Evaluation of immunological responses to third COVID-19 vaccine among people treated with sphingosine receptor-1 modulators and anti-CD20 therapy

ImportancePeople living with multiple sclerosis (MS) and other disorders treated with immunomodulatory therapies remain concerned about suboptimal responses to coronavirus disease 2019 (COVID-19) vaccines. Important questions persist regarding immunological response to third vaccines, particularly with respect to newer virus variants. ObjectiveEvaluate humoral and cellular immune responses to third COVID-19 vaccine dose in people on anti-CD20 therapy and sphingosine 1-phosphate receptor (S1PR) modulators, including Omicron-specific assays. DesignObservational study evaluating immunological response to third COVID-19 vaccine dose in volunteers treated with anti-CD20 agents, S1PR modulators, and healthy controls. Neutralizing antibodies against USA-WA1/2020 (WA1) and B.1.1.529 (BA.1) severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) were measured before and after third vaccine. Cellular responses to spike peptide pools generated from WA1 and BA.1 were evaluated. SettingMount Sinai Hospital ParticipantsPeople treated with anti-CD20 therapy or S1PR modulators and healthy volunteers ExposureTreatment with anti-CD20 therapy, S1PR modulator, or neither Main outcomes and measuresSerum neutralizing antibodies and ex vivo T cell responses against SARS-CoV-2 antigens. ResultsThis cohort includes 25 participants on anti-CD20 therapy, 12 on S1PR modulators, and 14 healthy controls. Among those on anti-CD20 therapy, neutralizing antibodies to WA1 were significantly reduced compared to healthy controls (ID50% GM post-vaccination of 8.1 {+/-} 2.8 in anti-CD20 therapy group vs 452.6 {+/-} 8.442 healthy controls, P<0.0001) and neutralizing antibodies to BA.1 were below the threshold of detection nearly universally. However, cellular responses, including to Omicron-specific peptides, were not significantly different from controls. Among those on S1PR modulators, neutralizing antibodies to WA1 were detected in a minority, and only 3/12 had neutralizing antibodies just at the limit of detection to BA.1. Cellular responses to Spike antigen in those on S1PR modulators were reduced by a factor of 100 compared to controls (median 0.0008% vs. 0.08%, p<0.001) and were not significantly "boosted" by a third injection. Conclusions and RelevanceParticipants on immunomodulators had impaired antibody neutralization capacity, particularly to BA.1, even after a third vaccine. T cell responses were not affected by anti-CD20 therapies, but were nearly abrogated by S1PR modulators. These results have clinical implications warranting further study.

Exome-wide association study to identify rare variants influencing COVID-19 outcomes: Results from the Host Genetics Initiative

Host genetics is a key determinant of COVID-19 outcomes. Previously, the COVID-19 Host Genetics Initiative genome-wide association study used common variants to identify multiple loci associated with COVID-19 outcomes. However, variants with the largest impact on COVID-19 outcomes are expected to be rare in the population. Hence, studying rare variants may provide additional insights into disease susceptibility and pathogenesis, thereby informing therapeutics development. Here, we combined whole-exome and whole-genome sequencing from 21 cohorts across 12 countries and performed rare variant exome-wide burden analyses for COVID-19 outcomes. In an analysis of 5,085 severe disease cases and 571,737 controls, we observed that carrying a rare deleterious variant in the SARS-CoV-2 sensor toll-like receptor TLR7 (on chromosome X) was associated with a 5.3-fold increase in severe disease (95% CI: 2.75-10.05, p=5.41x10-7). This association was consistent across sexes. These results further support TLR7 as a genetic determinant of severe disease and suggest that larger studies on rare variants influencing COVID-19 outcomes could provide additional insights. Author SummaryCOVID-19 clinical outcomes vary immensely, but a patients genetic make-up is an important determinant of how they will fare against the virus. While many genetic variants commonly found in the populations were previously found to be contributing to more severe disease by the COVID-19 Host Genetics Initiative, it isnt clear if more rare variants found in less individuals could also play a role. This is important because genetic variants with the largest impact on COVID-19 severity are expected to be rarely found in the population, and these rare variants require different technologies to be studies (usually whole-exome or whole-genome sequencing). Here, we combined sequencing results from 21 cohorts across 12 countries to perform a rare variant association study. In an analysis comprising 5,085 participants with severe COVID-19 and 571,737 controls, we found that the gene for toll-like receptor 7 (TLR7) on chromosome X was an important determinant of severe COVID-19. Importantly, despite being found on a sex chromosome, this observation was consistent across both sexes.

