to established systemic autoimmune rheumatic diseases, such as SSc, SLE, polymyositis/dermatomyositis (PM/DM), and rheumatoid arthritis (RA).

Anti-topo I is a highly specific disease marker of SSc when tested by immunodiffusion [4, 5] or IP as in the present study. It can be occasionally found in undiagnosed patients such as UCTD [22] or RP [5], at least partially, because autoantibodies are usually produced before clinical manifestation [23]. In one study, anti-topo I were tested by ELISA in 2,181 unselected individuals to find none was positive [24]. All these data support the high specificity of anti-topo I for SSc.

Reports on high prevalence of anti-topo I in SLE by ELISA and its association with SLE activity and nephritis [8, 9] challenged the general observation on SSc specificity of anti-topo I and triggered much confusion and many controversies [5, 10, 11]. When we tested 46 SLE sera (from Louisiana, not included in the present study) by a commercial anti-topo I ELISA, 41% were positive; however, only two of 19 were IP positive [10]. In the study that had 32 (25%) of 128 prevalence of anti-topo I in SLE [8], only four of 32 ELISA positives were double immunodiffusion positive, and data supporting the specificity of ELISA were limited. Some also reported 13% to 29% prevalence of anti-topo I in SLE [6, 7, 9, 25] whereas others reported low prevalence by ELISA [5, 11]. Thus, the prevalence of anti-topo I in SLE appears to depend on the source of antigens or ELISA kits. In some studies [8‐10], anti-topo I ELISA positives in SLE are detecting antibodies that are different from those detected by immunodiffusion and IP. False positives caused by anti-dsDNA/chromatin antibodies in SLE sera in ELISA for autoantibodies to DNA-binding proteins, such as Ku and replication protein A, are well documented [10, 26]. Thus, the most likely explanation appears to be that anti-topo I ELISA positives in SLE are false positives caused by antibodies to DNA/chromatin. Because topo I is a nucleotide sequence nonspecific DNA-binding protein, one scenario is that serum DNA binds to topo I coated on plate, and this is followed by anti-DNA/chromatin antibodies binding to DNA. A second scenario is that preformed serum anti-DNA/chromatin immune complex can bind to topo I via its DNA component. It is also possible that anti-topo I ELISA positives in SLE in some studies reflect detection of low-affinity antibodies or antibodies other than IgG class because of secondary antibody specificity. Alternatively, certain ELISA antigens may contain impurities as unrelated antigens, or some SLE sera recognize denatured topo I epitopes not present in native molecules and thus appear unreactive (negative) in immunodiffusion or IP.

Anti-topo I antibodies are positive in 1% to 3% of SLE patients, even by reliable methods such as immunodiffusion [8]. This may be explained by SLE-SSc overlap syndrome, not typical pure SLE [10, 27], as shown in the present study. Thus, anti-topo I by immunodiffusion or IP is specific for SSc, and cautious interpretation is required for anti-topo I ELISA positive results in SLE.

SSc patients can be classified based on autoantibody specificities that are associated with unique clinical subsets [3]. Coexistence of SSc-related autoantibodies is uncommon [3]; however, a combination of anti-topo I and anti-U1RNP appears to be an interesting and possibly clinically useful exception. In addition to cases reported mainly from Japan [27‐29], frequent association of anti-topo I and anti-U1RNP in a large Japanese and American cohorts also was observed [1, 2]. In one study, nine (12%) of 78 of anti-topo I-positive SSc had coexisting anti-U1RNP, and an additional three later developed anti-U1RNP [1]. Three patients in this cohort also had anti-Sm antibodies [27]. A study from Finland reported 12% of coexistence of anti-topo I and anti-U1RNP [30]. Detection of anti-topo I in MCTD patients indicates coexisting anti-topo I and anti-U1RNP [31]. Regarding the issue of race and coexistence of these two specificities in SSc, the prevalence was reported as 2% in Caucasian, 13% in African American, and 16% in Japanese in another U.S. cohort [2]. The 50% prevalence of anti-U1RNP in anti-topo I-positive African Americans in the present study is higher than that in other studies to date. Furthermore, prevalence of diffuse scleroderma in African Americans was low versus that in the previous study [2]. Three of four cases of anti-topo I + U1RNP-positive African American patients can be classified as SSc by using the ACR criteria based on the presence of pitting scars and ILD [16]; however, they lack sclerodermatous skin changes. Thus, this subset of patients might not be included in the studies that selected SSc patients based on diagnosis by physicians [2, 32, 33], sclerodactyly as a minimum requirement [34], or by using other SSc criteria [35]. They can be easily classified as "SLE with ILD and RP" because this is the common pattern of presentation among anti-U1RNP-positive SLE or MCTD. This subset could also be real anti-topo I-positive SLE without features of SSc described in some literature [8]. It may be clinically important to identify anti-topo I, in addition to anti-U1RNP, in these patients, because the former could be associated with severe ILD and scleroderma renal crisis [2, 3].

Anti-topo I detected by IP in unselected rheumatology patients is highly specific for SSc. Anti-topo I and anti-U1RNP frequently coexist in African American patients, and they are associated with a subset of overlap syndrome of SLE, SSc, and PM/DM, characterized by RP, pitting scars, and ILD without sclerodermatous changes.