Lymphoproliferation- a clue towards an underlying monogenic disorder of immune dysregulation- a retrospective analysis from a single center in India

Background: Lymphoproliferation is the persistent proliferation of lymphoid cells and it's incidence in inborn errors of immunity varies from 0.7 to 18%. Material(s) and Method(s): This is a retrospective analysis of patients referred to the department of Immunology, B. J. Wadia Hospital for Children, Mumbai between March 2017 to December 2022. Inclusion criteria consisted of 3 months duration of significant lymphadenopathy and/or splenomegaly or history of lymphoma. The clinical characteristics, laboratory and molecular findings of the included patients were analyzed. Result(s): A total of 66 patients were included. There was a male preponderance with male:female ratio of 25:8. Median age of onset of lymphoproliferation was 4.75 years(Range 1 year to 60 years). Splenomegaly was seen in 75%. Infections included recurrent pneumonia (14/66), recurrent ear infections(5/66), COVID(4/66), one episode of pneumonia(6/66), herpes zoster(3/66), recurrent subcutaneous abscess (3/66), abdominal koch(3/66), chronic sinusitis(2/66), dermatophytosis(2/66), esophageal candidiasis(2/66), recurrent malaria(1/66), recurrent varicella(1/66), cryptococcal meningitis(1/66), gram negative sepsis(1/66), BCG adenitis(1/66), pseudomonas osteomyelitis(1/66), impetigo (1/66), pseudomonas urinary tract infection (1/66), chicken pox(1/66), herpes keratitis(1/66), dengue(1/66), Other manifestations included Evans plus phenotype(10/66), Evans phenotype(8/66), Autoimmune hemolytic anemia(5/66), bronchiectasis(5/66), Type 1 diabetes(3/66), hyper reactive airway disease(2/66), inflammatory bowel disease(4/66), autoimmune thrombocytopenia(2/66), stroke(3/66), hemophagocytic lymphohistiocytosis(2/66), hypertriglyceridemia(2/66), hypothyroidism(2/66), celiac disease(1/66), Type 2 diabetes(1/66), autoimmune encephalitis(1/66), autoimmune hepatitis(2/66), anti-parietal cell antibody(1/66), arthritis(1/66), autoimmune enteropathy(1/66), systemic lupus erythromatosus(1/66), primary biliary cirrhosis requiring liver transplant(1/66), nephrotic syndrome(1/66), lymphoedema(1/66), hypersplenism(1/66), recurrent oral ulcers(1/66), gout(1/66), dermatitis(1/66), ovarian teratoma(1/66), alopecia areata(1/66). Hodgkin's lymphoma(HL) was the most common malignancy(9/66), followed by non Hodgkin lymphoma(NHL)(6/66), transformation from NHL to HL(1/66), Burkitt to T-cell lymphoma(1/66), HL to DLBCL(1/66), HL to anaplastic T-cell lymphoma(1/66). EBV driven lymphoproliferation was seen in biopsy of21/66. Genetic testing showed mutations in LRBA(11/66), PIK3CD(5/66), CTLA4(3/66), TET2(2/66), IL2RA (1/66), IL12RB1(1/66), BACH2(1/66), PRKCD(1/66), TNFSFR13B(1/66), TNFAIP3(1/66), FAS(2/66), FASL(1/66), Caspase8(1/66), CARD11(1/66), RTEL1(1/66), AICD(1/66), PIK3R1(1/66), IKBKB(1/66). Treatment included IVIG, chemotherapy, rituximab, sirolimus, abatacept, HSCT. Conclusion(s): All children with persistent lymphoproliferation, with or without autoimmunity and/or infections should be worked up for an underlying monogenic disorder of immune dysregulation. Lymphomas presenting at abnormal site and/or age, relapse and EBV driven lymphomas require further evaluation. Presence of monogenic cause helps in providing targeted therapy.Copyright © 2023 Elsevier Inc.

SARS-CoV-2 VACCINE-INDUCED T CELL RESPONSES IN CANCER PATIENTS UNDERGOING TREATMENT

Background: mRNA vaccines have proven useful in protecting vulnerable populations against SARS-CoV-2 infection. However, certain therapeutics, specifically those used in cancer treatment, reduce mRNA vaccine-induced humoral responses against SARS-CoV-2. The effects on T cell responses are not well characterized. Here, we evaluate SARS-CoV-2 spike-specific T cell responses over the course of one year in solid tumor patients in BC, Canada. Method(s): 18 female, solid-tumor patients from the BC Cancer Agency were enrolled in this prospective, cohort study, with 7 patients receiving cytotoxic chemotherapy and 11 patients receiving non-cytotoxic treatments. Whole blood was collected 1-month (T1) and one-year +/- 1-month (T2) post series completion (2 mRNA doses). Antigen-induced marker assays (AIM assays) were used to quantify CD4+ and CD8+ T cell responses, where whole blood was stimulated with ancestral or omicron SARS-CoV2 Spike peptide pools or unstimulated for 48 hours at 37degree C, fluorescently stained for activation markers CD25 and OX40 (CD4+ T cells) or CD69 and CD137 (CD8+ T cells), and analyzed using a 5-laser flow cytometer. Phenotyping of antigen-specific CD4+ T cells was done in parallel to assess the frequency of spike-specific Tregs, Th1, Th2, Th9, Th17, and Th17.1 cells. Result(s): All individuals had detectable levels of spike-specific CD4+ T cells at T2, while only 72.2% of individuals had detectable levels of spike-specific CD8+ T cells. Treatment type did not significantly impact the magnitude or phenotype of T cell responses, including those to Omicron. However, increased age was associated with decreased ancestral CD8+ T cell responses at T2. Further, ancestral and omicron responses were significantly different at T2, with decreased magnitude and altered phenotype of omicron-specific CD4+ T cells. Conclusion(s): Here, we report that solid tumor patients, treated with either chemotherapy or biologics, mount robust T cell immunity to SARS-CoV-2 following vaccination. Additional data is needed to determine if these responses correlate with antibody levels and clinical illness.

