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OBJECTIVES: To discover new and detect known antisynthetase autoantibodies (ASAs) through protein immunoprecipitation combined with gel-free liquid chromatography-tandem mass spectrometry (IP-MS). METHODS: IP-MS was performed using sera of individuals showing features of antisynthetase syndrome (ASyS) without (n=5) and with (n=12) previously detected ASAs, and healthy controls (n=4). New candidate aminoacyl-tRNA-synthetase (ARS) autoantigens identified through unbiased IP-MS were confirmed by IP-western blot. A targeted IP-MS assay for various ASA specificities was developed and validated with sera of patients with known ASAs (n=16), disease controls (n=20) and healthy controls (n=25). The targeted IP-MS assay was applied in an additional cohort of patients with multiple ASyS features or isolated myositis without previously detected ASAs (n=26). RESULTS: Autoantibodies to cytoplasmic cysteinyl-tRNA-synthetase (CARS1) were identified by IP-MS and confirmed by western blot as a new ASA specificity, named anti-Ly, in the serum of a patient with ASyS features. Rare ASAs, such as anti-OJ, anti-Zo and anti-KS, and common ASAs could also be identified by IP-MS. A targeted IP-MS approach for ASA detection was developed and validated. Application of this method in an additional cohort identified an additional patient with anti-OJ autoantibodies that were missed by line and dot immunoassays. DISCUSSION: CARS1 is the dominant cognate ARS autoantigen of the newly discovered anti-Ly ASA specificity. Rare and common ASA specificities could be detected by both unbiased and targeted IP-MS. Unbiased and targeted IP-MS are promising methods for discovery and detection of autoantibodies, especially autoantibodies that target complex autoantigens.
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Enfermedades Pulmonares Intersticiales , Miositis , Humanos , Autoanticuerpos , Autoantígenos , ARN de TransferenciaRESUMEN
ObjectiveMultiple spliceosome components are known autoantigens in systemic sclerosis (SSc). Here we aim to identify new and characterize rare anti-spliceosomal autoantibodies in patients with SSc without known autoantibody specificity. MethodsSera that precipitated spliceosome subcomplexes, as detected by immunoprecipitation-mass spectrometry (IP-MS), were identified from a database of 106 patients with SSc without known autoantibody specificity. New autoantibody specificities were confirmed with immunoprecipitation-western blot. The IP-MS pattern of new anti-spliceosomal autoantibodies was compared with anti-U1 RNP-positive sera of patients with different systemic autoimmune rheumatic diseases and anti-SmD-positive sera of patients with systemic lupus erythematosus (n = 24). ResultsThe NineTeen Complex (NTC) was identified and confirmed as new spliceosomal autoantigen in one patient with SSc. U5 RNP, as well as additional splicing factors, were precipitated by the serum of another patient with SSc. The IP-MS patterns of anti-NTC and anti-U5 RNP autoantibodies were distinct from those of anti-U1 RNP- and anti-SmD-positive sera. Furthermore, there was no difference in IP-MS patterns between a limited number of anti-U1 RNP-positive sera of patients with different systemic autoimmune rheumatic diseases. ConclusionAnti-NTC autoantibodies are a new anti-spliceosomal autoantibody specificity, here first identified in a patient with SSc. Anti-U5 RNP autoantibodies are a distinct but rare anti-spliceosomal autoantibody specificity. All major spliceosomal subcomplexes have now been described as target of autoantibodies in systemic autoimmune diseases.
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Lupus Eritematoso Sistémico , Enfermedades Reumáticas , Esclerodermia Sistémica , Humanos , Autoanticuerpos , Empalmosomas/química , Lupus Eritematoso Sistémico/diagnóstico , Anticuerpos Antinucleares , AutoantígenosRESUMEN
PURPOSE: In up to 20% of patients with systemic sclerosis (SSc) no known autoantibody specificity can be identified. Recently discovered autoantigens, such as telomeric repeat binding factor 1 (TERF1), as well as established autoantigens, like RuvBL1/2, are associated with telomere and telomerase biology. We aimed to identify new telomere- and telomerase-associated autoantigens in patients with SSc without known autoantibody specificity. METHODS: Unlabelled protein immunoprecipitation combined with gel-free liquid chromatography-tandem mass spectrometry (IP-MS) was performed with sera of 106 patients with SSc from two tertiary referral centres that had a nuclear pattern on HEp-2 indirect immunofluorescence without previously identified autoantibody. Telomere- or telomerase-associated proteins or protein complexes precipitated by individual sera were identified. Candidate autoantigens were confirmed through immunoprecipitation-western blot (IP-WB). A custom Luminex xMAP assay for 5 proteins was evaluated with sera from persons with SSc (n = 467), other systemic autoimmune rheumatic diseases (n = 923), non-rheumatic disease controls (n = 187) and healthy controls (n = 199). RESULTS: Eight telomere- and telomerase-associated autoantigens were identified in a total of 11 index patients, including the THO complex (n = 3, all with interstitial lung disease and two with cardiac involvement), telomeric repeat-binding factor 2 (TERF2, n = 1), homeobox-containing protein 1 (HMBOX1, n = 2), regulator of chromosome condensation 1 (RCC1, n = 1), nucleolar and coiled-body phosphoprotein 1 (NOLC1, n = 1), dyskerin (DKC1, n = 1), probable 28S rRNA (cytosine(4447)-C(5))-methyltransferase (NOP2, n = 1) and nuclear valosin-containing protein-like (NVL, n = 2). A Luminex xMAP assay for THO complex subunit 1 (THOC1), TERF2, NOLC1, NOP2 and NVL revealed high reactivity in all index patients, but also in other patients with SSc and disease controls. However, the reactivity by xMAP assay in these other patients was not confirmed by IP-WB. CONCLUSION: IP-MS revealed key telomere- and telomerase-associated proteins and protein complexes as autoantigens in patients with SSc.
