Antiphospholipid antibody testing.

Antiphospholipid antibodies (aPL) are a family of autoantibodies targeting phospholipid-binding proteins and are associated with several clinical settings, and most notably define the antiphospholipid syndrome (APS). These antibodies can be identified using a variety of laboratory tests, which include both solid-phase immunological assays and functional clotting assays that detect lupus anticoagulants (LA). aPLs are linked to a range of adverse medical conditions, such as thrombosis and complications affecting the placenta and fetus, potentially leading to morbidity and mortality. The specific aPL identified, along with the pattern of reactivity, correlates with the severity of these conditions. Therefore, laboratory testing for aPL is crucial for evaluating the risk of complications and for fulfilling certain classification criteria for APS, which are also applied as diagnostic markers in medical practice. This review provides an overview of the available laboratory tests currently for measuring aPL and discusses their clinical implications.

Predictive biomarkers of response to tocilizumab in giant cell arteritis (GCA): correlations with imaging activity.

In the recent EULAR recommendations, ultrasound examination is now recommended as a first-line imaging test in all patients with suspected giant cell arteritis (GCA) and the axillary arteries should be included in the standard exam. As an alternative to ultrasound evaluation, cranial and extracranial arteries can be examined using FDG-PET or MRI. The aim of our study was to observe in a retrospective case series whether there is a correlation between biomarkers and imaging activity in a population of patients followed in real life with GCA treated with prednisone (PDN) and tocilizumab (TCZ). We retrospectively enrolled 68 patients with newly diagnosed GCA between January 2020 and September 2021, followed in real life, who were examined at the Rheumatology Unit of the San Giovanni di Dio Hospital, Florence, Italy. Patients were evaluated at T0-T3-T6-T12-T18-T24 for the following blood tests: ESR, CRP, fibrinogen, platelet count, serum amyloid A (SAA), IL-6, and circulating calprotectin (MRP). Ultrasound examination of the temporal arteries and axillary arteries was assessed at T0 within 7 days of starting treatment with high-dose glucocorticoids and subsequently at T3-T6-T12-T18-T24. A scale from 0 to 3 with semi-quantitative tools (SUV max) was assessed at T0-T12-T24. The evaluation of the correlation coefficient between laboratory and imaging variables has shown that SAA and MRP have the most powerful correlation with the PET score (0.523 and 0.64), and MRP also has an excellent correlation coefficient with the Halo score (0.658). The evaluation of the ROC curves shows for a PET score 3 and SAA values higher than 26 mg/L, sensitivity of 81.5% and specificity of 84.1%, and for a PET score 3 and MRP values higher than 2.3 mcg/mL, sensitivity of 100% and specificity of 76.8%. In this study, we demonstrated that SAA and MRP can be useful as promising tools to detect GCA activity. The study demonstrates a good correlation between the two biomarkers and the imaging activity evaluated by the Halo and PET scores.

Reflecting on a decade of the international consensus on ANA patterns (ICAP): Accomplishments and challenges from the perspective of the 7th ICAP workshop.

The International Consensus on ANA Patterns (ICAP) is an ongoing international initiative dedicated to harmonizing technical and interpretation aspects of the HEp-2 IFA test. Comprised of internationally recognized experts in autoimmunity and HEp-2 IFA testing, ICAP has operated for the last 10 years by promoting accurate reading, interpretation, and reporting of HEp-2 IFA images by professionals involved in various areas related to autoimmune diseases, such as clinical diagnostic laboratories, academic research, IVD industry, and patient care. ICAP operates through continuous information exchange with the international community and encourages the participation of younger experts from all over the world. The 7th ICAP workshop has addressed several aspects that originated from this interaction with the international community and has effectively established objective goals and tasks to be delivered over the next two years. Some of these are outlined in this article, including the planning of three audio-visual educational modules to be posted at the www.anapattern.org website, the classification of two novel HEp-2 IFA patterns, the implementation of a project dedicated to continuously updating the information on the clinical and immunologic relevance of the HEp-2 IFA patterns, and the launch of two additional branches of the HEp-2 Clinical and Immunological (HEp-2 CIC) project.

Issues in autoantibody tests used in the classification criteria for autoimmune rheumatic diseases: the laboratory autoimmunologist's perspective.