Hairy Cell Leukemia in a Filipino Male during the COVID-19 Pandemic - Report of a Rare Case

Hairy cell leukemia (HCL) is a rare, chronic, mature B-cell lymphoproliferative disorder accounting for 2% of all leukemias. In this paper, we would like to present our experience in the management of HCL in a financially limited setting where other diagnostic tests and chemotherapy are unavailable. The case report aims to emphasize the recognition of the distinctive morphology of hairy cells in the peripheral blood in the consideration of the initial diagnosis. A 60-year-old Filipino male was incidentally found to have anemia, thrombocytopenia and an absolute neutrophilic count below 1,000 in a pre-operative clearance for elective herniorrhaphy. Blood smear revealed atypical lymphocytes with hair like cytoplasmic projections. CT-scan of the abdomen showed splenomegaly and prominent paraaortic nodes. Flow cytometry of the bone marrow aspirate was consistent with an involvement of a Mature B cell neoplasm markers CD19, CD20, CD22 and surface immunoglobulin lambda and hairy cell leukemia markers CD11c, CD103 and CD25. He responded to six-weekly sessions of Cladribine with remission of the bone marrow and hematologic parameters. HCL is a rare type of a mature B cell neoplasm characterized by pancytopenia, splenomegaly, bone marrow fibrosis and the presence of atypical lymphoid cells with hairy projections in blood, bone marrow and spleen. Immunophenotyping express CD11c, CD103, CD123, and CD25. BRAF V600E mutation is the disease defining genetic event. Cladribine and Pentostatin are the first line of treatment. Cases of leukemia can be easily overlooked because of the mild derangement in the complete blood count. A meticulous differential review of the atypical lymphocyte, is the first step in the diagnosis of this rare disease.Copyright © 2022, Philippine College of Physicians. All rights reserved.

Differential peripheral blood mononuclear cell reactivity against SARS-CoV-2 proteins in naive and previously infected subjects following COVID-19 vaccination

The decline in vaccine efficacy and the risk of reinfection by SARS-CoV-2 make new studies important to better characterize the immune response against the virus and its components. Here, we investigated the pattern of activation of T-cells and the expression of inflammatory factors by PBMCs obtained from naive and previously infected subjects following COVID-19 vaccination, after PBMCs stimulation with S1, RBD, and N-RBD SARS-CoV-2 proteins. PBMCs showed low levels of ACE2 and TMPRSS2 transcripts, which were not modulated by the exposure of these cells to SARS-CoV-2 proteins. Compared to S1 and RBD, N-RBD stimulation showed a greater ability to stimulate T-cell reactivity, according to CD25 and CD69 markers. Interestingly, T-cell reactivity was more pronounced in vaccinated subjects with prior SARS-CoV-2 infection than in vaccinated donors who never had been diagnosed with COVID-19. Finally, N-RBD stimulation promoted greater expression of IL-6 and IFN-gamma in PBMCs, which reinforces the greater immunogenic potential of this protein in the vaccinated subjects. These data suggest that PBMCs from previously infected and vaccinated subjects are more reactive than those derived from just vaccinated donors. Moreover, the N-RBD together viral proteins showed a greater stimulatory capacity than S1 and RBD viral proteins.Copyright © 2022

Severe lymphocyte dysfunction and impaired cellular immune response as a consequence of COVID-19

Introduction: A correct diagnosis of long-term complications in COVID-19 convalescents is of great importance for their therapeutic management. Aim(s): To identify long-term post-COVID-19 damages related to lymphocytes T to search prevention and treatments in future. Material(s) and Method(s): Data from 10 healthy volunteers, 23 patients (moderate/severe) COVID-19 were analysed. Responses of lymphocytes to LPS, ConA, CD3/CD28 mitogens and spike protein were measured in all groups during acute phase, 3- and 6-months post-hospitalization. Main lymphocyte T subpopulations (CD4, CD8) and CD25 expression on the lymphocyte subpopulations were also examined. Result(s): 6 of 13 severe patients were ventilated mechanically. The remaining 6 patients improved after high-flow nasal oxygen therapy and were discharged with normalized respiratory functions, the same as the mild/moderate COVID-19 group. We found that the decrease in total lymphocytes is equally due to the decrease in CD4 and CD8 cells. These changes persist in moderate and severely ill groups over 3 months post-COVID-19. In the case of CD4 lymphocytes, the changes concerned both naive (CD4+/CD25-) and activated (CD4+/CD25+) cells. A significant decrease in the CD8 cells was also observed, but only for the absolute number of naive CD8 lymphocytes (CD8+/CD25-). During the acute COVID-19 a significant impaired responses to ConA and CD3/CD28 were found. No difference in spike protein response was noticed. Conclusion(s): Long-term reductions (3- and 6-months) in the total count of lymphocytes in subpopulations and proliferative responses to mitogens suggest a severe impairment of the cellular immune response.

POST-GAM-COVID-VAC COMBINED VECTOR VACCINE CELLULAR AND HUMORAL IMMUNE RESPONSE.

Currently, as the SARS-CoV-2 pandemic evolves, there has been increasingly more attention paid to building natural and vaccine-induced immunity against SARS-CoV-2 and related disease known as COVID-19. Widespread preventive vaccination plays an important role in effectively protecting people from viral infections and can reduce national economic costs. Purpose - to study peripheral blood cell subset composition and magnitude of humoral response in vaccinated Gam-COVID-Vac subjects. The prospective study included 352 patients, of which 194 (119 women and 75 men) underwent an immunogram study and assessed level of anti-SARS-CoV-2 antibodies. In patients, the study of the lymphocyte subset composition and estimation of anti-SARS-CoV-2 antibodies was carried out at two time points - prior to vaccination and 90 days after inoculated component 1 of the Gam-COVID-Vac vaccine. In general, vaccination was well tolerated by patients, with no serious adverse events after immunization. The reaction to the vaccine (fever, malaise, headache, local reactions) was short-term (1-2 days) and more often noted after inoculated vaccine component 2. Comparatively analyzed immunogram parameters in females before and after vaccination revealed increased relative level of T-lymphocytes (CD3+), T-helper cell subset (CD3+CD4+), increased absolute and relative level of activated CD3+CD25+ T-lymphocytes, but decreased absolute and relative level of natural killer (CD3-CD56+CD16+) and natural killer T-cell (CD3+CD56+CD16+) cell subsets as well as decreased CD147 receptor expression on T-lymphocytes. Similar patterns were also found while examining the immunogram in males exepting increased level of lymphocytes and lowered CD147 expression on both T- and B-lymphocytes. No changes in the parameters of the immune T-cell arm was found. The high efficacy of the vaccine was confirmed by development of SARS-CoV-2-specific class G antiviral antibodies in 97.5% and 92.3% of vaccinated females and males, respectively. The data obtained evidence that: 1) vaccination induces a specific humoral immune response determined three months post-vaccination, and 2) it caused no serious disturbances in the immune system functioning, which could be reflected in the peripheral blood lymphocyte subset composition. Thus, the data presented allow to conclude that Gam-COVID-Vac is effective vaccine against SARS-CoV-2 infection.Copyright © 2022 Saint Petersburg Pasteur Institute. All rights reserved.

