A case series of diabetic ketoacidosis associated with SGLT2 inhibitors

Sodium-glucose cotransporter 2 inhibitors (SGLT2i) are increasingly initiated as treatment for type 2 diabetes due to favourable cardiorenal characteristics. However, studies have identified an increased risk of diabetic ketoacidosis (DKA). We carried out a retrospective, case-based study at East and North Herts NHS Trust between February 2018 and December 2020. Fifteen cases of SGLT2i associated DKA were identified in people with presumed type 2 diabetes;33.3% were classed as euglycaemic DKA with a blood glucose of <11mmol/L. All cases were associated with a significant precipitating factor including diarrhoea, vomiting, reduced oral intake and sepsis. One case was related to COVID-19. Two people were subsequently found to have raised islet autoantibodies suggesting type 1 diabetes or latent autoimmune diabetes in adults. It is important that awareness of SGLT2i associated DKA is raised among users and health care practitioners, including the recognition of euglycaemic DKA. Sick day rules should be emphasised and reiterated at clinical encounters. Non-specialists in primary care, oncology and in perioperative settings should be empowered to advocate for temporary withdrawal and there should be readier access to blood ketone monitoring when required. When SGLT2i associated DKA occurs, due consideration should be given to evaluate the diabetes classification and investigate the circumstances of the event. Copyright © 2023 John Wiley & Sons.Copyright © 2023 John Wiley & Sons, Ltd.

Dried Blood Spot Analyses for the Diagnosis of Anti-cytokine Autoantibodies

Background: High-titer neutralizing anti-cytokine autoantibodies have been shown to be involved in several acquired diseases, including pulmonary alveolar proteinosis, cryptococcal meningitis, and disseminated/extrapulmonary Nocardia infections (anti-GM-CSF autoantibodies), disseminated mycobacterial disease (anti-IFN-gamma autoantibodies), and some cases of severe COVID-19 infection (anti-type 1 interferons). Currently, patient blood samples are shipped via courier and require temperaturecontrolled conditions for transfer. This method is expensive and requires patients to have access to medical personnel to draw the blood. However, the well-established technique of collecting blood on a paper card as a dried blood spot (DBS) for diagnosis offers a point of care alternative which can be performed with a simple finger prick. This method is less invasive, cheaper, and allows for easy transport of patient samples. Method(s): 30 uL of whole blood from patients was blotted on filter paper and stored at 4C until use. The filter paper was hole punched and each punched spot was eluted with 150 uL of a 0.05% Tween PBS solution at room temperature overnight. The eluate was screened for anti-cytokine autoantibodies using a particle-based approach. Patient plasma was also screened in conjunction for comparison. Result(s): We confirmed the presence of autoantibodies in the DBS eluate from 4 previously diagnosed patients with anti-GM-CSF autoantibodies and 2 patients with anti-IFN-gamma autoantibodies. Functional studies showed the DBS eluate from a patient with anti-GM-CSF autoantibodies was able to block GM-CSF-induced STAT-5 phosphorylation in normal PBMC. As a proof of concept and to increase the number of patients evaluated, we also confirmed the presence of anti-cytokine autoantibodies using dried plasma eluate from 9 patients with known anti-GM-CSF autoantibodies and 9 patients with anti-IFN-gamma autoantibodies. Levels detected in DBS analyses were comparable to the levels found in plasma from the same patients not subjected to blotting and elution. Temperature studies showed that the autoantibodies were detected at similar levels when stored at 4C, 25C, and 40C for a week. Conclusion(s): The diagnosis of pathogenic anti-cytokine autoantibodies should be considered in the context of unusual or adult-onset infections, and screening for this diagnosis can be performed with dried blood spot testing.Copyright © 2023 Elsevier Inc.

Newborn-onset chronic urticaria as the first manifestation of NLRP3 mosaicism

Objectives: A 7-month-old boy presented with generalized urticaria since the first week of life, without any other clinical manifestation. Cow's milk allergy was ruled out. His development was normal for his age. Maternal history was significant for COVID-19 infection in the third trimester of pregnancy with mild symptoms. Family history was significant for dermatographism in a maternal uncle. Hives were migratory with no single lesion persisting more than 24 h. There were no recognizable triggers and only relieved for 1-2 days after each vaccination. Patient was treated with optimal doses of antihistamines without improvement. Method(s): Laboratory tests and further studies were performed Results: Laboratory tests were normal including complete blood testing, circulating autoantibodies and infectious studies. C-reactive protein level and erythrocyte sedimentation rate were elevated. Due to chronic urticaria of newborn onset unresponsive to antihistamines a monogenic autoinflammatory disease was suspected. A targeted gene panel covering causative genes revealed the unreported p.Gly307Ala variant in the NLRP3 gene with a variant allele frequency (VAF) of 3% compatible with gene mosaicism. NLRP3 variant was classified as "likely pathogenic" based on its location, where a different variant has been reported as causing a severe form of cryopyrin-associated periodic syndromes (CAPS), and bioinformatic analyses. As expected, the variant was absent in patient's parents supporting for its de novo nature. Vision and hearing exams were normal. Treatment with canakinumab will start soon. Discussion(s): CAPS are dominantly-inherited autoinflammatory diseases caused by gain-of-function NLRP3 variants. These variants are often germline, but in some reported cases the variants are postzygotic causing gene mosaicism as in the patient here described. We believe that the mild presentation in our patient, despite having a likely pathogenic variant, may be explained by the low VAF. The genetic diagnosis in our patient allowed early initiation of anti-IL-1 treatment, which probably will prevent the development of other CAPS manifestations.