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Esclerodermia Sistémica , Telomerasa , Humanos , Autoantígenos , Telomerasa/metabolismo , Autoanticuerpos , Telómero , Proteínas Nucleares/metabolismo , Proteínas de Ciclo Celular/metabolismo , ATPasas Asociadas con Actividades Celulares Diversas/metabolismo , Proteínas Portadoras , ADN Helicasas/metabolismo , Proteínas de Unión al ADN/metabolismo , Proteínas de Unión al ARNRESUMEN
OBJECTIVES: Antinuclear antibodies (ANAs) are associated with several autoimmune diseases. Indirect immunofluorescence (IIF) on human epithelial type 2 (HEp-2) cells is the golden standard for ANA detection in the clinic. In case of a positive HEp-2 IIF test result, follow-up tests are done to determine autoantibody specificity. For a fraction of the HEp-2 IIF-positive samples, the nature of the autoantigens remains uncharacterized. Our objective was to characterize autoantigens in such samples. METHODS: To characterize autoantigens in an unbiased way, we combined protein immunoprecipitation with liquid chromatography (LC) tandem mass spectrometry (MS/MS) sequencing. RESULTS: Using such approach we detected the Ki antigen, also referred to as PA28γ, in the immunoprecipitate of serum samples of three individuals with an autoimmune disease. The HEp-2 nuclear speckled IIF fluorescent signal of all three serum samples was abolished after pre-absorption of the serum with recombinant Ki antigen, confirming that autoantibodies against Ki underlie the HEp-2 IIF signal. CONCLUSIONS: Our data suggest that anti-Ki autoantibodies can underlie a nuclear speckled HEp-2 IIF pattern.
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Autoanticuerpos , Enfermedades Autoinmunes , Humanos , Técnica del Anticuerpo Fluorescente Indirecta/métodos , Espectrometría de Masas en Tándem , Autoantígenos , Anticuerpos Antinucleares , Enfermedades Autoinmunes/diagnósticoRESUMEN
INTRODUCTION: Fatigue is a major determinant of impaired quality of life in primary Sjögren syndrome (pSS) patients. Effective therapeutic strategies are lacking. OBJECTIVES: To review the potential benefit of rituximab, a chimeric anti-CD20 antibody, in the treatment of fatigue in pSS. METHODS: A systematic review on the effect of rituximab on fatigue-related outcome measures was conducted, retrieving evidence from CENTRAL (Cochrane Central Register of Controlled Trials), MEDLINE (via PubMed), EMBASE, and Scopus. RESULTS: No benefit of rituximab over placebo on any fatigue-related outcome measure could be demonstrated in the included trials. Significant effects were only observed when compared with baseline, but not when compared with placebo. CONCLUSIONS: The use of rituximab for the treatment of pSS-related fatigue cannot be supported by the currently available evidence.