POST-GAM-COVID-VAC COMBINED VECTOR VACCINE CELLULAR AND HUMORAL IMMUNE RESPONSE

Currently, as the SARS-CoV-2 pandemic evolves, there has been increasingly more attention paid to building natural and vaccine-induced immunity against SARS-CoV-2 and related disease known as COVID-19. Widespread preventive vaccination plays an important role in effectively protecting people from viral infections and can reduce national economic costs. Purpose - to study peripheral blood cell subset composition and magnitude of humoral response in vaccinated Gam-COVID-Vac subjects. The prospective study included 352 patients, of which 194 (119 women and 75 men) underwent an immunogram study and assessed level of anti-SARS-CoV-2 antibodies. In patients, the study of the lymphocyte subset composition and estimation of anti-SARS-CoV-2 antibodies was carried out at two time points - prior to vaccination and 90 days after inoculated component 1 of the Gam-COVID-Vac vaccine. In general, vaccination was well tolerated by patients, with no serious adverse events after immunization. The reaction to the vaccine (fever, malaise, headache, local reactions) was short-term (1-2 days) and more often noted after inoculated vaccine component 2. Comparatively analyzed immunogram parameters in females before and after vaccination revealed increased relative level of T-lymphocytes (CD3+), T-helper cell subset (CD3+CD4+), increased absolute and relative level of activated CD3+CD25+ T-lymphocytes, but decreased absolute and relative level of natural killer (CD3-CD56+CD16+) and natural killer T-cell (CD3+CD56+CD16+) cell subsets as well as decreased CD147 receptor expression on T-lymphocytes. Similar patterns were also found while examining the immunogram in males exepting increased level of lymphocytes and lowered CD147 expression on both T- and B-lymphocytes. No changes in the parameters of the immune T-cell arm was found. The high efficacy of the vaccine was confirmed by development of SARS-CoV-2-specific class G antiviral antibodies in 97.5% and 92.3% of vaccinated females and males, respectively. The data obtained evidence that: 1) vaccination induces a specific humoral immune response determined three months post-vaccination, and 2) it caused no serious disturbances in the immune system functioning, which could be reflected in the peripheral blood lymphocyte subset composition. Thus, the data presented allow to conclude that Gam-COVID-Vac is effective vaccine against SARS-CoV-2 infection.

EFFECT OF COVID-19 VACCINATION ON THE IMMUNE STATUS AND AUTOANTIBODY PROFILE IN WOMEN OF REPRODUCTIVE AGE.

In the context of the COVID-19 pandemic, scientific interest is growing in studying the impact of the proposed vaccination on women's reproductive health. As is known, alterations in the state of the immune system and activation of an autoimmune response can lead to reproductive failure in women and potential complications of subsequent pregnancy. Objective(s): to evaluate the effect of the "Gam-COVID-Vac" on the immune status parameters, the relationship of their changes and the specific immune response to vaccination with the dynamics of the level of autoantibodies in women of reproductive age. The prospective study included 120 women who were vaccinated against COVID-19 with the "Gam-COVID-Vac". The criteria for inclusion in the study were: the age from 18 to 49 years, the absence of COVID-19 in the anamnesis, a negative result of a study on SARS-CoV-2 by PCR and negative results of tests for antibodies of classes G and M to SARS-CoV-2 before vaccination, the absence of pregnancy and serious somatic diseases. The patients were examined twice: immediately before vaccination and 90-100 days after the introduction of the 1st component of the vaccine. The level of IgG antibodies to SARS-CoV-2 after vaccination was assessed using ELISA. Before and after vaccination, the levels of antiphospholipid, anti-nuclear, organ-specific and antihormonal autoantibodies were determined, peripheral blood lymphocytes were immunophenotyped to determine the main subpopulations (CD3, CD4, CD8, CD19, CD5, CD16, CD56), as well as the expression of activation markers of lymphocytes (HLA-DR, CD25, CD147) using monoclonal antibodies. The effectiveness and safety of the combined vector vaccine against COVID-19 were high. Specific IgG antibodies to SARS-CoV-2 were produced in 98.3% of vaccinated women, no serious adverse reactions were observed. After vaccination, there was an increase in the level of some autoantibodies within the reference ranges, only IgM antibodies to phosphatidylethanolamine (PE) and IgG antibodies to DNA increased above the reference values. However, this increase was transient. After vaccination, the following changes in the parameters of the immunogram were observed: an increase in the content of cells with CD3+CD25+, CD19+ phenotype in peripheral blood and a decrease in the content of cells with CD56+CD16+ phenotype within the reference ranges, a decrease in CD147+/CD3+. Weak correlations were noted between these changes in immunogram parameters and the levels of some autoantibodies. The specific antiviral immune response to vaccination did not correlate with the autoimmune response. Vaccination with "Gam-COVID-Vac" is effective and safe and does not lead to disorders in the immune system. The observed increase in the level of autoantibodies to PE and DNA is transient. Changes in the parameters of the immune status within the reference ranges may be due to vaccination and the development of a specific antiviral immune response. Copyright © 2022, SPb RAACI.