CD4+ T cells from preeclamptic patients cause hypertension, autoantibodies and cognitive dysfunction in a pregnant rat model

CD4+ T Cells from Preeclamptic patients with or without a history of COVID-19 during pregnancy cause hypertension, autoantibodies and cognitive dysfunction in a pregnant rat model Objective: Preeclampsia (PE) new onset hypertension (HTN) during pregnancy, is associated with increased autoantibodies, cerebral blood flow (CBF) impaired cognitive function and memory loss. We have shown adoptive transfer of placentalCD4+T cells from PE women into athymic nude pregnant rats causesHTNand autoantibodies associated with PE.COVID-19 (CV) during pregnancy is associated with increased diagnosis of PE. However, we do not know the role of CD4+ T cells stimulated in response to CV in contributing to the PE phenotype seen patients with a Hx of CV during pregnancy. Therefore, we hypothesize that adoptive transfer of placental CD4+ T cells from patients with a CV History (Hx) during pregnancy with PE causes HTN, increased CBF and cognitive dysfunction in pregnant athymic nude recipient rats. Study Design: Placental CD4+ T cells isolated from normotensive (NP), PE, Hx of CV normotensive (CV Hx NT), and Hx of CV with PE (CV Hx+PE) at delivery. One million CD4+ T cells were injected i.p. into nude athymic rats on gestational day (GD) 12. The Barnes maze and the novel object recognition behavioral assays were used to assess cognitive function on GDs 15-19. Blood pressure (MAP) and CBF were measured by carotid catheter and laser Doppler flowmetry on GD19, respectively. A two-way ANOVA was used for statistical analysis. Result(s):MAPincreased inCVHx+PE (111 +/- 4, n = 4) and PE recipient rats (115 +/- 2 mmHg, n = 5) compared to CV Hx NT (100 +/- 4, n = 5) and NP (99 +/- 3 mmHg, n = 4, P < .05). CV Hx+PE and PE exhibited latency with errors navigating in the Barnes maze compared to CV Hx NT and NP groups. Locomotor activity was decreased in CV Hx+PE (P < .05) compared to PE, CV Hx NT, and NP groups. CV Hx+PE and PE spent more time exploring identical objects compared to CV Hx NT and NP groups. PE and CV Hx+ PE had increased CBF compared to CV Hx NT and NP rats. Conclusion(s): Our findings indicate that pregnant recipients of CD4+ T cells from PE with or without a Hx CV during pregnancy cause HTN, increased CBF and cognitive dysfunction compared to recipients of NP or NT Hx COVID-19 CD4+ T cells.

Characteristics and Clinical Implications of Anti-IFN-alpha cytokine antibodies in partial Recombinase Activating Gene Deficiency patients before and during the COVID-19 Pandemic

Introduction: With the onset of the COVID-19 pandemic, there was increased attention on anti- IFN-alpha autoantibodies and its correlation with severe clinical outcomes in a large group of patients. However, this correlation has not been extensively investigated in patients with partial Recombinase Activating Gene Deficiency (pRD) who are known to have increased prevalence of anti- IFN-alpha autoantibodies. Therefore, there is a need to assess the presence of anti- IFN-alpha antibodies in pRD patients before and after the COVID-19 pandemic and explore the relationship between anti- IFN-alpha antibody presence and clinical outcomes. Method(s): Sera was collected from the whole blood after informed consent and Enzyme-Linked Immunosorbent Assay was conducted to confirm the presence of IgG-specific anti- IFN-alpha autoantibodies. Positive samples were determined as OD values above 3 standard deviations of the healthy donor OD mean. Result(s): Our cohort included both adult (n = 13) and pediatric (n = 9) patients with variants in RAG1 and RAG2. Eleven patients (50%) out of the 22 showed elevated anti- IFN-alpha autoantibodies levels. Five patients (23%) were defined as low positive for anti- IFN-alpha autoantibodies, and 6 patients had no autoantibody titers. Of the 22 patients, 16 were symptomatic with infectious and non-infectious complications including recurrent viral and/or bacterial infections, autoimmune cytopenias, and lymphoproliferation. Ten (63%) of the symptomatic patients demonstrated high anti-IFN-alpha autoantibodies titers. Of the 11 patients with no or low neutralizing anti- IFN-alpha autoantibodies levels, 5 were asymptomatic. In temporal comparison, 16 samples were collected pre-COVID-19 pandemic;8 samples were collected during the pandemic, 2 of which belonged to patients with samples collected before and during the pandemic. In the pre-pandemic cohort, 66% had anti- IFN-alpha autoantibodies. Conversely, during the COVID-19 pandemic, 89% had anti- IFN-alpha autoantibodies. Of note, one patient who had neutralizing anti- IFN-alpha autoantibodies remained positive both before and during the pandemic despite HSCT. Patient also had a SARS-CoV-2 infection in summer of 2022 with a mild clinical course. Conclusions & Next Steps: We observed persistence of anti-IFN-alpha autoantibodies in our cohort post-pandemic and even post-HSCT. It is unclear whether the presence of anti-cytokine antibodies are risk factor for severe COVID-19.Copyright © 2023 Elsevier Inc.