Shift of lung macrophage composition is associated with COVID-19 disease severity and recovery

Though it has been 2 years since the start of the Coronavirus Disease 19 (COVID-19) pandemic, COVID-19 continues to be a worldwide health crisis. Despite the development of preventive vaccines, very little progress has been made to identify curative therapies to treat COVID-19 and other inflammatory diseases which remain a major unmet need in medicine. Our study sought to identify drivers of disease severity and death to develop tailored immunotherapy strategies to halt disease progression. Here we assembled the Mount Sinai COVID-19 Biobank which was comprised of ~600 hospitalized patients followed longitudinally during the peak of the pandemic. Moderate disease and survival were associated with a stronger antigen (Ag) presentation and effector T cell signature, while severe disease and death were associated with an altered Ag presentation signature, increased numbers of circulating inflammatory, immature myeloid cells, and extrafollicular activated B cells associated with autoantibody formation. Strikingly, we found that in severe COVID-19 patients, lung tissue resident alveolar macrophages (AM) were not only severely depleted, but also had an altered Ag presentation signature, and were replaced by inflammatory monocytes and monocyte-derived macrophages (MoM{Phi}). Notably, the size of the AM pool correlated with recovery or death, while AM loss and functionality were restored in patients that recovered. These data therefore suggest that local and systemic myeloid cell dysregulation is a driver of COVID-19 severity and that modulation of AM numbers and functionality in the lung may be a viable therapeutic strategy for the treatment of critical lung inflammatory illnesses.

Proteomic Characterization of Acute Kidney Injury in Patients Hospitalized with SARS-CoV2 Infection

Acute kidney injury (AKI) is a known complication of COVID-19 and is associated with an increased risk of in-hospital mortality. Unbiased proteomics using biological specimens can lead to improved risk stratification and discover pathophysiological mechanisms. Using measurements of [~]4000 plasma proteins in two cohorts of patients hospitalized with COVID-19, we discovered and validated markers of COVID-associated AKI (stage 2 or 3) and long-term kidney dysfunction. In the discovery cohort (N= 437), we identified 413 higher plasma abundances of protein targets and 40 lower plasma abundances of protein targets associated with COVID-AKI (adjusted p <0.05). Of these, 62 proteins were validated in an external cohort (p <0.05, N =261). We demonstrate that COVID-AKI is associated with increased markers of tubular injury (NGAL) and myocardial injury. Using estimated glomerular filtration (eGFR) measurements taken after discharge, we also find that 25 of the 62 AKI-associated proteins are significantly associated with decreased post-discharge eGFR (adjusted p <0.05). Proteins most strongly associated with decreased post-discharge eGFR included desmocollin-2, trefoil factor 3, transmembrane emp24 domain-containing protein 10, and cystatin-C indicating tubular dysfunction and injury. Using clinical and proteomic data, our results suggest that while both acute and long-term COVID-associated kidney dysfunction are associated with markers of tubular dysfunction, AKI is driven by a largely multifactorial process involving hemodynamic instability and myocardial damage.

Quantitative chest CT combined with plasma cytokines predict outcomes in COVID-19 patients