A Single Center Experience of Simultaneous Heart-Kidney Transplantation

Purpose: Currently there are no UNOS guidelines regarding the selection criteria required for simultaneous heart-kidney transplant recipients (SHKT). As of 2018 our center has begun performing these dual transplants for appropriate candidates. We report on the criteria devised to guide SHKT candidate selection at our institution and the subsequent clinical outcomes. Method(s): This is a single center, retrospective study of 26 patients who received SHKT at our institution from Dec 2018 to Oct 2021. A multidisciplinary team composed of heart and kidney transplant medical and surgical members determined appropriate recipient-donor SHKT candidate pairs. Selection criteria for SHKT was established by our kidney transplant group and included an evaluation for chronic kidney disease (CKD) or evidence of acute kidney injury (AKI) with a prolonged course or requiring renal replacement therapy (RRT). The surgery was conducted according to our institution's standardized protocols. The majority of patients received IL2-RA and methylprednisolone induction therapy, and all patients received triple immunosuppression therapy with prednisone, mycophenolate mofetil and tacrolimus. Adjustments in long term therapy were made in collaboration between the heart and kidney transplant teams. Result(s): From Dec 2018 to Oct 2021, 26 patients underwent SHKT at our institution. 24 patients (92%) carried a diagnosis of chronic kidney disease (CKD) as defined as an eGFR <60 ml/min/1.73m2 for at least 90 days on at least two separate tests. Clinical risk factors for CKD, the presence of proteinuria, and renal imaging data were also taken into consideration when determining a diagnosis of CKD. Two patients (8%) carried a diagnosis of stage III AKI for at least 4 weeks and required renal replacement therapy during their hospital course. Of our 26 patients, one patient received a DCD donor and 12 patients (46%) received hepatitis C donors. 25 patients (96%) received induction therapy with IL2-RA. During the first 3 months post-transplant, the only patient who received ATG had 7 severe infections;11 patients (44%) and 13 patients (52%) who received IL2 -RA had no infections and <= 4 mild infections, respectively. One patient died due to COVID 19 pneumonia complicated by multisystem organ failure. For a median follow up period of 410 (187-707) days, 8% patients in the IL2-RA induction cohort experienced a 2R/3A heart rejection, 8% patients remained on HD due to primary kidney graft nonfunction, and the survival rate was 96%. Conclusion(s): UNOS guidelines regarding selection criteria for SHKT are an important next step in the care of heart transplant candidates with kidney disease, particularly as the number of SHKT performed yearly increase. Compared to the literature, our data supports the use of standardized criteria for SHKT selection and the use of IL2- RA as an induction strategy with excellent patient survival.

Higher Proinflammatory Cytokines Are Associated with Increased Antibody Titer After a Third Dose of SARS-CoV-2 Vaccine in Solid Organ Transplant Recipients

Purpose: Solid organ transplant recipients (SOTRs) are at increased risk for severe COVID-19 and exhibit lower antibody responses to SARS-CoV-2 vaccines. This study aimed to determine if pre-vaccination cytokine levels are associated with antibody response to SARS-CoV-2 vaccination. Method(s): A cross-sectional study was performed among 58 SOTRs before and after two-dose mRNA vaccine series, 35 additional SOTRs before and after a third vaccine dose, with comparison to 16 healthy controls (HCs). Anti-spike antibody was assessed using the IgG Euroimmun ELISA. Electrochemiluminescence detectionbased multiplexed sandwich immunoassays were used to quantify plasma cytokine and chemokine concentrations (n=20 analytes). Concentrations between SOTRs and HCs, stratified by ultimate antibody response to the vaccine, were compared using Wilcoxon-rank-sum test with false discovery rates (FDR) computed to correct for multiple comparisons. Result(s): In the study population, 100% of HCs, 59% of SOTRs after two doses and 63% of SOTRs after three doses had a detectable antibody response. Multiple baseline cytokines were elevated in SOTRs versus HCs. There was no significant difference in cytokine levels between SOTRs with high vs low-titer antibodies after two doses of vaccine. However, as compared to poor antibody responders, SOTRs who went on to develop a high-titer antibody response to a third dose of vaccine had significantly higher pre-third dose levels of several innate immune cytokines including IL-17, IL-2Ra, IL-6, IP-10, MIP-1alpha, and TNF-alpha (FDR <0.05). Conclusion(s): A specific inflammatory profile or immune state may identify which SOTRs are likely to develop stronger sero-response and possible protection after a third dose of SARS-CoV-2 vaccine.