2C5-surface modified nanocarriers targeting neutrophil extracellular traps

Background: Neutrophil extracellular traps (NETs) are composed of processed chromatin bound to granular and selected cytoplasmic proteins and released by neutrophils. NETs consist of smooth filaments composed of stacked nucleosomes. Fully hydrated NETs have a cloud-like appearance and occupy a space 10-15-fold larger than the volume of the cells they originate from. DNases are the enzymes that cleave extracellular DNA including NETs. Together with their protective role in microbial infections, NETs are involved in multiple pathological processes and represent key events in a variety of pathologies including cancer, autoimmunity, and cardiovascular disease. Sites of NETs concentration are dangerous for the host if the process of NETs formation becomes chronic or the mechanism of NETs removal does not work. NETosis has been linked to the development of periodontitis, cystic fibrosis, type 2 diabetes, COVID-19 or rheumatoid arthritis as well as cancer progression. Purpose(s): Thus, the destruction of NETs is of primary significance in many pathologies. In our approach, we are focusing on mimicking one of the natural mechanisms of destroying excessive NETs by delivering deoxyribonuclease I to the specific site of pathological NETs accumulation by modifying the nanoparticles using an anti-nucleosome monoclonal antibody (2C5). The antibody is specific to nucleosomes and can recognize histones in NETs. DNase I is U.S. Food and Drug Administration (FDA)-approved active component and is commonly used in therapeutic methods of modern medicine for cystic fibrosis to clear extracellular DNA fibers in the lungs and systemic lupus erythematosus. Recent findings have also shown the effectiveness of DNase I in the digestion of NETs. However, the low serum stability and fast deactivation by environmental stimuli have been considered as the limiting factors for clinical applications of DNase I, which can be overcome by its targeted specific delivery in pharmaceutical nanocarriers. Method(s): In this study, we generate NETs in vitro using human neutrophils and HL-60 cells differentiated into granulocyte-like cells. We used interleukin-8, lipopolysaccharide from E.Coli (LPS), phorbol myristate acetate (PMA), and calcium ionophore A23187 (CI) to generate the NETs. We confirmed the specificity of 2C5 toward NETs by ELISA, which showed that it binds to NETs with the specificity like that for purified nucleohistone substrate. We further utilized that feature to create two delivery systems (liposomes and micelles) for DNAse I enzyme to destroy NETs, which was confirmed by staining NETs with SYTOX Green dye and followed by flow cytometric measurements and microscopic images. Conclusion(s): Our results suggest that 2C5 could be used to identify and visualize NETs and serve as a ligand for NET-targeted diagnostics and therapies. Also, we proved that our carrier can successfully deliver DNase to NETs to provide their degradation.

Anti-interferon-alpha autoantibodies in patients with inborn errors of Immunity and rheumatic diseases before and during the COVID-19 pandemic

Introduction: Type 1 interferon (IFN) autoantibodies, such as anti-IFNalpha, have pathogenic significance in life-threatening COVID-19 pneumonia. Ten to twenty percent of severe COVID cases are associated with type I IFN autoantibodies. These autoantibodies likely pre-exist while others arise de novo relative to SARS-CoV-2 infection. It is unclear to what extent type I anti-IFN autoantibodies are induced by SARS-CoV-2 infection and contribute to COVID-19 severity. We investigated these phenomena in those with inborn errors of immunity (IEI) and rheumatic disease (RHE). Aim(s): We aim to compare the prevalence and neutralization ability of anti-IFNalpha autoantibodies in IEI and RHE patients using archived blood samples before and after the COVID-19 pandemic began. Method(s): We determined the presence of autoantibodies against IFNalpha in plasma samples by enzyme linked immunosorbent assay in 453 patients with IEI or RHE who were testing either before or after the COVID-19 pandemic began in March 2020. Using flow cytometry, we determined the function of IFNalpha autoantibodies in plasma to block CD4T cell activation by inhibiting STAT-1 phosphorylation. Result(s): We found that 25 patients with IEI or RHE were positive for anti-IFNalpha autoantibodies. 10 out of 229 patient samples collected before the pandemic (4.2%) tested positive whereas 15 out of 224 patient samples collected after the pandemic began (7.0%) were positive. Seven of the 25 patients (28%) who tested positive had neutralizing antibodies in plasma, which prevented STAT-1 phosphorylation in CD4T cells;all of these patients had partial recombination activating gene deficiency (pRD) except for one patient with autoimmunity, leukemia and selective IgA deficiency. One pRD patient had anti-IFNalpha autoantibodies with neutralization capacity before the pandemic, which persisted after hematopoietic stem cell transplantation (HSCT) with full immune reconstitution. The patient was immunized for SARS-CoV-2 before and after HSCT and acquired COVID-19 infection a year after HSCT. The patient was symptomatic but never hospitalized and fully recovered despite having anti-IFNalpha autoantibodies. Conclusion(s): Anti-IFNalpha autoantibody levels were comparable before and after the start of the COVID-19 pandemic in IEI and RHE patients but only 28% of cases were neutralizing. The clinical implications of these autoantibodies are yet to be determined.Copyright © 2023 Elsevier Inc.