Despite extraordinary international efforts to dampen the spread and understand the mechanisms behind SARS-CoV-2 infections, accessible predictive biomarkers directly applicable in the clinic are yet to be discovered. Recent studies have revealed that diverse types of assays bear limited predictive power for COVID-19 outcomes. Here, we harness the predictive power of chest CT in combination with plasma cytokines using a machine learning approach for predicting death during hospitalization and maximum severity degree in COVID-19 patients. Patients (n=152) from the Mount Sinai Health System in New York with plasma cytokine assessment and a chest CT within 5 days from admission were included. Demographics, clinical, and laboratory variables, including plasma cytokines (IL-6, IL-8, and TNF-) were collected from the electronic medical record. We found that chest CT combined with plasma cytokines were good predictors of death (AUC 0.78) and maximum severity (AUC 0.82), whereas CT quantitative was better at predicting severity (AUC 0.81 vs 0.70) while cytokine measurements better predicted death (AUC 0.70 vs 0.66). Finally, we provide a simple scoring system using plasma IL-6, IL-8, TNF-, GGO to aerated lung ratio and age as novel metrics that may be used to monitor patients upon hospitalization and help physicians make critical decisions and considerations for patients at high risk of death for COVID-19.

Circulating proteins to predict adverse COVID-19 outcomes

Predicting COVID-19 severity is difficult, and the biological pathways involved are not fully understood. To approach this problem, we measured 4,701 circulating human protein abundances in two independent cohorts totaling 986 individuals. We then trained prediction models including protein abundances and clinical risk factors to predict adverse COVID-19 outcomes in 417 subjects and tested these models in a separate cohort of 569 individuals. For severe COVID-19, a baseline model including age and sex provided an area under the receiver operator curve (AUC) of 65% in the test cohort. Selecting 92 proteins from the 4,701 unique protein abundances improved the AUC to 88% in the training cohort, which remained relatively stable in the testing cohort at 86%, suggesting good generalizability. Proteins selected from different adverse COVID-19 outcomes were enriched for cytokine and cytokine receptors, but more than half of the enriched pathways were not immune-related. Taken together, these findings suggest that circulating proteins measured at early stages of disease progression are reasonably accurate predictors of adverse COVID-19 outcomes. Further research is needed to understand how to incorporate protein measurement into clinical care.

Acute COVID-19 gene-expression profiles show multiple etiologies of long-term sequelae

Summary ParagraphTwo years into the SARS-CoV-2 pandemic, the post-acute sequelae of infection are compounding the global health crisis. Often debilitating, these sequelae are clinically heterogeneous and of unknown molecular etiology. Here, a transcriptome-wide investigation of this new condition was performed in a large cohort of acutely infected patients followed clinically into the post-acute period. Gene expression signatures of post-acute sequelae were already present in whole blood during the acute phase of infection, with both innate and adaptive immune cells involved. Plasma cells stood out as driving at least two distinct clusters of sequelae, one largely dependent on circulating antibodies against the SARS-CoV-2 spike protein and the other antibody-independent. Altogether, multiple etiologies of post-acute sequelae were found concomitant with SARS-CoV-2 infection, directly linking the emergence of these sequelae with the host response to the virus.

Fatal breakthrough infection after anti-BCMA CAR-T therapy highlights suboptimal immune response to SARS-CoV-2 vaccination in myeloma patients

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) vaccines are highly effective in healthy individuals. Patients with multiple myeloma (MM) are immunocompromised due to defects in humoral and cellular immunity as well as immunosuppressive therapies. The efficacy after two doses of SARS-CoV-2 mRNA vaccination in MM patients is currently unknown. Here, we report the case of a MM patient who developed a fatal SARS-CoV-2 infection after full vaccination while in remission after B cell maturation antigen (BCMA)-targeted chimeric antigen receptor (CAR)-T treatment. We show that the patient failed to generate antibodies or SARS-CoV-2-specific B and T cell responses, highlighting the continued risk of severe coronavirus disease 2019 (COVID-19) in vaccine non-responders. In the largest cohort of vaccinated MM patients to date, we demonstrate that 15.9% lack SARS-CoV-2 spike antibody response more than 10 days after the second mRNA vaccine dose. The patients actively receiving MM treatment, especially on regimens containing anti-CD38 and anti-BCMA, have lower antibody responses compared to healthy controls. Thus, it is of critical importance to monitor this patient population for serological responses. Non-responders may benefit from ongoing public health measures and from urgent study of prophylactic treatments to prevent SARS-CoV-2 infection.