INFLAMMATION OVERDRIVE: A CONFIRMED DIAGNOSIS OF HEMOPHAGOCYTIC LYMPHOHISTIOCYTOSIS RESULTING FROM COVID-19

SESSION TITLE: Unique Inflammatory and Autoimmune Complications of COVID-19 Infections SESSION TYPE: Rapid Fire Case Reports PRESENTED ON: 10/19/2022 12:45 pm - 1:45 pm INTRODUCTION: Coronavirus disease 2019 (COVID-19) can manifest as a severe immunologic syndrome known as hemophagocytic lymphohistiocytosis (HLH). HLH is a hyper-inflammatory state with a lethal mortality rate, especially when discovered late in the disease process. The optimal timely approach to diagnosis and treatment of secondary HLH in COVID-19 is unclear, however, risk stratification with Hscore using biomarkers can be useful to increase confidence in an HLH diagnosis. CASE PRESENTATION: A 36-year-old morbidly obese male with a history of well controlled mild intermittent asthma presented to the hospital complaining of a one week history of dyspnea and cough after failing outpatient COVID-19 treatment. Upon arrival, he was hypoxic on room air and was placed on non-invasive ventilation. He unfortunately decompensated further and was transferred to the intensive care unit where he was intubated for severe hypoxia and increased work of breathing. His course was complicated by superimposed bacterial pneumonia, vasopressor dependent septic shock, and anuric acute kidney injury requiring continuous renal replacement therapy. He remained profoundly hypoxic despite rescue therapy with multiple sessions of prone ventilation. On hospital day seventeen his platelets declined acutely and a serotonin release assay confirmed heparin-induced thrombocytopenia. His clinical status remained tenuous into the third week of admission. Notably, he developed persistent fever with associated bicytopenia and elevated lactate dehydrogenase, D-dimer, fibrinogen, triglycerides, and aspartate aminotransferase. His calculated Hscore was 189. Hematology recommended initiating HLH therapy with daily dexamethasone and etoposide, however the latter was held due to the patient's rapid hemodynamic decline. The patient succumbed to illness after a twenty-day hospitalization. His HLH was confirmed with a positive postmortem soluble-IL-2-receptor test. DISCUSSION: Proposals of routine HLH screening in critically ill patients are endorsed to promote early detection of this morbid condition. Calculating Hscore using vital signs, imaging, laboratory tests, and patient history can guide suspicion of diagnosis, since HLH-specific markers are often not feasible. Hscores more than 169 correspond to 93% sensitivity and 86% specificity in HLH diagnosis. Immunosuppression is standard therapy with hematology guidance due to the complex pathophysiology and limited research. CONCLUSIONS: This case emphasizes the importance of understanding the relationship between COVID-19 and secondary HLH. A timely diagnosis is vital in order to attempt to effectively treat a syndrome that carries a 65% mortality rate. Reference #1: Dimopoulos G, Mast Q. de, Markou N, et al. Favorable Anakinra responses in severe COVID-19 patients with secondary hemophagocytic lymphohistiocytosis. Cell Host Microbe 2020;doi: 10.1016/j.chom.2020.05.007. PubMed PMID: 32411313. Reference #2: Bauchmuller K, Manson JJ, Tattersall R, et al. Haemophagocytic lymphohistiocytosis in adult critical care. J Intensive Care Soc 2020;21:256–68. Reference #3: Schnaubelt, Sebastian MDa,∗;Tihanyi, Daniel MDb;Strassl, Robert MDc;Schmidt, Ralf MDc;Anders, Sonja MDb;Laggner, Anton N. MDa;Agis, Hermine MDd;Domanovits, Hans MDa Hemophagocytic lymphohistiocytosis in COVID-19, Medicine: March 26, 2021 - Volume 100 - Issue 12 - p e25170 doi: 10.1097/MD.0000000000025170 DISCLOSURES: No relevant relationships by Kristina Catania No relevant relationships by Katie Kennedy No relevant relationships by Josef Kinderwater No relevant relationships by MaryKate Kratzer no disclosure submitted for Ogugua Obi;

Hemophagocytic Lymphohistiocytosis after COVID-19 Vaccination: A Case Report

Aim and background: Cases of thrombotic thrombocytopenia induced by coronavirus disease 2019 (COVID-19) vaccines have been reported recently. Herein, we describe hemophagocytic lymphohistiocytosis (HLH) following COVID-19 vaccination. Case report: A 35-year-old male, chronic alcoholic, 3 years into abstinence received first dose Covishield vaccine. He started developing a fever, testicular pain, diminished sensorium requiring invasive ventilation, and decreased urine output 4 days after getting vaccinated. Initial workup for NCCT brain and HRCT chest was normal, tropical fever panel was negative, cultures for blood and endotracheal aspirate were sterile, liver and renal functions showed mild derangement, CSF study was normal. Ultrasound examination of the abdomen revealed mild hepatosplenomegaly, mild testicular swelling, and suprainguinal lymphadenopathy, with no focus of infection. Subsequently, he developed bicytopenia with haemoglobin 9.0 g/dL and platelet counts 50 × 109/L, ferritin 2130 μg/L, triglyceride 353 mg/dL, and decreased fibrinogen 1.41 g/L. Bone marrow as well as lymph node biopsy showed haemophagocytosis with engulfment of neutrophils, lymphocytes, and normoblasts making HLH a likely diagnosis. Soluble CD25 and NK cell function could not be performed. Extensive evaluation was done to look into the etiology of HLH. SARS-CoV-2 reverse transcriptase-polymerase chain reaction (RT-PCR) test was negative. RT-PCR test for Epstein-Barr virus (EBV), influenza A (H1N1, H3N2), influenza B, cytomegalovirus (CMV) performed from endotracheal aspirate (ETA) was negative. Similarly, the RT-PCR test from serum samples for EBV, Parvo B-19, CMV, and from CSF sample for EBV, Parvo B-19, CMV, and HSV-1 was negative. Hepatitis B, C, and HIV serologies were negative. Culture and sensitivity repeated from blood, ETA and urine was sterile. Autoimmune panel including complements levels were negative. Peripheral smear, bone marrow, and lymph node biopsy were normal and did not reveal abnormal or malignant cells. He had persistent fevers to 38.6°C during the first 6 days of his admission, with a rise in his ferritin to 1950 μg/L. The patient received steroids but not etoposide. By the 8th day, his fevers resolved, with improvement in his lethargy and malaise. Two weeks later, his ferritin had reduced to 510 μg/L, platelet count rose to 180 × 109/L, and repeat ultrasound abdomen demonstrated resolution of his splenomegaly. In our patient, there was no clear precipitant of HLH other than the Covishield vaccine. There was no evidence of an infection or malignancy. Due to our patient's clinical stability, resolution of symptoms, and improvement of HLH parameters he did not require HLH specific therapy. It is unclear if he had a pre-existing genetic predisposition to HLH as genetic testing is pending, however, it is unlikely as he has reached the age of 35 and suffered from previous viral infections without developing HLH.

Immunotherapy: ALLOGENEIC USE OF TREG CELLS OBTAINED FROM PEDIATRIC THYMIC TISSUE (THYTREG) AS A CELLULAR THERAPY TO SUPPRESS EXACERBATED IMMUNE RESPONSES