Italian ancestral HLA haplotype predisposing to severe COVID-19 by low spike and high IFNalpha binding affinity

Background/Objectives: Validated association between COVID-19 and the most obvious candidate genes, e.g. HLA, is still missing. A weak association with class I HLA-C*04:01 was found for infection in Sardinians and for severity in another mixed population. Auto-antibodies to interferon type I have been implicated in the severity of COVID-19 in two studies. Method(s): The binding affinity between HLA molecules and SARS-CoV-2 spike protein and IFNalpha subunits was evaluated in silico. The presence of antibodies against one or more of the 12 IFNalpha subunits was evaluated in 160 hospitalized COVID-19 patients. The 10 most frequent haplotypes in the Italian population were tested in 1.997 SARS-CoV-2 infected patients (hospitalized versus not hospitalized). Result(s): The presence of auto-antibodies against at least one IFNalpha subunit was detected in 26% of patients. The haplotype A*24:02-B*35:02-C*04:01-DRB1*11:04-DQB1*03:01 was found to predispose to severity (p = 0.0018;p = 0.07 after Bonferroni correction) in patients <50 years. The haplotype includes alleles able to bind spike with low affinity (i.e. C*04:01 and DRB1*11:04) and IFNalpha with high affinity (i.e. DRB1*11:04). Conclusion(s): One of the 10 most frequent ancestral haplotype of the Italian population predisposes to severity likely reducing both innate immunity through IFNalpha auto-antibodies induction and adaptive immunity through weaker spike protein presentation.

Myocarditis and autoimmunity.

INTRODUCTION: Autoimmune myocarditis may develop due to heterogeneous causes. Myocarditis is often caused by viral infections, but it can also be caused by systemic autoimmune diseases. Immune checkpoint inhibitors and virus vaccines induce immune activation, and they can cause the development of myocarditis, as well as several immune-related adverse events. The development of myocarditis is dependent on the genetic factors of the host, and the major histocompatibility complex (MHC) may be an important determinant of the type and severity of the disease. However, non-MHC immunoregulatory genes may also play a role in determining susceptibility. AREA COVERED: This review summarizes the current knowledge of the etiology, pathogenesis, diagnosis, and treatment of autoimmune myocarditis with a particular focus on viral infection, autoimmunity, and biomarkers of myocarditis. EXPERT OPINION: An endomyocardial biopsy may not be the gold standard for the diagnosis of myocarditis. Cardiac magnetic resonance imaging is useful in diagnosing autoimmune myocarditis. Recently identified biomarkers of inflammation and myocyte injury are promising for the diagnosis of myocarditis when measured simultaneously. Future treatments should focus on the appropriate diagnosis of the etiologic agent, as well as on the specific stage of the evolution of immune and inflammatory processes.

Autoimmunity in Long Covid and POTS.

Orthostatic intolerance and other autonomic dysfunction syndromes are emerging as distinct symptom clusters in Long Covid. Often accompanying these are common, multi-system constitutional features such as fatigue, malaise and skin rashes which can signify generalized immune dysregulation. At the same time, multiple autoantibodies are identified in both Covid-related autonomic disorders and non-Covid autonomic disorders, implying a possible underlying autoimmune pathology. The lack of specificity of these findings precludes direct interpretations of cause and association, but their prevalence with its supporting evidence is compelling.

Rare Case of Immune-Mediated Necrotizing Myopathy With Isolated Dysphagia as Presenting Symptom

Introduction: Immune mediated necrotizing myopathy (IMNM) is a rare, but progressive disease that accounts for about 19% of all inflammatory myopathies. Dysphagia occurs in 20-30% of IMNM patients. It often follows proximal muscle weakness and ensues in the later stages of the disease. We report a rare case of IMNM, presenting with dysphagia as the initial symptom, followed by proximal muscle weakness. Case Description/Methods: A 74-year-old male with a past medical history of coronary artery disease, hypertension, and hyperlipidemia presented to the ED with 2-3 weeks of intractable nausea, vomiting, and dysphagia for solids and liquids. Vital signs were stable, and initial labs displayed an AST of 188 U/L and ALT of 64 U/L with a normal bilirubin. Computed tomogram of the chest, abdomen, and pelvis were negative. An esophagram showed moderate to severe tertiary contraction, no mass or stricture, and a 13 mm barium tablet passed without difficulty. Esophagogastroduodenoscopy exhibited a spastic lower esophageal sphincter. Botox injections provided no significant relief. He then developed symmetrical proximal motor weakness and repeat labs demonstrated an elevated creatine kinase (CK) level of 6,357 U/L and aldolase of 43.4 U/L. Serology revealed positive PL-7 autoxantibodies, but negative JO-1, PL-12, KU, MI-2, EJ, SRP, anti-smooth muscle, and anti-mitochondrial antibodies. Muscle biopsy did not unveil endomysial inflammation or MHC-1 sarcolemmal upregulation. The diagnosis of IMNM was suspected. A percutaneous endoscopic gastrostomy feeding tube was placed as a mean of an alternative route of nutrition. He was started on steroids and recommended to follow up with outpatient rheumatology. He expired a month later after complications from an unrelated COVID-19 infection. Discussion(s): The typical presentation of IMNM includes painful proximal muscle weakness, elevated CK, presence of myositis-associated autoantibodies, and necrotic muscle fibers without mononuclear cell infiltrates on histology. Dysphagia occurs due to immune-mediated inflammation occurring in the skeletal muscle of the esophagus, resulting in incoordination of swallowing. Immunotherapy and intravenous immunoglobulin are often the mainstay of treatment. Our patient was unique in presentation with dysphagia as an initial presenting symptom of IMNM, as well as elevated enzymes from muscle breakdown. It is critical as clinicians to have a high degree of suspicion for IMNM due to the aggressive nature of the disease and refractoriness to treatment.