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Síndrome de Sjögren , Fatiga/tratamiento farmacológico , Fatiga/etiología , Humanos , Calidad de Vida , Rituximab/uso terapéutico , Síndrome de Sjögren/complicaciones , Síndrome de Sjögren/diagnóstico , Síndrome de Sjögren/tratamiento farmacológicoRESUMEN
Anti-transcription intermediary factor 1 (TIF1)-γ autoantibodies are robustly linked with cancer-associated DM in adults. This review aims to give an overview of the physiological context of TIF1-γ and to determine whether there is a pathophysiological link between anti-TIF1-γ autoantibodies and the occurrence of cancer. Detection of anti-TIF1-γ autoantibodies has a high sensitivity and specificity for cancer-associated DM in adults and is therefore useful for both diagnosis and cancer risk stratification. The function of the autoantigen, TIF1-γ, may provide insight into the mechanism behind this association. TIF1-γ is a ubiquitously present protein involved in various biological pathways, including TGF-ß signalling. In cancer, it can act either as a tumour suppressor or promoter, depending on the cellular context and cancer stage. Evolving data provide pathophysiological insights, linking anti-TIF1-γ autoantibodies to both the anti-tumour response and to muscle and skin damage. TIF1-γ expression is increased in muscle and skin tissue of patients with DM. Mutations or loss-of-heterozygosity in TIF1-γ alleles in malignant tissue may result in the expression of tumour-specific neo-antigens stimulating autoantibody production. The newly formed autoantibodies are hypothesized to cross-react with antigens in muscle and skin, driving the development of DM. Based on the current evidence, anti-TIF1-γ autoantibodies should be considered warning lights of a potential tumour autoantigen and should alert the physician to the possibility of an underlying cancer.
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Autoanticuerpos , Autoantígenos/inmunología , Dermatomiositis/inmunología , Factores de Transcripción/inmunología , HumanosRESUMEN
Mucormycosis is an aggressive invasive fungal infection that occurs rarely in immunocompetent but frequently in immunocompromised patients. We present a case of a 68-year-old patient with cutaneous mucormycosis due to Rhizopus pusillus. He was initially hospitalized for invasive pulmonary aspergillosis and diabetes mellitus secondary to acute graft-versus-host treatment with glucocorticoids after allogeneic hematopoietic stem cell transplantation for acute myeloid leukemia. Treatment with liposomal amphotericin B and posaconazole was initiated but the patient developed septic shock with multiple organ failure and died 5 days later. The risk factors, clinical presentation, treatment, and prognosis of cutaneous mucormycosis in hematopoietic stem cell and solid organ transplant patients are discussed.
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Trasplante de Células Madre Hematopoyéticas/efectos adversos , Infecciones Fúngicas Invasoras/diagnóstico , Mucormicosis/diagnóstico , Piel/microbiología , Anciano , Anfotericina B/uso terapéutico , Antifúngicos/uso terapéutico , Resultado Fatal , Humanos , Huésped Inmunocomprometido , Infecciones Fúngicas Invasoras/tratamiento farmacológico , Masculino , Mucormicosis/tratamiento farmacológico , Insuficiencia Multiorgánica , Rhizopus/aislamiento & purificación , Choque Séptico , Piel/patología , Triazoles/uso terapéuticoAsunto(s)
Autoanticuerpos/inmunología , Dermatomiositis/diagnóstico , Dermatomiositis/inmunología , Adolescente , Artralgia/diagnóstico , Artralgia/etiología , Creatina Quinasa/análisis , Dermatomiositis/tratamiento farmacológico , Femenino , Técnica del Anticuerpo Fluorescente/métodos , Glucocorticoides/uso terapéutico , Humanos , Metilprednisolona/uso terapéuticoAsunto(s)
Miositis , Polimiositis , Enfermedades Reumáticas , Reumatología , Adolescente , Adulto , Humanos , Estados UnidosAsunto(s)
Candida albicans , Candidiasis/microbiología , Tenosinovitis/microbiología , Adulto , Mano/microbiología , Humanos , MasculinoRESUMEN
A high prevalence of antinuclear antibodies (ANA) in COVID-19 has been insinuated, but the nature of the target antigens is poorly understood. We studied ANA by indirect immunofluorescence in 229 individuals with COVID-19. The target antigens of high titer ANA (≥1:320) were determined by immunoprecipitation (IP) combined with liquid-chromatography-mass spectrometry (MS). High titer ANA (≥1:320) were found in 14 (6%) of the individuals with COVID-19. Of the 14 COVID-19 cases with high titer ANA, 6 had an underlying autoimmune disease and 5 a malignancy. IP-MS revealed known target antigens associated with autoimmune disease as well as novel autoantigens, including CDK9 (in systemic sclerosis) and RNF20, RCC1 and TRIP13 (in malignancy). The novel autoantigens were confirmed by IP-Western blotting. In conclusion, in depth analysis of the targets of high titer ANA revealed novel autoantigens in systemic sclerosis and in malignant disease.