Classification criteria of autoimmune rheumatic diseases are an important means to define homogenous groups of patients that can be compared across studies for clinical trials and research purposes. The measurement of autoantibodies is a relevant aspect in the definition of classification criteria, with a significant weight in the scores necessary to classify patients with autoimmune rheumatic diseases. The impact of autoantibodies has gradually increased over the years, contributing to the evolution and improvement of the classification criteria. However, these criteria often do not take into consideration how autoantibodies are measured, i.e. differences in diagnostic accuracy of the methods. This is a critical point especially when obsolete analytical methods that are no longer used in many clinical laboratories are taken into consideration. In this review we have critically examined assays and methods for the determination of autoantibodies that are (or could be) included among the classification criteria of autoimmune rheumatic diseases in light of more recent evidence and technology evolution.

Evaluation of a New Multiparametric Microdot Array-Based Immunoassay Panel for Systemic Autoimmune Disease Diagnosis.

BACKGROUND: The early reliable detection and quantification of autoantibodies play an important role in autoimmune disease diagnosis and in disease-course monitoring. New technologies, such as the multiplexed determination of autoantibodies, have recently been introduced and are being adopted more frequently. The aim of this study was to evaluate the ability of a new microdot array-based multiparametric assay (ZENIT AMiDot CTD panel, A. Menarini Diagnostics, Firenze, Italy) to correctly classify patients with autoimmune rheumatic diseases (ARDs) and compare it to a fluorescence enzyme immunoassay (FEIA) for the detection of anti-ENAs. METHODS: The study included 69 consecutive samples from patients with ARDs that were analyzed using two different methods (FEIA and AMiDot) to detect anti-CENP B and six anti-ENA antibodies: anti-Scl-70, anti-SSB/La, anti-Jo-1, anti-U1-RNP, anti-Ro52, and anti-Ro60. The control group sera came from sixty-eight blood donors. Tests were run on the automated slide processor ZENIT FLOW, and then the slides were imaged and analyzed using ZENIT fast. RESULTS: Since the samples were selected for at least one antibody positivity with an ARD diagnosis, we did not calculate clinical sensitivity but only specificity, which was 98.53%, ranging from 90% for anti-SSB/La antibodies to 100% for anti-CENP B ones. Mean agreement among the methods assessed by Cohen's kappa was 0.816 ± 0.240. CONCLUSIONS: The assay demonstrated good clinical performance and may be considered a valuable aid in detecting ARD patients, offering an alternative to methods such as FEIA which are largely in use today.

Do Ultrasound Lung Abnormalities Correlate to Biomarkers and Male Gender in Rheumatoid Arthritis Patients? A Monocentric Cross-Sectional Study.

Background: Lung ultrasound (LUS) is a tool of growing interest in Rheumatoid Arthritis (RA) oligo- symptomatic ILD to avoid. Objective: We aimed to evaluate (i) the prevalence of pleural (PLUS) and parenchymal (PAUS) abnormalities in LUS in the RA population and their possible correlation to biomarkers; (ii) the predictivity of gender, smoking habits, previous infections (past COVID-19 tuberculosis), and treatments; (iii) the differences in LUS between sexes. Methods: We collected the data of 155 (15 early and 140 late) RA patients with mild respiratory symptoms, evaluating PLUS and PAUS, in fourteen lung areas and also summing the scores (LUS-T). Results: Only 13/155 (8.4%) were completely negative; LUS correlated to age (all parameters p 0.0001), rheumatoid factor IgM (PLUS p 0.0006, PAUS p 0.02, LUS-T p 0.001) and ACPA (p 0.001, 0.006, 0.001, respectively), and PLUS also correlated to IL6 (p 0.02). The male gender was predictive of all LUS evaluations (p 0.001, 0.05, 0.001, respectively), which were higher than in women (p 0.001, 0.01, 0.001, respectively). Other potential risk factors were independent, except biological treatments, which showed a low predictivity to PLUS (p < 0.05). Conclusions: We can conclude that LUS is a useful technique in RA low respiratory symptoms and correlates with age, the most important RA biomarkers, and male sex.

Advanced Atrioventricular Block in Athletes: Prevalence and Role of Anti-Ro/Sjögren Syndrome-Related Antigen A Antibodies.