Lymphocyte Populations and Autoantibodies, in a Cohort of Patients with Rheumatoid Arthritis: A Long-Term Study of Covid-19 Cases and Non-Covid-19 Controls

Objectives: To assess the immunological [Lymphocyte populations (LP) and Autoantibodies (Ab)] and clinical profile of rheumatoid arthritis (RA) patients who suffered from COVID-19 compared with non-COVID-19 RA patients. Method(s): A nested case-control study of RA patients treated under a strict follow-up model. RA patients and confirmed COVID-19 infection (last 24 months) and RA patients without the infection were included. Subgroups of cases: Long COVID (LC): symptoms after infection for >=4 weeks;Post COVID syndrome (PCS): symptoms for >=12 weeks;and patients with symptoms alpha4 weeks. Sociodemographic, clinical, and paraclinical variables of RA and COVID-19 infection (in cases) were captured. Antinuclear antibodies (ANA), anticardiolipin antibodies, lymphocyte populations (BD FACSDuetTM-BDFACSLyricTMmultiparameter flow cytometry) T cells, B cells, and NK were evaluated. Univariate and bivariate analyzes (STATA 17) were done. Result(s): 300 patients were included (148 cases/152 controls;87.3% women). Median age 59 years (IQR 11). 71.86% were in low disease activity. There were no significant differences in sociodemographic and clinical characteristics between cases and controls. Cases had a time since infection of 18.5 months (IQR 7). Of the total cases, 69%presented LC and 63%PCS.No significant differences were found between cases and controls in the lymphocyte population nor in the antibodies evaluated. There were no differences in the immune profile when comparing patients with LC and PCS with those with symptoms alpha4 weeks after COVID-19 infection. Conclusion(s): No differences were found in the behavior of the immunological profile (independent of symptoms of LC and PCS) in RA patients under strict follow up, evaluated long-term after infection with those who did not have COVID-19. This suggest that patients returned to their baseline homeostatic state, something that has not yet been reported up to now. These results should be replicated in populations with different RA characteristics.

The First Reported Case of Lorlatinib-Associated Diabetic Ketoacidosis

Introduction: Lorlatinib is a third-generation tyrosine kinase inhibitor that inhibits anaplastic lymphoma kinase (ALK) and c-ros oncogene 1 (ROS1). Although 2-10% of patients with non-small cell lung cancer developed hyperglycemia in phase 2 and 3 studies of lorlatinib, only one case has subsequently reported hyperglycemia >500 mg/dL, and no cases of diabetic ketoacidosis (DKA) have been previously reported. Phase 1 trials in neuroblastoma are ongoing. Case Description: A 34-year-old woman with ALK-mutated paraspinal neuroblastoma presented with DKA 14 months after initiation of lorlatinib. Prior to starting lorlatinib, her hemoglobin A1c had been 5.0% (n: < 5.7%). After 12 months of therapy, her A1c increased to 7.8%, prompting the initiation of metformin 500 mg daily. However, two months later she was admitted for DKA with a blood glucose of 591 mg/dL (n: 65-99 mg/dL), CO2 17 mmol/L (n: 20-30 mmol/L), anion gap 18 (n: 8-12), moderate serum ketones, and 3+ ketonuria. Her A1c was 14.8%, C-peptide was 1.2 ng/mL (n: 1.1-4.3 ng/mL), and her glutamic acid decarboxylase-65 and islet antigen-2 autoantibodies were negative. She was also found to be incidentally positive for COVID-19 but was asymptomatic without any oxygen requirement. The patient's DKA was successfully treated with IV insulin infusion, and she was discharged after 3 days with insulin glargine 27 units twice daily and insulin aspart 16 units with meals. One month later, her hemoglobin A1c had improved to 9.4%, and the patient's oncologist discontinued lorlatinib due to sustained remission of her neuroblastoma and her complication of DKA. After stopping lorlatinib, her blood glucose rapidly improved, and she self-discontinued all her insulin in the following 3 weeks. One month later, she was seen in endocrine clinic only taking metformin 500 mg twice daily with fasting and post-prandial blood glucose ranging 86-107 mg/dL. Discussion(s): This is the first reported case of DKA associated with lorlatinib. This case highlights the importance of close glucose monitoring and the risk of severe hyperglycemia and DKA while on lorlatinib therapy. Discontinuation of lorlatinib results in rapid improvement of glycemic control, and glucose-lowering treatments should be promptly deescalated to avoid hypoglycemia.Copyright © 2023