Background & Aim: Due to its immunomodulatory potential, therapy based on the transfer of regulatory T cells (Tregs) has acquired great interest in the treatment of diseases in which it is necessary to restore immune homeostasis. Until now, autologous Treg cell therapy has proven to be safe, but the employment of blood as the source of Treg presents several limitations in terms of Treg recovery and the quality of the employed Tregs. Our group has developed a new technology to produce massive amounts of GMP Treg derived from the pediatric thymic tissue discarded in pediatric cardiac surgeries (thyTreg) that could overcome the main obstacles. Indeed, we are employing thyTreg cells with success in a clinical trial as autologous cell therapy in transplanted children. Given the large amounts of thyTreg that can be obtained from a single thymus, the main objective of this work is to evaluate the immunogenicity of thyTreg and confirm that its immature phenotype makes possible the allogeneic use of this cellular therapy in order to treat a range of immune diseases and patients. Methods, Results & Conclusion: The thyTreg obtained in the laboratory using the protocol developed by our group exhibit high viability (>90%) and high purity (>80%) in terms of CD25+FoxP3+ expression. ThyTreg have been observed to express low levels of immunogenicity markers (CD40L, CD80, CD86) by flow cytometry. Moreover, in vitro models of thyTreg co-culture with allogeneic peripheral blood mononuclear cells (PBMC) from healthy donors have been performed to i) determine if thyTreg generate an immunogenic response on PBMC, and ii) evaluate the capacity of thyTreg to suppress the proliferation of allogeneic PBMC. Even that the HLA disparity in the allogeneic cocultures between thyTreg and PBMC was high (13 of the 21 typed pairs had HLA <4/12 concordance), thyTreg did not induce the expression of activation markers (CD25, CD69) nor the proliferation or the production of pro-inflammatory cytokines (IFN-g) by allogeneic PBMCs. Moreover, thyTreg greatly inhibit the proliferation of allogeneic CD4 and CD8 T cells, reaching levels of around 70% inhibition of proliferation at a 1: 1 ratio. The results suggest that allogenic thyTreg are not immunogenic and are capable of exerting their suppressive function in an allogeneic context, indicating their possible off-the-shelf use as a treatment for transplant rejection, graft-versus- host disease, autoimmune diseases or the cytokine release syndrome characteristic of severe COVID-19 patients.

ADULT-ONSET STILL'S DISEASE AND MACROPHAGE ACTIVATION SYNDROME FOLLOWING COVID-19 PNEUMONIA

CASE: A 41-year-old woman with recent COVID-19 pneumonia presented to the hospital with several months of fever, polyarthralgia, and weight loss. She reported waxing and waning shoulder, elbow, wrist, hip, knee, and ankle pain without identifiable triggers. She had no pertinent medical or family history. Vital signs were only notable for fever of 40C which recurred daily. Exam revealed tenderness to palpation of multiple joints;her skin had no rash, purpura, or nodules. Hepatosplenomegaly and axillary lymphadenopathy were noted. Infectious workup was negative for bacterial, viral, fungal, mycobacterial, parasitic, and protozoal infections. Initial studies demonstrated hemoglobin 8.2 mg/dL, lymphopenia, and aspartate transaminase 58 U/L. Flow cytometry, excisional lymph node biopsy, and bone marrow biopsy were negative for lymphoproliferative disease. Rheumatologic workup revealed elevated ferritin, triglycerides, Interleukin-6, soluble Interleukin-2 receptor (sIL-2R), and “C-X-C Motif Chemokine Ligand 9” (CXCL9);extensive rheumatologic serologies were otherwise negative. Her clinical picture was consistent with Macrophage Activation Syndrome (MAS). She also met diagnostic criteria for Adult-Onset Still's Disease (AOSD) given arthralgia, fever, lymphadenopathy, splenomegaly, abnormal liver function test (LFT), and otherwise negative workup. Her presentation suggested COVID-19 triggered AOSD which triggered MAS. We administered intravenous immune globulin (IVIG) and high-dose steroids. She clinically improved and was discharged with oral steroids. She returned to the hospital two months later for fever, arthralgia, and faint, evanescent rash with elevated erythrocyte sedimentation rate, C-reactive protein, ferritin, lactate dehydrogenase, and LFT consistent with an AOSD flare. She received intravenous steroids and Anakinra. Symptoms resolved, and she was discharged with plans to continue Anakinra and oral steroids. At followup, she had resolution of all symptoms. IMPACT/DISCUSSION: COVID-19 has many chronic complications, including triggering of underlying rheumatic disease. This sequence of events suggests that COVID-19 Pneumonia triggered an underlying diagnosis of AOSD. AOSD should be considered in the differential diagnosis of patients with quotidian fever and arthralgia following COVID-19 infection. AOSD is a diagnosis of exclusion and requires ruling out infectious, malignant, and rheumatic etiologies. AOSD may trigger MAS, a dysregulated immune response to underlying inflammation, and should be considered in patients with suspected infection refractory to treatment who have fever, splenomegaly, cytopenias, and elevated ferritin, triglycerides, sIL-2R, and CXCL9. CONCLUSION: COVID-19 has many chronic complications. AOSD may manifest after COVID-19 infection and should be considered in the differential diagnosis of patients with persistent fever and arthralgia. MAS should be suspected in patients with systemic inflammation refractory to treatment. AOSD may cause MAS.

Antibody patterns and T-cell responses to SARS-CoV-2 in Swedish primary health care workers during the Covid-19 pandemic

During the first period of the Covid-19 pandemic, most of the immunological studies on SARS-CoV-2 were based on hospitalized-and intensive care unit patients. In this study, a healthy population continuously exposed to the virus, Swedish primary health care workers (n = 156), were monitored for 6 months and the development of antibody patterns and T-cell responses to SARS-CoV-2 were evaluated. In addition to blood sampling, demographic-and clinical information such as PCR-tests, self-reported symptoms, underlying medical conditions, and medications were collected. Multivariate statistical analysis using OPLS-DA showed that Covid-19 infection was associated with SARS-CoV-2 specific IgG antibodies, T-cell responses, male sex, hypertension, and higher BMI and contrary, that not contracting the infection, was associated with female sex, no-or only SARS-CoV-2 specific IgA antibodies, smoking and airborne allergy. Analysis with Cytometry by Time-of-flight (CyTOF) revealed a unique cytotoxic CD4+ T cell population in participants with IgG-dominated antibody responses which expressed CD25, CD38, CD69, CD194, CD279, CTLA-4 and granzyme B. 10% of the study participants had only SARS-CoV-2 specific IgA antibodies with no detectable SARS-CoV-2 specific IgG antibodies. These IgA antibodies could partially neutralize the virus in vitro and none of the participants with this antibody pattern contracted Covid-19 during the study period. These results have the potential to further help us understand the immunological responses to SARS-CoV-2 infection.