Divergent early antibody responses define COVID-19 disease trajectories

A damaging inflammatory response is strongly implicated in the pathogenesis of severe COVID-19 but mechanisms contributing to this response are unclear. In two prospective cohorts, early non-neutralizing, afucosylated, anti-SARS-CoV-2 IgG predicted progression from mild, to more severe COVID-19. In contrast to the antibody structures that predicted disease progression, antibodies that were elicited by mRNA SARS-CoV-2 vaccines were low in Fc afucosylation and enriched in sialylation, both modifications that reduce the inflammatory potential of IgG. To study the biology afucosylated IgG immune complexes, we developed an in vivo model which revealed that human IgG-Fc{gamma}R interactions can regulate inflammation in the lung. Afucosylated IgG immune complexes induced inflammatory cytokine production and robust infiltration of the lung by immune cells. By contrast, vaccine elicited IgG did not promote an inflammatory lung response. Here, we show that IgG-Fc{gamma}R interactions can regulate inflammation in the lung and define distinct lung activities associated with the IgG that predict severe COVID-19 and protection against SARS-CoV-2. One Sentence SummaryDivergent early antibody responses predict COVID-19 disease trajectory and mRNA vaccine response and are functionally distinct in vivo.

Serological response to COVID-19 vaccination in IBD patients receiving biologics

ObjectiveThe impact of medications on COVID-19 vaccine efficacy in IBD patients is unknown, as patients with immunosuppressed states and/or treated with immunosuppressants were excluded from vaccine trials. To address this, we evaluated serological responses to COVID-19 vaccination with the SARS-CoV-2 spike (S) mRNA BNT162b2 (Pfizer-BioNTech) and mRNA-1273 (NIH-Moderna) vaccines in IBD patients enrolled in an ongoing SARS-CoV-2 sero-survey at the Icahn School of Medicine at Mount Sinai in New York City. DesignWe obtained sera from 48 patients who had undergone vaccination with one or two vaccine doses. Sera were tested for SARS-CoV-2 anti-RBD total immunoglobulins and IgG (Siemens COV2T and sCOVG assays), anti-Spike IgG (in-house ELISA), and anti-nucleocapsid antibodies (Roche). ResultsAll IBD patients (15/15) who completed two-dose vaccine schedules achieved seroconversion to high levels. Two IBD patients with history of COVID-19 infections and who were seropositive at baseline seroconverted to high levels after the first dose. Concurrent biologic use was 85% (41/48), including 33% of patients (16) on TNF antagonist monotherapy, 42% (17) on vedolizumab monotherapy, 6% (3) on vedolizumab combination therapy with thiopurine, and 8% (4) ustekinumab; 1 patient was receiving guselkumab for psoriasis. Three patients (6%) were on oral steroids at the time of vaccination. ConclusionIBD patients receiving biologics can seroconvert with robust serological responses after complete Pfizer-BioNTech and NIH-Moderna COVID-19 vaccination. In IBD-patients with previous SARS-CoV-2 seroconversion, a single dose of either vaccine can induce high index values, mirroring findings from the general population.

Application of a 27-protein candidate cardiovascular surrogate endpoint to track risk ascendancy and resolution in COVID-19.

BackgroundThere is an urgent need for tools allowing the early prognosis and subsequent monitoring of individuals with heterogeneous COVID-19 disease trajectories. Pre-existing cardiovascular (CV) disease is a leading risk factor for COVID-19 susceptibility and poor outcomes, and cardiac involvement is prevalent in COVID-19 patients both during the acute phase as well as in convalescence. The utility of traditional CV risk biomarkers in mild COVID-19 disease or across disease course is poorly understood. We sought to determine if a previously validated 27-protein predictor of CV outcomes served a purpose in COVID-19. MethodsThe 27-protein test of residual CV (RCV) risk was applied without modification to n=860 plasma samples from hospitalized and non-hospitalized SARS-CoV-2 infected individuals at disease presentation from three independent cohorts to predict COVID-19 severity and mortality. The same test was applied to an additional n=991 longitudinal samples to assess sensitivity to change in CV risk throughout the course of infection into convalescence. ResultsIn each independent cohort, RCV predictions were significantly related to maximal subsequent COVID-19 severity and to mortality. At the baseline blood draw, the mean protein-predicted likelihood of an event in subjects who died during the study period ranged from 88-99% while it ranged from 8-36% in subjects who were not admitted to hospital. Additionally, the test outperformed existing risk predictors based on commonly used laboratory chemistry values or presence of comorbidities. Application of the RCV test to sequential samples showed dramatic increases in risk during the first few days of infection followed by risk reduction in the survivors; a period of catastrophically high cardiovascular risk (above 50%) typically lasted 8-12 days and had not resolved to normal levels in most people within that timescale. ConclusionsThe finding that a 27-protein candidate CV surrogate endpoint developed in multi-morbid patients prior to the pandemic is both prognostic and acutely sensitive to the adverse effects of COVID-19 suggests that this disease activates the same biologic risk-related mechanisms. The test may be useful for monitoring recovery and drug response.