BACKGROUND: Advanced atrioventricular block (AVB), that is, higher than second-degree Mobitz-1, is an abnormal finding in athletes. Despite intensive investigation, in several cases the pathogenesis remains unknown, but frequently pacemaker implantation is still indicated. Increasing evidence points to circulating anti-Ro/Sjögren syndrome-related antigen A (SSA) antibodies cross-reacting with L-type calcium channel and inhibiting the related current as an epidemiologically relevant and potentially reversible cause of isolated AVB in adults. The aim of the study was to determine the prevalence of anti-Ro/SSA-associated advanced AVBs in a large sample of young athletes. METHODS AND RESULTS: A total of 2536 consecutive athletes aged <40 years without a history of cardiac diseases/interventions were enrolled in a cross-sectional study. Resting and exercise electrocardiography was performed, and those presenting any AVB were further evaluated by 24-hour Holter ECG. Athletes with second-degree AVBs and their mothers underwent anti-Ro/SSA testing. Moreover, purified immunoglobulin G from subjects with anti-Ro/SSA-positive and anti-Ro/SSA-negative advanced AVB were tested on L-type calcium current and L-type-calcium channel expression using tSA201 cells. The global prevalence of advanced AVB in the overall sample was ≈0.1%, but the risk considerably increased (2%) when intensely trained postpubertal male subjects were selectively considered. While none of the athletes with advanced AVB showed heart abnormalities, in 100% of cases anti-Ro/SSA antibodies were detected. Ex vivo experiments showed that immunoglobulin G from anti-Ro/SSA-positive but not -negative subjects with advanced AVB acutely inhibit L-type calcium current and chronically downregulate L-type-calcium channel expression. CONCLUSIONS: Our study provides evidence that advanced AVB occurs in young athletes, in most cases associated with anti-Ro/SSA antibodies blocking L-type calcium channels. These findings may open new avenues for immunomodulating therapies to reduce the risk of life-threatening events in athletes, avoiding or delaying pacemaker implantation.

Comparison of ELISA with automated ECLIA for IL-6 determination in COVID-19 patients: An Italian real-life experience.

Objectives: Coronavirus disease 2019 (COVID-19) has a wide spectrum of clinical severity. A cytokine storm is associated with COVID-19 severity. Of these, IL-6 is significantly associated with higher mortality and is also a marker for predicting disease prognosis. IL-6 may act as a target for therapeutics and, a blockade of IL-6 function by Tocilizumab has been described as a treatment of the inflammatory process COVID-19-related. This study aims to describe our experience comparing two different methods, in detail Human IL-6 Instant ELISA and the Elecsys IL-6 based on ECLIA, for the IL-6 assessment. Design and methods: IL-6 levels from serum samples of 104 COVID-19 patients, admitted to the AOU Careggi (Hospital in Florence -Italy), were assessed by using the two above-mentioned methods, and the results were analysed through Passing-Bablok regression fit and Bland-Altman plot. Results: The regression exhibited a linear relation between the methods with a regression equation (y = - 0.13 + 0.63 x; 95 % C.I. intercept = - 0.13 to 4.55; 95 % C.I. slope = 1.03 to 1.26 with R2 = 0.89, p > 0.05), showing a positive slope. The agreement of the two methods reported a bias of -25.0 pg/mL. Thus, the two methods correlate but do not agree in terms of numeric results. Conclusions: The two assays showed good comparability. However, because of the extremely wide linear range of the ECLIA, its throughput and its capacity for immune profiling, it represents an interesting emerging technology in the immunology field.

Real-Life Comparison of Four JAK Inhibitors in Rheumatoid Arthritis (ELECTRA-i Study).