Pbmc Immunophenotyping and Plasma Inflammatory Profile of Children with Long Covid

Background: Long COVID can be developed by individuals after an infection with SARS-CoV-2 as described by the WHO. Although this condition is more commonly described in adults, it can occur in children and adolescents with a wide range of estimated prevalence of 1-25%. Little is known about the role of the immune system in long COVID. However, one of the main hypotheses about the underlying mechanism in long COVID is that there is an immune and inflammatory dysregulation that persists after the acute infection. The objective of this study is to compare immune cells populations, and inflammatory biomarkers in paediatric populations with and without long COVID. Method(s): We analyzed 55 blood samples from the pediaCOVID cohort (Hospital Germans Trias i Pujol), which includes more than 130 children diagnosed with long COVID and 23 controls. We measured different immune cell populations using spectral cytometry with a panel of 37 cellular markers, and 42 inflammatory markers using Luminex or ELISA. EdgeR was used for statistical analysis of the spectral data;p-values of inflammatory markers were calculated using the likelihood ratio test and they were corrected for multiple comparisons. Result(s): The study cohort had a median age of 14.3 (IQR, 12.5-15.2) and 69.1% female. Patients had at least 3 symptoms associated with long COVID (median [IQR];10 [7-16]). The most common symptom was asthenia/fatigue (98.2%). Compared to the control cohort, children with long COVID had increased numbers of CD4+CD8+ T cells, IgA+CD21+CD27+ memory B cells, and IgA+CD21-CD27- memory B cells, while CD4+ TEMRA cells (CD45RA+, CCR7-), intermediate monocytes (CD14+, CD16+) and classical monocytes (CD14+, CD16-) were decreased (all p< 0.05;q=n.s.). None of the 42 inflammatory biomarkers showed significant differences between children with and without long COVID. Conclusion(s): The results of this study suggest that specific populations of peripheral blood immune cells might be involved in the mechanisms underlying prolonged COVID in children and adolescents. The increase in both IgA+CD21-CD27- and IgA+CD21+CD27+ memory B cells could be associated with the persistence of viral antigen in the gut and/or gut dysbiosis. Moreover, the decrease in CD4+ TEMRA cells could be related to autoantibodies against G-protein coupled receptors (GPCRs), since this cell population can express GPR56, and autoantibodies against GPCRs were previously reported to be elevated in adults with long COVID.

New Onset Autoimmune Disease More Common after Covid-19

Background: Viral infections including SARS-CoV-2 may trigger autoimmune disease through T-cell-mediated autoimmune response through molecular mimicry-cross-reactive T-cell recognition or bystander T-cell activation. Autoantibodies have been detected in patients with COVID-19 and some human proteins have homologous regions with SARS-CoV-2 peptides that could function as autoantigens. While there are scattered reports of various autoimmune diseases diagnosed after COVID-19, the risk is not known. Method(s): TriNetX (a global federated health research network providing access to electronic medical records across 72 large healthcare organizations) was utilized to define a cohort of adults 18 years or older seen on or after January 1, 2020 with at least one follow-up visit after an index date. Exposure was defined as COVID-19 diagnosis by ICD10 code or positive laboratory test. Controls did not have COVID-19 (by the same criteria) and were propensity score-matched to patients who had COVID-19 by age and female sex. Index date was the date of COVID-19 diagnosis or first provider visit for any reason during the study period for controls. Outcomes (see table) were assessed starting one month after index date (to exclude prior undiagnosed autoimmune disease) until one year after. Patients with a specific outcome prior to the index date or within one month after the index date were excluded from the analysis for that outcome. Incidence by COVID-19 exposure status and risk ratios for each outcome were assessed. Result(s): 4,016,472 patients were included (2,008,236 in both groups). Overall, mean (SD) age was 49.2 (17.9) and 57.7% were female. Patients who had COVID-19 were more likely to be white (63 vs 56.9%;p< 0.001). Rheumatoid arthritis, psoriasis and type 1 diabetes mellitus had the highest incidence after COVID-19 (0.24, 0.22 and 0.19%, respectively). While the incidence of most of the autoimmune diseases assessed were low in both groups, the risk ratios for all but one condition (Grave's) showed statistically significant higher risk in patients after COVID-19 than in those without COVID-19 (see table). Risk ratios were highest for polyarteritis nodosa (4.43, 3.27-6.01), reactive arthritis (3.56, 2.05-6.2) and ANCA-associated vasculitis (3.36, 2.6-4.34). Conclusion(s): Autoimmune diseases were more likely to be diagnosed within the first year after COVID-19 than in age-, sex-matched controls. Future work will assess the validity of autoantibodies in predicting autoimmune disease after COVID-19. (Table Presented).