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Enfermedades Autoinmunes , COVID-19 , Neoplasias , Esclerodermia Sistémica , Humanos , Autoanticuerpos/análisis , Anticuerpos Antinucleares , Autoantígenos , Quinasa 9 Dependiente de la Ciclina , Proteínas Nucleares , Proteínas de Ciclo Celular , Factores de Intercambio de Guanina Nucleótido , ATPasas Asociadas con Actividades Celulares DiversasRESUMEN
INTRODUCTION: Myositis-specific autoantibodies (MSAs) are thought to be mutually exclusive in patients with idiopathic inflammatory myopathies (IIM) based on studies with immunoprecipitation-based (IP) detection methods. Recently, detection of multiple MSAs in unique patients is increasingly reported, but the extent of this phenomenon remains unclear. METHODS: At our centre, we reviewed results from two line immunoassays and one dot immunoassay in 145 IIM patients and 240 controls for the presence of multiple MSAs. Pubmed and Embase were systematically searched for articles mentioning detection of multiple MSAs in IIM patients, published until February 2019. We assessed the frequency, detection method, the precise combinations and clinical phenotypes of participants with multiple MSAs. RESULTS: At our centre, detection of multiple MSAs occurred in 3.4-8.3% of patients with IIM, depending on the assay. However, no cases with full concordance across all three assays were identified. Forty-four articles reported detection of multiple MSAs, representing a total of 133 cases, including four patients with a connective tissue disease other than IIM and two healthy controls. In 101 cases all MSAs were detected using only one detection method: 40 cases with IP-based methods (most frequently used technique) and 61 cases with other assay types. In most cases the phenotype of patients with multiple MSAs matched the predicted presentation associated with one MSA and in few cases the phenotype matched with both MSAs. CONCLUSION: Detection of multiple MSAs in unique IIM patients is less rare than commonly accepted. Specificity issues of the commercially available multiplex immunoassays may, at least partly, explain the higher frequency compared to IP-based methods. 'True multiple MSA-positive' patients may exist, though they are most likely rare.
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Autoanticuerpos , Miositis , Polimiositis , Humanos , Miositis/inmunología , Fenotipo , Polimiositis/inmunologíaRESUMEN
Systemic sclerosis (SSc) affects the upper gastrointestinal (GI) system in 90% of patients. High-resolution manometry (HRM) assesses esophageal dysmotility, but its role in diagnosis and follow-up remains unclear. The objectives of this systematic review were to investigate the role of HRM in the assessment of SSc-associated upper GI involvement and to evaluate the correlation between HRM abnormalities and clinical characteristics and the effects of therapeutic interventions on HRM findings. Fifteen articles were included. Most (11/15) studies were of very good or good quality. Most studies assessed correlations between esophageal symptoms and esophageal dysmotility. Two studies assessed the effectiveness of buspirone and reported HRM findings. Studies assessing upper GI symptoms using validated questionnaires, such as the University of California Scleroderma Clinical Trial Consortium Gastrointestinal Tract 2.0 or Gastrointestinal Symptoms Severity Index score, found an association between absent contractility on HRM and upper GI symptoms, but even asymptomatic patients often have esophageal body dysmotility on HRM. Esophageal dysmotility positively correlates with the presence of interstitial lung disease on high-resolution computed tomography and reduced diffusion capacity (< 0.8 of predicted value). Trials investigating the effect of buspirone demonstrate both increased lower esophageal sphincter resting pressure and reduced upper GI symptoms. Most studies report on limited patient numbers and retrospective data. Potential bias was minimized using quality appraisal. HRM findings correlate to upper GI symptoms when assessed by validated questionnaires and can detect response to therapy in buspirone trials. Esophageal body dysmotility on HRM positively correlates with the presence of interstitial lung disease. KEY POINTS: ⢠Esophageal body dysmotility on HRM correlates with presence of ILD. ⢠HRM findings seem to correspond to clinical symptom alleviation in interventional trials, but data are still limited. ⢠At present HRM, a procedure with a high negative burden to the patient, offers little to no role in the therapeutic strategy.
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Trastornos de la Motilidad Esofágica/diagnóstico , Trastornos de la Motilidad Esofágica/fisiopatología , Tracto Gastrointestinal/fisiopatología , Manometría/métodos , Esclerodermia Sistémica/fisiopatología , Trastornos de la Motilidad Esofágica/complicaciones , Humanos , Esclerodermia Sistémica/complicacionesRESUMEN
Objectives: To review the therapeutic option of Rituximab, a chimeric anti-CD20 antibody, in systemic autoimmune rheumatic diseases (SARDs) such as systemic lupus erythematosus, systemic sclerosis, primary Sjögren syndrome and idiopathic inflammatory myopathy. Methods: A non-systematic review was conducted. Results: The specific role and indication of rituximab in SARDs has been the subject of multiple trials in recent years. Evidence supports the use of rituximab in moderate-to-severe refractory systemic lupus erythematosus, diffuse skin involvement in systemic sclerosis and systemic involvement in primary Sjögren syndrome. Several guidelines have adopted these indications. In addition, there is a consensus about the use of rituximab in refractory myositis. The role of rituximab in interstitial lung disease associated with these SARDs needs to be further explored. Conclusion: Rituximab is a treatment option in several SARDs. Upcoming trials, use in daily practice and the safety profile are elaborated on.