Background: Real-world evidence of the efficacy and adverse events of JAK inhibitor treatment (Tofacitinib, Baricitinib, Upadacitinib, and Filgotinib) in rheumatoid arthritis is still limited. Methods: We studied 115 patients from the Rheumatology Unit of S. Giovanni di Dio Hospital affected by D2T-RA, according to the 2010 EULAR criteria. Out of the 115 patients, 17 had been treated with Baricitinib 8 mg/daily, 32 with Filgotinib 200 mg/daily, 21 with Tofacitinib 10 mg/daily, and 45 with Upadacitinib 15 mg/daily. We evaluated the clinical response after 3, 6, and 12 months of treatment and the follow-up from September 2022 to September 2023. All patients were evaluated according to the number of tender joints (NTJs), number of swollen joints (NSJs), visual analog scale (VAS), global assessment (GA), health assessment questionnaire (HAQ), Disease Activity Score (DAS28), and CDAI. Furthermore, laboratory parameters of efficacy and tolerability were evaluated. Results: All treatments demonstrated a statistically significant decrease in the DAS28 and CDAI scores, tender and swollen joint counts, VAS, HAQ, and patient global assessment (PGA) after 3, 6, and 12 months of treatment. All treatments showed similar behavior, and statistically significant decreases in circulating calprotectin, TNFα, and IL-6 were observed for all drugs after 12 months of treatment. In addition, soluble urokinase plasminogen activator receptor (suPAR) values showed significant differences at baseline and after 12 months of treatment for Filgotinib: 4.87 ± 4.53 vs. 3.61 ± 0.9 (0.009) and Upadacitinib: 6.64 ± 7.12 vs. 4.06 ± 3.61 (0.0003), while no statistically significant differences were found for Baricitinib: 3.4 ± 0.1 vs. 3.78 ± 0.1 and Tofacitinib: 3.95 ± 1.77 vs. 2.58 ± 0.1. The TC/HDL-C ratio (atherogenic index) showed significant differences when comparing Baricitinib vs. Filgotinib (0.0012), Filgotinib vs. Tofacitinib (0.0095), and Filgotinib vs. Upadacitinib (0.0001); furthermore, the LDL-C/HDL-C ratio in the Filgotinib group did not change (2.37 ± 0.45 vs. 2.35 ± 2.13 (NS)) after 12 months of treatment. Venous Thrombotic Events (VTEs) and major adverse cardiovascular events (MACEs) accounted for 1% of adverse events after treatment with Baricitinib. Herpes zoster reactivation accounted for 1% of adverse events after treatment with Filgotinib and Tofacitinib, while non-melanoma skin cancer (NMSC) accounted for 1% of adverse events after Upadacitinib treatment. Conclusions: Our real-world data from patients with RA show differences in some laboratory parameters and in the impact of lipid metabolism in JAK inhibitor treatment.

Harmonization of ANA testing challenge: quantification strategy to accurately predict end-point titers avoiding serial dilution.

Despite the advantages of automated systems for antinuclear antibody (ANA) analysis, the prediction of end-point titers avoiding serial dilutions is still in progress. The aims of this study were to set a conversion table providing discriminant ranges of fluorescence signal intensity values (FI) corresponding to the end-point titers and validate this tool in a real-life laboratory setting. Eight hundred ninety-four serum samples were analyzed for ANA using Image Navigator System. In order to classify FI into non-overlapping groups corresponding to conventional end-point titers, statistical discriminant analysis was used. Validation study was performed calculating agreement and error rates between visual readings and conversion table of 1119 routine ANA positive samples. Setting of FI ranges corresponding to the end-point titers for different staining patterns was computed. For samples showing single pattern, the overall agreement between visual readings and conversion table was 98.4% for all titers ranging from 1:160 to 1:2560, of which 68.0% had the same titer and 30.4% were within ± one titer difference. Concordance rates according to ANA patterns were as follows: (1) nuclear 98.4%, of which 67.0% had the same titer and 31.4% ± one titer; (2) cytoplasmic 100%, of which 72.7% had the same titer and 27.3% than ± one titer; (3) mitotic 66.6%, of which 33.3% had more ± one titer. Our study developed a quantification method for autoantibodies titers assessment based on just one single sample dilution instead of traditional serial dilution approach, providing significant advantages in routine laboratory in terms of reduction in hand-on time and harmonization of results.

Adopting the International Consensus on ANA Patterns (ICAP) classification for reporting: the experience of Italian clinical laboratories.

