Immune Changes in Severe COVID-19 Pulmonary Infections

Study Overview
=================
Brief Summary
-----------------
SARS-CoV-2 outbreak causes a spectrum of clinical patterns that varies from asymptomatic infection to mildly symptomatic manifestations and more-severe forms that need intensive care. Until now, the immune response to SARS-CoV-2 virus infection has been poorly reported to help decision for immune modulation therapies. As a consequence, trials have been designed to test both anti-inflammatory molecules as steroids or anti-bodies against IL-6, and others proposing to boost immunity with interferon beta based on similar inclusion criteria. The immune response to infective agents including viruses may have a complex time evolution with early and late phases corresponding to different patterns, oscillating between pro-inflammation and immune-depression. The potential window to improve outcome in COVID-19 by therapeutic intervention aimed at a fine tuning between immune toxicity and immunodepression requires a longitudinal assessment during the course of illness, especially for the patients who develop acute respiratory failure. Immune monitoring of both innate and adaptive immunity would then be essential to appropriately design clinical trials. The whole blood cells evaluation was recorded according to the time intervals between the onset of symptoms and the sampling after ICU admission. Patients' care was standardized, especially with regard to ventilation, sedation, and antimicrobial treatment. In this study the investigators prospectively perform a longitudinal study of both innate and adaptive immunity on patients admitted to ICU for an COVID-19 related acute respiratory failure. The data will be analyzed in reference to the onset of initial symptoms and also to the admission in ICU. The primary end point is the evolution of the characterization of monocytes and their subsets in term of number and expression of HLA-DR. A similar approach is used for lymphocytes and their subtypes with in addition, an ex vivo testing of their capabilities to be stimulated by SARS-CoV-2 viral proteins in term of TNFalpha, INFgamma, and IL1beta production. The secondary end-point was to test the association with outcomes and other non-specific markers of inflammation as CRP (C reactive protein), PCT (procalcitonin), DDimers and ferritin.

Detailed Description
-----------------
The most severe form of COVID-19 treated in intensive care for acute respiratory failure may have a poor prognosis. Both the level of IL-6 and the severity of the lymphopenia have been associated to the poor prognosis. Better knowledge of the time evolution of the circulating immune cells subpopulations and functions will help to best tailor the treatment: anti-inflammatory strategy at the initial phase might be rapidly shifted to immune stimulation when immunodepression is diagnosed. It is then essential to assess the patients' immune status using flowcytometry methods to characterize both innate and adaptive immunity of the whole blood circulating immune peripheral blood mononuclear cells (PBMC). After cell staining with the adequate cell markers, the flowcytometry (NAVIOS® Flow Cytometer (Beckman Coulter) allowed to analyze the number and the function of the cells with an adequate gating strategy and the Kaluza® software v2.1 (Beckman Coulter). The data were then grouped by time intervals referring to the onset of symptoms and also to the ICU admission. The trend for innate immunity (monocytes number and subpopulations, HLA-DR expression) and for adaptive immunity (lymphocytes and subpopulations) will be analyzed. Since it is unknown if whole blood CD3/CD4 and CD3/CD8 lymphocytes elicit an exhaustion pattern and/or an abnormal response, an ex vivo testing of their reactivity for SARS-CoV-2 viral proteins will be performed. This test of polyfunctionality will characterize the intracellular cytokine expression (IL-1 beta, TNFalpha, and INFgamma) both for CD4 and CD8 T cells.

Official Title
-----------------
Whole Blood Immune Cells Characterization in Critically Ill COVID-19 Patients: A Prospective Study

Conditions
-----------------
SARS-CoV-2, Immune System Disorder

Participation Criteria
=================
Eligibility Criteria
-----------------
Inclusion Criteria: confirmed COVID-19 a positive RT- PCR, a highly suggestive thoracic CTScan, severe hypoxemia Exclusion Criteria: none

Ages Eligible for Study
-----------------
Minimum Age: 18 Years

Sexes Eligible for Study
-----------------
All

Accepts Healthy Volunteers
-----------------
No

Study Plan
=================
How is the study designed?
-----------------

Arms and Interventions

| Participant Group/Arm | Intervention/Treatment |
| --- | --- |
| Immuno Cohort<br>COVID-19 confirmed ICU patients, sedated and ventilated, sequential characterization of circulating immune cells over their ICU stay. Every 2 to 3 days, fresh whole blood aliquots from routine blood counts were processed on a Flow Cytometer (Beckman Coulter) to determine immune cells subpopulations | |
| Cortico Cohort<br>COVID-19 confirmed ICU patients, sedated and ventilated, sequential characterization of circulating immune cells over their ICU stay. According to RECOVERY study, early routine administration of dexamethasone 6 mg/day over 10 days. Every 2 to 3 days, fresh whole blood aliquots from routine blood counts were processed on a Flow Cytometer (Beckman Coulter) to determine immune cells subpopulations | |

What is the study measuring?
-----------------
Primary Outcome Measures

| Outcome Measure | Measure Description | Time Frame |
| --- | --- | --- |
| Changes in monocytes HLA-DR expression | circulating immune cell characterization | through ICU stay, an average of 30 days | 
| Changes in lymphocytes subpopulations numbers | circulating immune cell characterization | through ICU stay, an average of 30 days | 
| Changes in monocytes number | circulating immune cell characterization | through ICU stay, an average of 30 days | 

Secondary Outcome Measures

| Outcome Measure | Measure Description | Time Frame |
| --- | --- | --- |
| TNFalpha level | stimulation by SARS-CoV-2 viral proteins | 4 hours | 
| INFgamma level | stimulation by SARS-CoV-2 viral proteins | 4 hours | 
| IL1beta level | stimulation by SARS-CoV-2 viral proteins | 4 hours | 
| SOFA score | Sequential Organ dysfunction assessement, ranging from 0 (better) to 24 (worst) outcome | through ICU stay, an average of 30 days | 
| number of recorded deaths | mortality | through study completion, an average of 6 months | 
| presence of pneumonia | infectious complications | through ICU stay, an average of 30 days | 
| presence of bacteremia | infectious complications | through ICU stay, an average of 30 days | 
| presence of urinary tract infection | infectious complications | through ICU stay, an average of 30 days | 
| C reactive protein | inflammation marker | through ICU stay, an average of 30 days | 
| D Dimers | inflammation marker | through ICU stay, an average of 30 days |