Welcome to the 13th International Congress on Autoimmunity interactive program

Abstract Body

Patients with COVID-19 with lung or systemic involvement present coagulation abnormalities, such as prolongation of prothrombin time and activated partial thromboplastin time, increased D-dimer levels, and, in some cases, severe thrombocytopenia. These patients are at high risk for thromboembolic events (arterial or venous) and thrombotic microangiopathy. The incidence of thromboembolic events in patients with COVID-19 is probably underestimated because of the asymptomatic presentation and the failure to perform systematic imaging studies. Thrombotic microangiopathy has been found in most of the few autopsies that have been performed to date, and the presence of pulmonary thromboembolism and deep vein thrombosis is striking in many of them. This hypercoagulability situation resembles antiphospholipid syndrome (APS), especially in its most severe form, catastrophic APS.

Although it has been described that up to 87.7% of patients with severe forms of COVID-19 were positive for lupus anticoagulant (LA) during their stay in the Intensive Care Units, the prevalence and clinical association of the presence of antiphospholipid antibodies (aPL) and other molecules related to APS is not sufficiently known.

Recently, we have been able to demonstrate that blood levels of β2-glycoprotein I (GPI) are much lower in patients with COVID-19 than in the general population. Moreover, none of the patients who had normal β2GPI levels had respiratory failure or died. We have also found that the aPL prevalence in patients with COVID-19 is similar to that in controls of the same age (except for IgA-aβ2GPI, whose prevalence is significantly higher) without being associated with the incidence of thrombotic events or other complications of the disease.

Lack of β2GPI would impede regulatory function of coagulation and platelet aggregation, leaving patients without weapons to control a thrombotic storm. This situation would be clinically and functionally equivalent to APS, although autoantibodies would not be involved.

Abstract Body

Antiphospholipid syndrome is an autoimmune disease characterized by thrombosis and pregnancy morbidity occurring in patients persistently positive for aPL. Current therapeutic options remain confined to long-term anticoagulation with vitamin K antagonists. The future holds much promise with the identification of novel potential targets, many of which are currently under investigation. The challenge will be to design prospective randomized controlled clinical trials to provide the evidence necessary to support integration of these therapies into clinical practice.

Abstract Body

Antiphospholipid syndrome (APS) is an autoimmune disease mediated by the presence of antiphospholipd antibodies (aPL). These antibodies are able to provoke clotting episodes and pregnancy loss through the activation of different pathogenic pathways. Following this argument, the ideal treatment for the disease should aim to suppress or abrogate aPLs, like an immunosuppressive treatment can do. However, immediately from the 80s, it became clear that anticoagulation is the most effective treatment because it is able to directly impair the clotting of the blood, while aPLs seemed more resistant than other autoantibodies to the immunosuppressive treatments. During the following decades, the problems and difficulties linked to the prolonged anticoagulant therapy, the long term effects of the disease and of its various clinical manifestations (including catastrophic APS, CAPS, and the so-called "non-criteria manifestations") became evident. These observations prompted the scientific community to look for possible "disease-modifying drugs" as immunosuppressors distinct from the "symptomatic ones" like anticoagulants that impair the pathological process without removing the true causes of the process.

A good observation point is to look at Systemic Lupus Erythematosus (SLE) patients with APS that usually take combined immunosuppressive treatments because of their primary disease. Some of these patients were shown to become aPL negative and could withdraw anticoagulant treatment without thrombosis recurrence. At first, hydroxychloroquine (HCQ) was studied for its immunosuppressive effect on APS patients, probably taking advantage of its known antithrombotic effect, facilitating the physicians to prescribe it. HCQ was shown able to significantly decrease the aPL titers, but only in the long term (2-5 years). Belimumab is reported to decrease the aPL levels in SLE patients taking it for their disease, again after a long observation time. According to some case reports, Rituximab is effective in persistent aPL related thrombocytopenia.