The indirect immunofluorescence assay on HEp-2 cells (HEp-2 IFA) is still considered the reference method to detect anti-nuclear antibodies (ANA) because of its high sensitivity and represents a relevant tool for the diagnosis of autoimmune rheumatic diseases. During the last decade, the International Consensus on ANA Patterns (ICAP) initiative promoted harmonization and understanding of HEp-2 IFA staining pattern nomenclature, as well as promoting their use in patient care by providing interpretation for HEp-2 IFA test results. In conjunction with a nationwide survey on the evolution of autoantibody diagnostics in autoimmune rheumatic diseases, we focused on the adherence of the Italian laboratories to the ICAP nomenclature analyzing its lights and shadows. The recent ICAP-oriented report, largely used today among Italian laboratories, also represents a further step in harmonizing and improving communication with the clinicians, adding value to laboratory findings and helping with critical clinical decisions.

Different Biomarkers of Response to Treatment with Selective Jak-1 Inhibitors in Rheumatoid Arthritis.

BACKGROUND: Rheumatoid arthritis (RA) is a systemic autoimmune disease that causes progressive joint damage. The Janus kinase (JAK) inhibitors (JAK-I) represent a new therapeutic option for RA patients, blocking the intracellular JAK-STAT pathway. Today, no studies have been conducted to determine whether new biomarkers could better reflect disease activity in patients treated with JAK-I than traditional disease activity indicators. Thus, the aim of our study was to determine additional disease activity biomarkers in RA patients receiving selective JAK-1 inhibitors. METHODS: we enrolled 57 patients with RA: 34 patients were treated with Upadacitinib (UPA) and 23 patients with Filgotinib (FIL). All patients were evaluated for clinimetry with DAS28 and Crohn's Disease Activity Index (CDAI), number of tender and swollen joints, Visual Analogic Scale (VAS), Physician Global Assessment (PhGA), and Health Assessment Questionnaire (HAQ), at baseline and at the 12th week of treatment. Lymphocyte subpopulations, complete blood count, erythrocyte sedimentation rate (ESR), C-Reactive Protein (CRP), anti-cyclic citrullinated peptide antibodies (APCA), rheumatoid factor (RF) IgM, interleukin 6 (IL-6), circulating calprotectin (cCLP), tumor necrosis factor α (TNFα), soluble urokinase Plasminogen Activator Receptor (suPAR), complement functional activity were measured at baseline and after the 12th week of treatment. RESULTS: in both groups of patients, we documented a significant reduction in the clinimetric parameters DAS28, CDAI, number of tender joints, number of swollen joints, VAS, PhGA, and HAQ. Moreover, significant differences were reported for laboratory parameters of ESR, CRP, IL-6, suPAR, cCLP, and PLT/L ratio in both groups. However, no difference was demonstrated between the two groups for changes in renal, hepatic, and lipid parameters. CONCLUSIONS: the suPAR and cCLP levels may lead towards a different therapeutic choice between UPA and FIL, with the expression of two different RA pathophenotypes directing FIL towards a lymphocyte-poor form and UPA towards a myeloid form of RA.

Autoantibody status according to multiparametric assay accurately estimates connective tissue disease classification and identifies clinically relevant disease clusters.

OBJECTIVE: Assessment of circulating autoantibodies represents one of the earliest diagnostic procedures in patients with suspected connective tissue disease (CTD), providing important information for disease diagnosis, identification and prediction of potential clinical manifestations. The purpose of this study was to evaluate the ability of multiparametric assay to correctly classify patients with multiple CTDs and healthy controls (HC), independent of clinical features, and to evaluate whether serological status could identify clusters of patients with similar clinical features. METHODS: Patients with systemic lupus erythematosus (SLE), systemic sclerosis (SSc), Sjogren's syndrome (SjS), undifferentiated connective tissue disease (UCTD), idiopathic inflammatory myopathies (IIM) and HC were enrolled. Serum was tested for 29 autoantibodies. An XGBoost model, exclusively based on autoantibody titres was built and classification accuracy was evaluated. A hierarchical clustering model was subsequently developed and clinical/laboratory features compared among clusters. RESULTS: 908 subjects were enrolled. The classification model showed a mean accuracy of 60.84±4.05% and a mean area under the receiver operator characteristic curve of 88.99±2.50%, with significant discrepancies among groups. Cluster analysis identified four clusters (CL). CL1 included patients with typical features of SLE. CL2 included most patients with SjS, along with some SLE and UCTD patients with SjS-like features. CL4 included anti-Jo1 patients only. CL3 was the largest and most heterogeneous, including all the remaining subjects, overall characterised by low titre or lower-prevalence autoantibodies. CONCLUSION: Extended multiparametric autoantibody assay allowed an accurate classification of CTD patients, independently of clinical features. Clustering according to autoantibody titres is able to identify clusters of CTD subjects with similar clinical features, independently of their final diagnosis.