New-Onset Thyroid Eye Disease After COVID-19 Infection in Patient with Graves' Disease Post-Radioactive Iodine Treatment

Introduction: Coronavirus disease 2019 (COVID-19) has been associated with dysregulation of the immune system and abnormal thyroid function. The aim of this novel case report is to inform physicians of the possibility that COVID-19 infection may precipitate thyroid eye disease (TED) in patients with Graves' Disease (GD) even after treatment with radioactive iodine (RAI). Case Description: In this report, we describe a patient with GD treated with RAI who developed TED after COVID-19 infection. The patient was initially diagnosed with GD in 2018. A thyroid uptake scan (I-123) was consistent with GD with moderately elevated uptake. She was initially managed with methimazole and atenolol and was eventually treated with RAI (16.32 millicurie I-131) in February 2021. She had post-ablative hypothyroidism managed with levothyroxine. The patient contracted COVID-19 in January 2022. In February 2022, the patient started experiencing eye irritation, dryness, protrusion of eyes, eyelid swelling, and visual disturbances. Thyroid stimulating hormone (TSH) receptor auto-antibodies (7.33 IU/L, normal < /=1.00 IU/L) and thyroid stimulating immunoglobulin (4.30 IU/L, normal < /=1.00 IU/L) were elevated. TSH was normal (2.180 mIU/L, normal 0.270 - 4.200 mIU/L) on levothyroxine 125 mcg daily. She was later diagnosed with TED. Discussion(s): GD is an autoimmune thyroid disorder related to the presence of TSH receptor-stimulating antibodies and is often associated with ocular symptoms. Activation of an autoimmune response during COVID-19 infection, may induce onset or relapse of GD. A study using the national health insurance service database in South Korea noted an increase in the incidence of subacute thyroiditis in 2020 in association with the COVID-19 pandemic. TED is usually seen in patients with GD. Radioactive iodine is widely used in the treatment of GD and has been associated with development or worsening of TED. There are published cases of TED occurring in patients with GD after receiving COVID-19 vaccine. It is thought that the inflammatory syndrome induced by the adjuvants could induce molecular mimicry, which could trigger TED. In most cases this adverse effect was transient, lasting a few months after treatment. There have been case reports of TED occurring after 3 to 21 days of COVID-19 vaccination in patients with controlled GD. Symptoms improved in 4-8 months. Development of TED in patients with GD who have been treated with RAI typically occurs soon after RAI therapy. For TED to occur in a GD patient 11 months after receiving RAI therapy is unusual. COVID-19 infection appears to have been the trigger for this patient's eye disease. This is highly unusual and has not been published to our knowledge.Copyright © 2023

Is COVID-19 severity unrelated to antinuclear antibodies?

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) induces an overreaction of the immune system, resulting in the production of auto-antibodies. Several studies have reported that autoantibodies are prevalent in COVID-19 patients. In our study, antinuclear antibodies were evaluated in patients with COVID-19. We examined 131 sera from patients (>17-year-old) with confirmed COVID-19. Samples were collected prior to receiving any medication and antinuclear antibodies (ANA) levels were measured by the indirect immunofluorescence (IIF) method. Furthermore, the immunoblotting test was used to determine the presence of anti-nuclear antigen antibodies. The IIF-ANA test was positive in 36.4% (48/131) of patients. Overall, non-ICU patients had higher IIF-ANA titers than ICU patients. In particular, ICU patients had fewer nuclear, cytoplasmic, and mitotic IIF-ANA antibodies than non-ICU patients. In conclusion, COVID-19 patients frequently have ANA possibly reflecting the immune dysregulation due to several damaged cells by SARS-CoV-2 virus.

Dermatomyositis with Specific Autoantibodies Appearing after Sars-Cov-2 Infection or Mrna Vaccination: Hints at a Shared Pathogenetic Mechanism