Other observations on immunosuppressive treatment in APS, are related to the treatment of CAPS and microangiopathies associated to aPLs. In these conditions the use of corticosteroids and immunosuppressants (besides anticoagulants) is already included in the recommendations and guidelines. Lately also the inhibition of complement system by Eculizumab and the inhibition of the mTOR pathway by Sirolimus were suggested by experimental data and by some clinical reports.

Background and Aims

COVID-19 thrombosis resembles the antiphospholipid syndrome, characterized by vascular or gestational thrombosis and presence of antiphospholipid antibodies (aPL). The appearance of aPL in infections, the permanence or transience of these autoantibodies and their clinical repercussion has been widely studied. However, this situation has not yet been clarified in COVID-19.

Methods

A prospective study with 360 COVID-19 patients enrolled from the onset of the disease and followed-up for 6 months was performed. Criteria aPL included in Sidney classification aPL, extra-criteria aPL, including anti-B2GPI IgA and anti-phosphatidylserine/prothrombin IgG/M and anti-SARS-CoV-2 antibodies were determined at acute phase and >12 weeks later. A cohort of 143 healthy volunteers of the same age-range distribution was used as a reference group.

Results

The study of the prevalence of aPL in COVID-19 patients and the reference population did not show significant differences. The presence of aPL in both determinations was associated with thrombosis (OR: 2.33 and 3.71). Strong agreement for presence of classic aPL and anti-B2GPI IgA in first and second serum samples (Weighted kappa: 0.92) was observed. aPL-associated thrombosis appeared significantly later than non-apl related thrombosis with a median of 17 days after hospital admission (IQR: 6–28) vs. 4 days for the rest of thrombosis (IQR: 3–7). COVID-19 did not seem to induce aPL de novo, since anti-SARS-CoV-2 antibodies levels increased, aPL hardly changed.

Conclusions

In COVID-19 thrombosis at least two overlapping mechanism could co-exist, an early cytokine-storm related and a later-aPL-mediated thrombosis, with SARS-CoV2 infection as a second hit.

Background and Aims

Anti-lipid autoimmune diseases pose a difficult differential diagnosis with relevant clinical implications. There is a need of new tissue-specific autoantibodies for a more precise diagnosis. Our objective was to determine the incidence of myelin basic protein (MBP) antibodies detected on peripheral nerve by indirect immunoflourescence (IIF) and of antiphospholipid antibodies (aPL) in the serum of neurological patients.

Methods

Sera of 77 patients with suspected central (CNS) or peripheral nervous system (PNS) disorders were tested using neurology mosaics (Euroimmun, Lubeck, Germany) with tissue sections of cerebellum, plexus myentericus and peripheral nerve. All samples were tested for serum aPL antibodies (IgG and IgM anti-cardiolipin and anti-β2-glycoprotein I) using multiplex flow analyzer (Bio-Rad Laboratories, Hercules, CA). Positive results for aPL antibodies were consider as ≥ 7 UI/mL.

Results

Twenty-seven out of 77 (35%) had positive MBP antibodies. Of these 27, 29% (n=8) had positive aPL. Among these 8 patients, 5 were women (62.5%), median age 53 years (range, 33 to 71 years old). The most predominant aPL antibody found was IgM anti-β2-glycoprotein. The values obtained (median±SD) were as follows: IgG anti-cardiolipin: 29.32±69.82; IgM anti-cardiolipin: 11.25±4.26; IgG anti-β2-glycoprotein: 103.55±80.11; IgM anti-β2-glycoprotein: 14.10±57.55. The clinical manifestations included CNS (n=16) or PNS (n=11) signs and symptoms: white matter lesions, epileptic activity, MS-like syndrome, optic neuropathy, chronic migraines and peripheral neuropathies.

Conclusions

Peripheral anti-MBP autoantibodies were present in a third of our cohort with mainly CNS manifestations, pointing to a more extensive anti-lipid reaction. A further third of them had antiphospholipid syndrome, which has relevant therapeutic consequences (thromboprophylaxis).