Gastrointestinal involvement attenuates COVID-19 severity and mortality

Given that gastrointestinal (GI) symptoms are a prominent extrapulmonary manifestation of coronavirus disease 2019 (COVID-19), we investigated the impact of GI infection on disease pathogenesis in three large cohorts of patients in the United States and Europe. Unexpectedly, we observed that GI involvement was associated with a significant reduction in disease severity and mortality, with an accompanying reduction in key inflammatory proteins including IL-6, CXCL8, IL-17A and CCL28 in circulation. In a fourth cohort of COVID-19 patients in which GI biopsies were obtained, we identified severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) within small intestinal enterocytes for the first time in vivo but failed to obtain culturable virus. High dimensional analyses of GI tissues confirmed low levels of cellular inflammation in the GI lamina propria and an active downregulation of key inflammatory genes including IFNG, CXCL8, CXCL2 and IL1B among others. These data draw attention to organ-level heterogeneity in disease pathogenesis and highlight the role of the GI tract in attenuating SARS-CoV-2-associated inflammation with related mortality benefit.

Cytotoxic lymphocytes are dysregulated in multisystem inflammatory syndrome in children

Multisystem inflammatory syndrome in children (MIS-C) presents with fever, inflammation and multiple organ involvement in individuals under 21 years following severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection. To identify genes, pathways and cell types driving MIS-C, we sequenced the blood transcriptomes of MIS-C cases, pediatric cases of coronavirus disease 2019, and healthy controls. We define a MIS-C transcriptional signature partially shared with the transcriptional response to SARS-CoV-2 infection and with the signature of Kawasaki disease, a clinically similar condition. By projecting the MIS-C signature onto a co-expression network, we identified disease gene modules and found genes downregulated in MIS-C clustered in a module enriched for the transcriptional signatures of exhausted CD8+ T-cells and CD56dimCD57+ NK cells. Bayesian network analyses revealed nine key regulators of this module, including TBX21, a central coordinator of exhausted CD8+ T-cell differentiation. Together, these findings suggest dysregulated cytotoxic lymphocyte response to SARS-Cov-2 infection in MIS-C.

Mapping Systemic Inflammation and Antibody Responses in Multisystem Inflammatory Syndrome in Children (MIS-C)

Initially, the global outbreak of COVID-19 caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) spared children from severe disease. However, after the initial wave of infections, clusters of a novel hyperinflammatory disease have been reported in regions with ongoing SARS-CoV-2 epidemics. While the characteristic clinical features are becoming clear, the pathophysiology remains unknown. Herein, we report on the immune profiles of eight Multisystem Inflammatory Syndrome in Children (MIS-C) cases. We document that all MIS-C patients had evidence of prior SARS-CoV-2 exposure, mounting an antibody response with normal isotype-switching and neutralization capability. We further profiled the secreted immune response by high-dimensional cytokine assays, which identified elevated signatures of inflammation (IL-18 and IL-6), lymphocytic and myeloid chemotaxis and activation (CCL3, CCL4, and CDCP1) and mucosal immune dysregulation (IL-17A, CCL20, CCL28). Mass cytometry immunophenotyping of peripheral blood revealed reductions of mDC1 and non-classical monocytes, as well as both NK- and T-lymphocytes, suggesting extravasation to affected tissues. Markers of activated myeloid function were also evident, including upregulation of ICAM1 and Fc{gamma}R1 in neutrophil and non-classical monocytes, well-documented markers in autoinflammation and autoimmunity that indicate enhanced antigen presentation and Fc-mediated responses. Finally, to assess the role for autoimmunity secondary to infection, we profiled the auto-antigen reactivity of MIS-C plasma, which revealed both known disease-associated autoantibodies (anti-La) and novel candidates that recognize endothelial, gastrointestinal and immune-cell antigens. All patients were treated with anti-IL6R antibody or IVIG, which led to rapid disease resolution tracking with normalization of inflammatory markers. One Sentence SummaryThis study maps the cellular and serological immune dysfunction underlying a novel pediatric inflammatory syndrome associated with SARS-CoV-2.