Neurosarcoidosis after infection with COVID-19, a case report and review of the literature

Objective: N/A Background: The full scope of the mid- and long-term effects of SARS-CoV2 infection is currently being reported. The immune response might contribute not only to the development of ARDS, but also to other systemic complications after the acute setting. Some disorders, including those of autoimmune or presumed autoimmune etiology, have been reported to be triggered, exacerbated, or unmasked during the COVID-19 pandemic. Sarcoidosis is a multi-systemic inflammatory disorder believed to occur due to an exaggerated immune response to unknown antigens in the setting of genetic susceptibility. We present a case of neuro-sarcoidosis after COVID-19. Design/Methods: Descriptive study, case report. Results: A 51-year-old right-handed female presented with multiple cranial neuropathies and paresthesia after a mild case of COVID-19. Her symptoms included vertigo, hypoacusis, balance issues, left facial palsy, and paresthesia in her upper extremities. Her brain MRI with contrast showed bilateral enhancement of the VII and VIII cranial nerves. CSF analysis showed mild protein elevation and elevated CD4:CD8 ratio. Serum sIL-2R was also elevated. Her chest CT scan was abnormal, prompting a lymph node biopsy that was consistent with non-caseating granulomas. A diagnosis of probable neuro-sarcoidosis was made and she showed improvement with steroids. She was later started on methotrexate as a steroid sparing agent in the outpatient setting. Conclusions: To our knowledge, neuro-sarcoidosis has not been previously described in temporal association with COVID-19. It might be that this infection acts as one of the triggers for sarcoidosis. Some common pathways shared by these conditions could explain the possibility of such a trigger. These pathways include the ACE2 receptor, the TMRPPS gene, and certain cytokines. When aberrant, causing incomplete clearance of an antigen, these pathways might lead to the formation of granulomas. Further research surrounding the non-immediate effects of the novel coronavirus is needed to better delineate possible autoimmune consequences of this serious infection.

GDF-15 IS A PREDICTOR OF MORTALITY IN CHRONIC KIDNEY DISEASE PATIENTS WITH COVID-19 INFECTION

BACKGROUND AND AIMS: A cytokine storm drives the pathogenesis of severe coronavirus disease (COVID-19) and several biomarkers with different mechanisms of action have been linked to mortality. Chronic kidney disease (CKD) emerged as a very common risk factor for severe COVID-19. Indeed, CKD patients are at increased risk of premature death from many causes, including, but not limited to, cardiovascular disease (CVD) and infections. In this study, we aimed to investigate the associations between the growth differentiation factor 15 (GDF-15), an established cardiovascular and inflammatory biomarker and outcomes in CKD patients hospitalized for COVID-19. METHOD: A retrospective study on COVID-19 hospitalized subjects in the acute phase of the disease. A broad range of cytokines (CD25, IL-18, TNF-α, TNF RI, TNF RII, GDF-15, IL-7, LIF, IL-6, CHITINASE3-LIKE1, RAGE and Pentraxin-3) were assessed in plasma (Luminex, ELISA) collected upon hospitalization. A total of 77 subjects were divided into two groups according to their estimated glomerular filtration rate (eGFR, by CKD-EPI formula), ≥45 mL/min (n = 44), or <45 mL/min (n = 33). RESULTS: We found no statistical differences between the two groups in terms of demographic features. Among comorbidities, we found a higher percentage of patients with diabetes in the eGFR < 45 group. Likewise, the serum tests upon admission showed in the eGFR < 45 group a higher value of neutrophilic count. Upon hospital admission, the patient groups were comparable in terms of symptoms, time from symptom onset to admission and death or discharge, radiological evidence of pneumonia and respiratory parameters and time of hospitalization. Furthermore, there were no statistical differences between medical therapy during hospitalization, need for respiratory support with Continuous Positive Airways Pressure or Non-Invasive Mechanical Ventilation, or death rather than discharge as the clinical outcome. Serum levels of 20 different compounds were measured in COVID-19 patients admitted to the hospital 4-5 days after the onset of symptoms. Interestingly, we found that patients with lower renal function (eGFR < 45 mL/min) had a significant increase of GDF-15, CD-25 and RAGE and, furthermore, higher serum levels of these molecules were detected in non-survivor patients and in those who needed ventilation. Also, TNFα, TNFR I, TNFR II, IL-7 and LIF had a significant increase in patients with eGFR < 45 mL/min with more elevated levels in non-survivor patients. In univariate analysis low and mid-low GDF-15 quartiles (<4.45 ng/mL) were associated with lower mortality risk, while mid-high and high quartiles (>4.45 ng/mL) were associated with higher mortality risk (Figure 1). Independent association between GDF-15 quartiles and mortality risk was confirmed in Cox model adjusted for eGFR, age, fever, dyspnoea and P/F [hazard ratio (HR) 2.28, 95% confidence interval (CI) 1.53-3.39, P < 0.0001) The strength of association between GDF-15 quartiles and mortality risk was increased in patients with eGFR < 45 mL/min/1.73 m2 (HR 2.53, CI 1.34-4.79) compared with the other eGFR group (HR 1.99, CI 1.17-3.39) (Table 1). CONCLUSION: Our results demonstrate that GDF-15 is an independent predictor of COVID-19 mortality in CKD patients. Given the reported increase of this cytokine with age and its possible mechanistic role in various pathological conditions, our findings suggest that GDF-15 signalling pathway inhibitors may be included as possible therapeutic candidates for COVID-19 in CKD. (Table Presented).