Background and Aims

We tested the hypothesis that antiphospholipid antibodies (aPL) at high titers remain positive over time, while low titers fluctuate. We also evaluated the correlation between two assay platforms and between aPL and platelet-bound C4d (PC4d) as they are both associated with thrombosis in systemic lupus erythematosus.

Methods

Consecutive samples sent to Exagen for aPL testing during a 5-year period were analyzed. Serum anti-cardiolipin (aCL) and anti-beta2 glycoprotein-1 antibodies (b2GP1) were measured by chemiluminescence (QUANTA Flash; Werfen) or ELiA fluorescence enzyme immunoassay (Phadia; ThermoFisher Scientific). Anti-phosphatidylserine/prothrombin complex antibodies (aPS/PT) were measured by ELISA (QUANTA Lite; Werfen). Assays were performed following manufacturers' instructions. PC4d was measured by flow cytometry following Exagen’s standard operating procedures.

Results

Most positive samples with aCL and b2GP1 high titers - but not low titers - were positive at a retest. aPS/PT followed a similar trend. aCL and b2GP1 measured with two different technologies were highly correlated (Table). PC4d and IgG of the 3 aPL were at best moderately correlated even when analysis included only positive samples (coefficient: 0.20-0.27).

Conclusions

In the large Exagen laboratory dataset, high titers aPL are often persistently positive, allowing an earlier diagnosis and risk assessment at the time of the initial screening. The high correlation between two methodologies suggest that these findings are independent of assay platform. The low to moderate correlation between PC4d and aPL suggests additive value to evaluate association with thrombosis in autoimmune diseases.

Background and Aims

Non-criteria Anti-phospholipid antibodies have gained great interest in recent years for their role in the Antiphospholipid Syndrome as well as for their possible value in patients negative for anti-phospholipid antibodies (aPL) of the Sydney consensus and Lupus Anticoagulant (LA).

Here we study the presence of the non-criteria aPL among patients fulfilling APS clinical manifestations. We also aim to analyze the differential clinical characteristics of isolated positive patients for non-criteria aPL.

Methods

We studied a cohort of 838 patients who met the clinical APS classification criteria obtained from the 4487 patients with suspected-APS evaluated in 2017 at the Hospital 12 de Octubre. All the patients were evaluated for Anti-PS/PT(IgG/M), IgA anti-Beta 2 glycoprotein I, aPL and LA were evaluated.

A control group of 220 age and gander match heathy donors was use for comparisons.

Results

A total of 314 patients were positive for at least one aPL, of them 137(16%) met the Sidney laboratory criteria, 50 were anti-PS/PT isolated positive and 71 were isolated positive for the IgA anti-Beta 2 glycoprotein I. Patients isolated positive for LA were 54. Inter-rater agreement between anti-PS/PT(IgG/M) and LA was moderate: Kappa index 0.51 (CI:0.44-0.57). The inclusion of non-criteria aPL identifies 42% of the patient's positives for any aPL.

Conclusions

The inclusion of non-criteria aPL may help in APS diagnosis. LA and Anti-PS/PT identify different types of patients. Although 58% of LA + were also PS/PT +, there were isolated positives for both determinations with clearly different clinical characteristics.

Background and Aims

Antiphospholipid syndrome (APS) is a multisystemic autoimmune disorder characterized by thrombotic events and/or gestational morbidity in antiphospholipid antibodies (aPL) carriers. By a single-center study, the presence of circulating immune-complexes (CIC) formed by beta-2-glycoprotein-I (B2GP1) and IgG/IgM anti-B2GPI antibodies (B2-CIC) was associated with clinical manifestations related to APS but not included in the classification criteria (livedo reticularis, thrombocytopenia, low levels of complement factors). The aims of this multicentric study were to evaluate the prevalence and association of B2-CIC with APS-related characteristics.

Methods

A multicentric, cross-sectional, retrospective, and observational study was performed with 303 patients who met the APS classification criteria, recruited from 6 European Union hospitals. The presence of B2-CIC (IgG/IgM isotypes) and their association with clinical manifestations and biomarkers related to the disease activity were assessed.