A Streamlined CyTOF Workflow To Facilitate Standardized Multi-Site Immune Profiling of COVID-19 Patients

Mass cytometry (CyTOF) represents one of the most powerful tools in immune phenotyping, allowing high throughput quantification of over 40 single parameters at single-cell resolution. However, wide deployment of CyTOF-based immune phenotyping studies are limited by complex experimental workflows and the need for specialized CyTOF equipment and technical expertise. Furthermore, differences in cell isolation and enrichment protocols, antibody reagent preparation, sample staining and data acquisition protocols can all introduce technical variation that can potentially confound integrative analyses of large data-sets of samples processed across multiple labs. Here, we describe the validation of a streamlined whole blood CyTOF workflow that addresses many of these challenges and facilitates wider adoption of CyTOF immune monitoring across sites with limited technical expertise or sample-processing resources or equipment. Our workflow utilizes commercially available reagents including the Fluidigm MaxPar Direct Immune Profiling Assay, a dry tube 30-marker immunophenotyping panel, and SmartTube Proteomic Stabilizer, which allows for simple and reliable fixation and cryopreservation of whole blood samples. We validate a workflow that allows for streamlined staining of whole blood samples with minimal processing requirements or expertise at the site of sample collection, followed by shipment to a central CyTOF core facility for batched downstream processing and data acquisition. We further demonstrate the application of this workflow to characterize immune responses in a cohort of hospitalized COVID-19 patients, highlighting key disease-associated changes in immune cell frequency and phenotype.

A tertiary center experience of multiple myeloma patients with COVID-19: lessons learned and the path forward

BackgroundThe COVID-19 pandemic, caused by SARS-CoV-2 virus, has resulted in over 100,000 deaths in the United States. Our institution has treated over 2,000 COVID-19 patients during the pandemic in New York City. The pandemic directly impacted cancer patients and the organization of cancer care. Mount Sinai Hospital has a large and diverse multiple myeloma (MM) population. Herein, we report the characteristics of COVID-19 infection and serological response in MM patients in a large tertiary care institution in New York. MethodsWe performed a retrospective study on a cohort of 58 patients with a plasma-cell disorder (54 MM, 4 smoldering MM) who developed COVID-19 between March 1, 2020 and April 30, 2020. We report epidemiological, clinical and laboratory characteristics including persistence of viral detection by polymerase chain reaction (PCR) and anti-SARS-CoV-2 antibody testing, treatments initiated, and outcomes. ResultsOf the 58 patients diagnosed with COVID-19, 36 were hospitalized and 22 were managed at home. The median age was 67 years; 52% of patients were male and 63% were non-white. Hypertension (64%), hyperlipidemia (62%), obesity (37%), diabetes mellitus (28%), chronic kidney disease (24%) and lung disease (21%) were the most common comorbidities. In the total cohort, 14 patients (24%) died. Older age (>70 years), male sex, cardiovascular risk, and patients not in complete remission (CR) or stringent CR were significantly (p<0.05) associated with hospitalization. Among hospitalized patients, laboratory findings demonstrated elevation of traditional inflammatory markers (CRP, ferritin, D-dimer) and a significant (p<0.05) association between elevated inflammatory markers, severe hypogammaglobulinemia, non-white race, and mortality. Ninety-six percent (22/23) of patients developed antibodies to SARS-CoV-2 at a median of 32 days after initial diagnosis. Median time to PCR negativity was 43 (range 19-68) days from initial positive PCR. ConclusionsDrug exposure and MM disease status at the time of contracting COVID-19 had no bearing on mortality. Mounting a severe inflammatory response to SARS-CoV-2 and severe hypogammaglobulinemia were associated with higher mortality. The majority of patients mounted an antibody response to SARS-CoV-2. These findings pave a path to identification of vulnerable MM patients who need early intervention to improve outcome in future outbreaks of COVID-19.