Background. Environmental factors such as infections and vaccines are known to trigger dermatomyositis (DM), and during the recent SARS-CoV-2 pandemic this has become even clearer. SARS-CoV-2 infection may share features with anti-MDA5 DM, such as rapidly progressive lung involvement, cutaneous lesions and cytokine release syndrome. A few case reports of DM following SARSCoV-2 vaccination have been published, suggesting the onset of an aberrant immune response leading to DM with specific autoantibody signatures and severe organ impairment. Methods. Clinical and laboratory data of the 2 case reports were obtained from electronic clinical charts in Humanitas Research Hospital (Rozzano, Milan, Italy). Autoantibody analysis was performed by protein-immunoprecipitation for anti-MDA5 and immunoblot for anti-Ro52 and TIF1gamma antibodies as per protocol. Results. Case report 1 is a 71-year-old woman who developed fever, cough, and anosmia, which resolved spontaneously in two weeks, but did not undergo a nasopharyngeal swab, while her relatives were diagnosed with SARS-CoV-2 infection. When symptoms improved, she developed arthralgia and skin lesions on her face, chest, and hands for which she started topical treatment, with negative SARSCoV-2 nasopharyngeal swab and positive serum test for IgG against SARS-CoV-2 spike protein. For the persistence of the skin rash and arthralgia, she was admitted to our Department in March 2021. Blood tests showed mild elevation of C reactive protein (2.1 mg/L -normal value NV<5), aspartate (84 UI/L) and alanine aminotransferase (133 UI/L -NV<35), ferritin (595 ng/ml -NV<306), troponin I (19 ng/L -NV<14), and BNP (251 pg/ml -NV<100) with normal complete blood cell count, creatine kinase, C3 and C4. IgG antibodies for SARS-CoV-2 spike protein were confirmed to be elevated (96 AU/ml -NV<15). Autoantibodies associated with connective tissue diseases were tested and only anti-MDA5 antibodies were positive at immunoprecipitation. A punch biopsy of a Gottron-like lesion on the left hand showed leukocytoclastic vasculitis. We observed reduced capillary density with neoangiogenesis and ectasic capillaries at the nailfold capillaroscopy. EKG and ecocardiography were normal, while cardiac magnetic resonance detected abnormalities in the parametric sequences, consistent with signs of previous myocarditis. A lung CT scan revealed pulmonary emphysema while respiratory function tests demonstrated reduced volumes (FVC 82%, FEV1 64%, inadequate compliance CO diffusion test). Based on the biochemical and clinical findings, a diagnosis of anti-MDA5-associated DM with skin and heart involvement was made and treatment with low-dose methylprednisolone (0.25 mg/kg daily) and azathioprine 100 mg was started, then switched to mycophenolate because not effective on skin lesions. Case report 2 is an 84-year-old woman with history of colon cancer (surgical treatment) and oral lichen treated with low doses steroids in the last 2 years. After the 2nd dose of SARS-CoV-2 mRNA vaccination, in March 2021 she developed skin rash with V-sign, Gottron's papules, periungueal ulcers, muscle weakness and fatigue, thus she performed a rheumatologic evaluation. Blood tests showed mild elevation of creatine kinase (484 UI/L, NV <167), CK-MB (9.6ng/ml, NV <3.4), BNP (215 pg/ml -NV<100) with normal values of complete blood cell count, C3 and C4. Anti-Ro52kDa and TIF1gamma were positive at immunoblot, thus we confirmed a diagnosis of DM. The clinical evaluation also showed active scleroderma pattern at nailfold capillaroscopy, normal echocardiography, bronchiectasia but not interstitial lung disease at lung CT, and normal respiratory function tests (FVC 99%, FEV1 99%, DLCO 63%, DLCO/VA 81%). A PET-CT scan was performed to exclude paraneoplastic DM, and treatment with steroids and mycophenolate was started. Conclusions. SARS-CoV-2 may induce mechanisms for escaping the innate immunity surveillance and causing autoimmune diseases, but more clinical and functional studies are needed to demonstrate this possible association.

Could SARS-CoV-2 Trigger the Formation of Antinuclear Antibodies?

Objective: The effect of severe acute respiratory syndrome-coronavirus-2 (SARS-CoV-2) infection on autoimmunity in both disease and post-disease stages has not been fully explained. There is not enough information about the evaluation of autoimmune antibodies in convalescent SARS-CoV-2 patients. This study aimed to investigate the presence and types of autoantibodies in post-illness coronavirus disease-2019 (COVID-19) patients and to compare them with indirect immunofluorescence assay (IIF)-antinuclear antibody (ANA) results before SARS-CoV-2 infection. Material(s) and Method(s): Twenty-four COVID-19 patients with known and reported ANA test results prior to SARS-CoV-2 infection were included in this study. Patients' IIF-ANA, extractable nuclear antigen blot and anti-dsDNA tests were studied three and nine months after SARS-CoV-2 infection. Result(s): Three months after SARS-CoV-2 infection, 41.66% of patients had a positive IIF-ANA test. When we compared these results with pre-infection ANA results, 3 patients (12.5%) were variable. The first case was chromosomal granular positive before infection and was found to be homogeneous, and cytoplasm was speckled positive after infection. Additionally, Scl-70, DFS70, and anti-dsDNA were found to be positive. We think that lupus symptoms were triggered after COVID-19. The second case had negative ANA before infection, while the ANA was antinuclear membrane positive (2+) three months after infection. Also, anti-RNP/Sm was detected as positive. The third case had negative ANA before infection, and was detected to have speckled weakly positive ANA three months after infection. However, autoantibody positivity was not detected. Conclusion(s): As a result, these data support the idea that SARS-CoV-2 infection may trigger autoimmunity and be associated with the development of autoantibodies.Copyright © 2022 by the Turkish Society of Immunology. Turkish Journal of Immunology published by Galenos Publishing House.

4th Global Conference on Myositis, GCOM

The proceedings contain 255 papers. The topics discussed include: molecular pathology of mitochondrial disorders;defining causal genes at MHC in SLE - implications for myositis and other diseases that share MHC risk;role of mitochondria in skeletal muscle dysfunction in myositis;selective T cell depletion for inclusion body myositis: why and how;inclusion body myositis in 2022: from physiopathogenesis to clinical trials;autoantibodies and complement in experimental IMNM: from pathogenesis to therapy?;reliability of immunoassays for myositis autoantibodies;when JM patients lose their 'J': transition challenges in myositis car;fatigue and well-being of children with chronic inflammatory disease;physical fitness in long-term JDM;Eular Covid and COVAX registries' update: focus on myositis;and outcomes, biomarkers, and novel treatments for the skin in dermatomyositis.