Results

The prevalence of B2-CIC in APS patients was 39.3%. B2-CIC-positive patients with thrombotic APS had a higher incidence of heart valve thickening and dysfunction (OR: 9.6, p=0.015), triple aPL positivity (OR: 1.83; p=0.027), and thrombocytopenia (OR: 2.32; p=0.007), as well as lower levels of blood platelets, C3 and C4 (p=0.001, p<0.001 and p<0.001) compared to those B2-CIC negative. These differences were not observed in B2-CIC-positive patients with isolated gestational APS or in patients negative for B2-CIC.

Conclusions

Patients with thrombotic events and positive for B2-CIC had higher platelets and complement consumption than those negative, suggesting a greater degree of platelet activation associated with an increase of thrombocytopenia. B2-CIC could act as a possible biomarker of disease activity in APS patients.

Background and Aims

Vitamin D receptor (VDR) polymorphisms have been implicated as significant contributors in microbiota variation from the general population and patients with specific disease entities, but not specifically in Systemic Lupus Erythematosus (SLE) or Multiple Sclerosis (MS)

Methods

Cross-sectional fecal microbiome sampling of SLE and MS patients at a University referral centre, from September 2019 to February 2020. Microbiome composition was obtained by NGS sequencing of the V4 region of the 16S rRNA gene region. VDR polymorphisms FokI and TaqI were assayed by real-time PCR.

Results

48 patients were enrolled (29 MS; 19 SLE). 37 (77%) were females; SLE and MS patients were characterized by high relative abundance of Bacteroidota and Firmicutes, as has often been reported for the general population. There was no difference in microbiome composition between both disease groups. In both SLE and MS, FokI polymorphisms were not associated with abundance of specific phyla of fecal microbiome (Table 1 and 2). In MS, but not in SLE, TaqI CC genotype was associated with higher relative abundance of Proteobacteria (p-value= 0,025) (Table 3 and 4). Firmicutes/Bacteroidota ratio were not associated with FokI and TaqI polymorphisms

Conclusions

VDR polymorphisms may be associated with specific fecal microbiota abundance in MS patients, but not SLE patients. These preliminary exploratory results should be further investigated

Background and Aims

Hematological immune related adverse events (irAE) are rare, affecting 0.5% of patients on anti-PD(L)1, mainly represented by neutropenia, hemolytic anemia or immune thrombocytopenia. Warm type autoimmune hemolytic anemia (AIHA) is the most described presentation, while cold agglutinin disease (CAD) seems much rarer.

Methods

We report the case of a patient with CAD occurring during treatment with anti-PD-1 and review the literature.

Results

A 72-year-old patient with metastatic lung cancer developed grade 3 seronegative immune-induced cholangitis after 37 nivolumab infusions. Since liver tests rapidly normalized under ursodeoxycholic acid, without corticosteroid therapy, she was rechallenged with nivolumab. Ten infusions later, she complained about dyspnea and asthenia, and developed de novo cold-induced livedo. Biology revealed deep hemolytic anemia with hemoglobin 4.2 g/dl, positive Coombs test (IgG and C3d) and cold agglutinins, together with eosinophilia (2.98 G/L) and mild cholestasis. Comprehensive etiological workout was negative. She recovered after nivolumab discontinuation and high dose steroids (2 mg/kg).

Apart from our case, only seven ICI-induced CAD were reported: four women and three men, with a median age of 64 years, all occurring under anti-PD(L)-1 (3 under pembrolizumab, two nivolumab and two atezolizumab). The median time to onset was 15 days with grade ≥ 3 for 6/7 and one death. Coombs was positive in 6/7 cases. None of the cases described any other associated irAE. Four cases were steroid-refractory and required rituximab.

Conclusions

ICI-induced CAD is a rare and serious irAE. A late onset is possible and steriod sensitivity seems to be greater than in non-ICI-related CAD.