Rheumatoid factor isotypes in rheumatoid arthritis diagnosis and prognosis: a systematic review and meta-analysis.

OBJECTIVE: The first biomarker associated with the rheumatoid arthritis is rheumatoid factor (RF) and since the earliest reports a role has been proposed in the diagnosis and in the prediction of clinical features and outcome. The study of RF isotypes has further attempted to improve diagnostic accuracy and identify specific subgroups of patients. The main objective of this study is to provide an analysis of the literature on the role of RF isotypes in the diagnosis and prognosis of rheumatoid arthritis (RA). METHODS: We performed a systematic literature review and meta-analysis on the role of RF isotypes in RA (only in English, from PubMed, search terms: "rheumatoid factor isotypes", "diagnosis", "prognosis" and "rheumatoid arthritis", last search 31 July 2022, two independent assessment of quality and biases, results included in tables and in the meta-analysis). RESULTS: Thirty-six articles were examined (7517 patients). Testing all RF isotypes with latex test or nephelometry allows for the highest sensitivity (68.6%, 95% CI 66.2% to 71.0%); nonetheless, the determination of IgA isotype provides the highest specificity (91.4%, 95% CI 90.8% to 92.0%) and the highest positive likelihood ratio (7.7, 95% CI 5.7 to 10.4). When testing IgM isotype the highest diagnostic OR (21.7, 95% CI 16.1 to 29.3) is reached. When analysing anti-citrullinated protein antibodies, RF isotype determination increases diagnostic accuracy. On the other hand, these do not provide relevant prognostic information, as results are conflicting. CONCLUSIONS: Testing RF allows the highest sensitivity, while IgA isotype the highest specificity and positive likelihood ratio for RA diagnosis. On the other hand, determination of RF isotypes dose not allow prognostic information, as data are limited and heterogeneous.

An Italian nationwide survey on the evolution of autoantibody diagnostics in autoimmune rheumatic diseases.

OBJECTIVES: The advent of new technologies and the discovery of new antigenic targets of autoantibodies has led to significant changes in autoimmune diagnostics worldwide. To address the extent to which autoimmunology laboratories adhere to such innovation in testing and reporting practices, the Italian Society of Clinical Pathology and Laboratory Medicine launched a national survey to assess the current status of autoimmune diagnostics and to provide direction for further harmonisation. METHODS: A questionnaire covering topics related to the diagnosis of autoimmune systemic rheumatic diseases was distributed to 152 Italian autoimmunology laboratories. The 59 closed-answer questions were subdivided into four main sections: 1. the setting (university, hospital or private laboratory) and the number of tests carried out; 2. the technologies used and their level of automation; 3. the analytical phase (antibody tests and methods), including awareness of the International Consensus on ANA Patterns (ICAP) initiative; 4. reporting of results and clinician relations. RESULTS: A total of 121 laboratories (79.6%) responded to the survey (15% universities, 70 hospitals, and 15% private laboratories). Indirect immunofluorescence is used by 94.8% of respondents, chemiluminescence by 78.4%, fluoro-immuno-enzymatic assays by 67.5%, immunodot by 52.6%, line-immunoassay by 47.4%, addressable laser bead immunoassay by 10.3% and radioimmunological methods by 10.2%. The great majority of respondents implemented complete automation of the listed methodologies. 65% of participants state that they add an interpretative comment in the report. 45% of participants enjoy a collaborative relationship with clinicians; counselling activities are provided by almost half of participants. CONCLUSIONS: Survey results indicated that almost all respondent laboratories have broadened their antibody panel and that high-throughput technologies have been widely introduced. Gaps identified by the survey include a still incomplete compliance with guidelines in antibody profiles (e.g. in antiphospholipid syndrome and rheumatoid arthritis and reporting of test results. Awareness of these differences provides insights that may further contribute to achieving harmonisation in autoimmune diagnostics.