An inflammatory cytokine signature helps predict COVID-19 severity and death

The COVID-19 pandemic caused by infection with Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) has led to more than 100,000 deaths in the United States. Several studies have revealed that the hyper-inflammatory response induced by SARS-CoV-2 is a major cause of disease severity and death in infected patients. However, predictive biomarkers of pathogenic inflammation to help guide targetable immune pathways are critically lacking. We implemented a rapid multiplex cytokine assay to measure serum IL-6, IL-8, TNF-, and IL-1{beta} in hospitalized COVID-19 patients upon admission to the Mount Sinai Health System in New York. Patients (n = 1484) were followed up to 41 days (median 8 days) and clinical information, laboratory test results and patient outcomes were collected. In 244 patients, cytokine measurements were repeated over time, and effect of drugs could be assessed. Kaplan-Meier methods were used to compare survival by cytokine strata, followed by Cox regression models to evaluate the independent predictive value of baseline cytokines. We found that high serum IL-6, IL-8, and TNF- levels at the time of hospitalization were strong and independent predictors of patient survival. Importantly, when adjusting for disease severity score, common laboratory inflammation markers, hypoxia and other vitals, demographics, and a range of comorbidities, IL-6 and TNF- serum levels remained independent and significant predictors of disease severity and death. We propose that serum IL-6 and TNF- levels should be considered in the management and treatment of COVID-19 patients to stratify prospective clinical trials, guide resource allocation and inform therapeutic options. We also propose that patients with high IL-6 and TNF- levels should be assessed for combinatorial blockade of pathogenic inflammation in this disease.

Pathophysiology of SARS-CoV-2: targeting of endothelial cells renders a complex disease with thrombotic microangiopathy and aberrant immune response. The Mount Sinai COVID-19 autopsy experience

BACKGROUNDSevere Acute Respiratory Syndrome Coronavirus-2 (SARS-CoV-2) and its associated clinical syndrome COVID-19 are causing overwhelming morbidity and mortality around the globe, disproportionately affecting New York City. A comprehensive, integrative autopsy series that advances the mechanistic discussion surrounding this disease process is still lacking. METHODSAutopsies were performed at the Mount Sinai Hospital on 67 COVID-19 positive patients and data from the clinical records were obtained from the Mount Sinai Data Warehouse. The experimental design included a comprehensive microscopic examination carried out by a team of expert pathologists, along with transmission electron microscopy, immunohistochemistry, RNA in situ hybridization, as well as immunology and serology assays. RESULTSLaboratory results of our COVID-19 cohort show elevated inflammatory markers, abnormal coagulation values, and elevated cytokines IL-6, IL-8 and TNF. Autopsies revealed large pulmonary emboli in four cases. We report microthrombi in multiple organ systems including the brain, as well as conspicuous hemophagocytosis and a secondary hemophagocytic lymphohistiocytosis-like syndrome in many of our patients. We provide electron microscopic, immunofluorescent and immunohistochemical evidence of the presence of the virus and the ACE2 receptor in our samples. CONCLUSIONSWe report a comprehensive autopsy series of 67 COVID-19 positive patients revealing that this disease, so far conceptualized as a primarily respiratory viral illness, also causes endothelial dysfunction, a hypercoagulable state, and an imbalance of both the innate and adaptive immune responses. Novel findings reported here include an endothelial phenotype of ACE2 in selected organs, which correlates with clotting abnormalities and thrombotic microangiopathy, addressing the prominent coagulopathy and neuropsychiatric symptoms. Another original observation is that of macrophage activation syndrome, with hemophagocytosis and a hemophagocytic lymphohistiocytosis-like disorder, underlying the microangiopathy and excessive cytokine release. We discuss the involvement of critical regulatory pathways.