CHRONIC INFLAMMATION MIGHT PROTECT HAEMODIALYSIS PATIENTS FROM SEVERE COVID-19

BACKGROUND AND AIMS: Patients on haemodialysis (HD) are expected to have excess mortality in coronavirus disease 2019 (COVID-19). This was challenged by a recent study reporting HD patients to have comparable mortality and decreased ICU admissions when hospitalized with COVID-19. It was speculated that an altered immune system due to chronic inflammation might protect HD patients from severe COVID-19. Therefore, we designed a study to describe the peripheral blood immune phenotype in HD patients and respective controls with COVID-19. METHOD: Sixty-four patients (31 HD, 33 non-HD) with PCR-confirmed severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection and 16 control patients (10 HD, 6 non-HD) were prospectively included. According to symptoms, COVID-19 patients were categorized as asymptomatic/mild and moderate/severe COVID-19 phenotypes. Cytokine profiling and immune phenotyping were performed. RESULTS: Th1 and Th17 plasma cytokine levels were highly increased in HD patients without SARS-CoV-2 infection and were not significantly regulated during COVID-19. In non-HD COVID-19 patients, these cytokines increased significantly with disease severity. While all patients with moderate/severe COVID-19 showed hallmarks of COVID-19 such as decreased CD3+, CD4+ and CD8+ and CD4+CD25hiFoxP3+ regulatory T cells, significantly increased CD38+CD8+ effector memory and CD38+CD8+ TEMRA T cells were detected in HD compared to non-HD patients with moderate/severe COVID-19. Furthermore, CD161+CD8+ T cells decreased significantly in non-HD COVID-19 patients dependent on disease severity, but not in HD patients. Dynamics of B cells and subtypes were comparable in HD and non-HD COVID-19 patients. Significantly fewer moderate/severe COVID-19 HD patients needed ICU treatment [1/13 (7.7%) HD, 12/24 (50%) non-HD], whereas no difference in mortality was observed [4/31 (12.9%) HD, 6/33 non-HD (18.2%)]. CONCLUSION: HD patients might be protected from severe COVID-19 due to their chronic inflammatory state with increased CD38+CD8+ effector memory and TEMRA T cells as well as CD161+CD8+ T cells.

IMMUNE EXHAUSTION IS ASSOCIATED with PERSISTENT SARS-CoV-2 VIREMIA and SEVERE DISEASE

Background: SARS-CoV-2 viremia is associated with adverse outcomes in COVID-19. The immunologic mediators of this relationship remain under-explored. In this study, we aimed to evaluate the correlation between immune exhaustion markers, SARS-CoV-2 viremia clearance and clinical outcomes. Methods: We included 126 participants with confirmed SARS-CoV-2 infection who were hospitalized at an urban hospital in Boston, Massachusetts, during the first surge of the COVID-19 pandemic in early 2020. Plasma samples from days 0, 3, and 7 of hospitalization were available for analyses. The plasma SARS-CoV-2 viral load was determined by reverse transcription quantitative PCR (RT-qPCR). Proteomics data were generated using the Olink platform and neutralization level was assessed using a pseudovirus neutralization assay. Viremia persistence was defined as >40 copies/ml (detection limit) if the baseline detectable viremia was <1000 copies/ml, or >100 copies/ml (quantification limit) if the baseline viremia was ≥1000 copies/ml at day 7 of admission. Partial least-squares discriminant analysis (PLS-DA) was used to select exhaustion markers that could distinguish viremia persistence and clearance. An exhaustion score was generated based on features selected by PLS-DA and was divided into four quartiles. Differentially expressed proteins between 1st and 4th quartiles were determined by linear model adjusting for baseline characteristics. R (4.1.0) was used for statistics. Results: Viremia persistence was associated with a higher level of baseline viremia, a higher rate of severe diseases and mortality within 28 days of follow-up. Viremia persistence was associated with elevation of certain exhaustion protein markers including TIM3, PDL1, LGALS9, LAG3 and IL2RA. With PLS-DA, we selected TIM3, PDL1, and LGALS9 into the exhaustion score modeling. A higher exhaustion score was associated with higher baseline viremia, persistent viremia, severe disease, and death (Figure). When compared to the lowest exhaustion score (1st quartile), the highest exhaustion score (4th quartile) was associated with elevation in proteins belonging to IL-18 signaling pathway, lung fibrosis, and immune evasion in COVID-19. The immune exhaustion level was not associated with the neutralization level. Conclusion: In participants with COVID-19, soluble exhaustion markers are associated with delayed viremia clearance, immune evasion independent of humoral immunity development, and adverse outcomes.

GDF-15 AS A PREDICTOR of MORTALITY in COVID-19: IMPLICATIONS for TREATMENT

Background: A cytokine storm drives the pathogenesis of severe COVID-19 and has therefore prompted the use of cytokine/transduction pathway inhibitors in the treatment of disease. However, numerous markers with different mechanisms of action have been linked to mortality, complicating the understanding of disease pathogenesis and the elaboration of therapeutic strategies. Methods: Retrospective study on COVID-19 hospitalized subjects in the acute phase of disease. A broad range of cytokines (CD25, IL-18, TNF-α, TNF RI, TNF RII, GDF-15, IL-7, LIF, IL-6, CHITINASE3-LIKE1, RAGE and Pentraxin-3) was assessed on plasma samples (Luminex, ELISA) collected upon hospitalization. Subjects were divided into two groups according to their clinical in-hospital death (Survivors: S;Non-Survivors: NS). Comparisons between groups were performed by Fisher's exact test or Mann-Whitney U test as appropriate. The association between each variable and mortality was analysed through univariate and multiple logistic regression models. Subsequently, survival analysis was conducted with Cox proportional hazard models. Results: 77 hospitalised Covid-19 patients were enrolled: 42 S and 35 NS (Figure 1A). As expected, in the NS group we found a higher proportion of subjects with fever and dyspnoea upon admission, development of ARDS and need of PEEP respiratory support (Figure 1A). NS also displayed significantly higher blood neutrophils/lymphocytes, C-reactive protein, LDH and procalcitonin as well as lower PaO2/FiO2 and peripheral O2 saturation values at admission (Figure 1A). In keeping with these findings, CD25, IL-18, IL-6, TNF-α, TNFRI, TNFRII, GDF-15, IL-7, LIF and Chitinase3-Like1, Pentraxin-3 and RAGE were significantly higher in NS than S (Figure 1B) and were associated to mortality in univariate regression models. In the multivariate regression model GDF-15 and fever were the two more relevant features associated with mortality (Figure 1C). In the survival analysis GDF-15 was the strongest predictor of mortality (HR 2,26, 1,55-3,31;p<0,01 reference group bottom quartile Figure 1D, E). Conclusion: Our in-depth characterization of the cytokine storm demonstrates that GDF-15 is an independent predictor of Covid-19 mortality. Given the reported increase of this cytokine with age and its possible mechanistic role in various pathological conditions, our findings suggest that GDF-15 signalling pathway inhibitors may be included as possible therapeutic candidates for Covid-19.