Immunogenicity after two and three doses of mRNA vaccine in patients with cancer treated with exclusive radiotherapy.

BACKGROUND AND PURPOSE: Data on immunoresponse after SARS-CoV-2 vaccines for patients treated with exclusive radiotherapy (RT) are scarce. Since RT may affect the immune system, we conducted the MORA trial (Antibody response and cell-mediated immunity of MOderna mRNA-1273 vaccine in patients treated with RAdiotherapy). MATERIALS AND METHODS: Data regarding humoral and cellular immune response of patients treated with RT were prospectively collected after the second and third dose of mRNA vaccines. RESULTS: Ninety-two patients were enrolled. With a median of 147 days after the second dose, the median SARS-CoV-2 IgG titer was 300 BAU/mL: six patients were seronegative (Spike IgG titer ≤ 40 BAU/mL), whereas 24, 46 and 16 were poor responders (Spike IgG titer:41-200 BAU/mL), responders (Spike IgG titer:201-800 BAU/mL) and ultraresponders (Spike IgG titer > 800 BAU/mL), respectively. Among seronegative patients, two patients were negative also for cell mediated response, as tested with IFN-γ release Assay (IGRA) test. With a median of 85 days after the third dose, the median SARS-CoV-2 IgG titer was 1632 BAU/mL in 81 patients: only two patients were seronegative, whereas 16 and 63 patients were responders and ultraresponders, respectively. Among the 2 persistently seronegative patients, IGRA test was negative in one who had previously received anti-CD20 therapy. Documented paucisymptomatic (n = 3) or asymptomatic (n = 4) infection occurred after the third dose, during the Omicron wave. CONCLUSION: In patients treated with exclusive RT, even during the Omicron breakthrough, robust humoral response and clinical protection from severe SARS-CoV-2 disease were achievable with three doses of mRNA vaccine.

Testing for isotypes does not help differentiating rheumatoid arthritis from other rheumatoid factor positive diseases.

Rheumatoid factors (RFs) are useful for diagnosis and classification of rheumatoid arthritis (RA). Nephelometric and turbidimetric techniques, which detect total RF but do not reveal the antibody isotype, are common diagnostic methods in clinical routine. Given the recent development of isotype-specific immunoassays, the detection of IgG, IgM, and IgA RFs represents an interesting challenge. The aim of the study was to evaluate whether specific RF tests performed as a second step after traditional nephelometry could help differentiating RA from other RF-positive diseases. We tested 117 consecutive serum samples that were RF-positive at nephelometry (BNII nephelometric analyzer, Siemens) for IgA, IgG, and IgM RF isotypes by a fluoroimmunoenzymatic assay (FEIA) on the Phadia 250 instrument (ThermoFisher). Fifty-five subjects had RA and 62 presented non-RA diagnoses. Eighteen sera (15.4%) were positive only by nephelometry, two were positive only for IgA RF, and the remaining 97 sera were all positive for IgM RF isotype (with or without IgG and IgA RF). Positive findings did not correlate with RA or non-RA diagnosis. Spearman rho correlation coefficient between nephelometric total RF and IgM isotype was moderate (0.657), and weak between total RF and IgA (0.396) and IgG (0.360) isotypes. Despite its low specificity, measurement of total RF by nephelometry still seems to be the method that performs best. As IgM, IgA, and IgG RF isotypes showed only a moderate correlation with total RF measurement, their diagnostic use as a second level test remains controversial.

Soluble Urokinase Plasminogen Activator Receptor (suPAR) in Autoimmune Rheumatic and Non Rheumatic Diseases.

The soluble urokinase plasminogen activator receptor (suPAR) is the bioactive form of uPAR, a membrane-bound glycoprotein, and it is primarily expressed on the surface of immunologically active cells. Mirroring local inflammation and immune activation, suPAR has gained interest as a potential prognostic biomarker in several inflammatory diseases. Indeed, in many diseases, including cancer, diabetes, cardiovascular diseases, kidney diseases, and inflammatory disorders, higher suPAR concentrations have been associated with disease severity, disease relapse, and mortality. Our review describes and discusses the supporting literature concerning the promising role of suPAR as a biomarker in different autoimmune rheumatic and non-rheumatic diseases.