Coronavirus disease 2019 (COVID-19) cases continue to rise across the globe. Over 54.3 million people have been infected with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) – the causative agent of COVID-19 – and 1.31 million have lost their lives.
Scientists in Taiwan recently published a report on their use of anti-neutrophil strategies to treat acute respiratory distress syndrome (ARDS) – a life-threatening condition that can arise in critical COVID-19 cases – in the journal Frontiers in Pediatrics.
Most cases of COVID-19 are mild and cause cold and flu-like symptoms: a fever, a cough and shortness of breath. Sometimes these are accompanied by a sore throat and tiredness, often with gut symptoms like abdominal pain and diarrhea, anosmia (loss of smell) and loss of taste. However, a significant proportion of cases develop complications such as ARDS, septic shock and thrombosis, often culminating in multiple organ failure and death.
Risk factors for adverse outcomes include a range of things: a person’s age (with those above 65 years being particularly compromised) and the presence of chronic medical conditions such as diabetes, cardiovascular disease and chronic obstructive pulmonary disease (COPD).
Role of neutrophils in immunity and inflammation
The innate immune response against SARS-CoV-2 is heavily dependent on neutrophils, a type of white blood cell that leads the body’s immune response to pathogens. Typically, in any infection, neutrophils are recruited to the damaged tissue, which they infiltrate through rolling, adhesion, or transmigration.
Once there, they are activated and secrete several effector chemicals that cause pathogen destruction, besides engulfing pathogens by phagocytosis. Some of the inflammatory mechanisms by which they destroy infectious pathogens include superanion release by respiratory burst, degranulation with protease release, and neutrophil extracellular trap (NET) formation.
However, these processes associated with neutrophil-induced inflammation can also worsen tissue damage even as it protects against the pathogen. As such, high neutrophil counts are characteristic of multiple inflammatory conditions (both acute and chronic) as well as autoimmune disorders, among which are asthma, COPD, atherosclerosis and thrombosis, autoimmune hepatitis and inflammatory bowel disease.
In this case, the SARS-CoV-2 RNAs act as pathogen-associated molecular patterns (PAMPs). These are recognized by Toll-like receptors (TLRs) such as TLR3, TLR7, TLR8, and TLR9. This causes interferon pathways to be activated, along with many pro-inflammatory cytokines like IL-6.
Dysregulated immunity and inflammation in severe infection
Initially, the innate immune response by activated neutrophils and macrophages protects the body against SARS-CoV-2, but as the viral load increases, exuberant inflammatory responses set in. In conjunction with the destruction of cells by the actively replicating virus in severe disease, this leads to massive tissue damage.
The damaged and dying cells induce dysregulated immune responses resulting in a cytokine storm. Again, positive feedback loops such as that formed by IL-1β and NETs exacerbate the inflammation. Nerve damage may occur due to direct viral invasion as well as from neutrophil-secreted reactive oxygen species (ROS) and NETs.
The effect of activated macrophages and neutrophils is thus to induce the severe non-cardiogenic pulmonary edema called ARDS. Thus, the ratio of neutrophils to lymphocytes is used as a marker of high risk for complications, including ARDS in COVID-19 patients.
Earlier papers have suggested the use of antivirals and anti-inflammatory therapies suppress viral replication and prevent immune dysregulated responses in COVID-19.
Neutrophilic inflammation the hallmark of ARDS
The authors quote another paper, “Neutrophil infiltration is the defining hallmark of ARDS.” This is true of bronchoalveolar lavage fluid, which contains both high levels of neutrophils and microparticles derived from them, as well as other chemokines that activate neutrophil recruitment cascades.
Neutrophil counts are usually high in COVID-19, rising in proportion to disease severity. These cells are key to the cytokine storm. Thus, neutrophil inhibitors of various kinds are being examined for their possible value in mitigating severe COVID-19-related ARDS.
Chemokine inhibitors and anti-EMP2 antibodies
Neutrophils outside the vascular compartment display higher chemokine binding, with activation of proteinase-activated receptors (PARs) on many innate immune, endothelial and epithelial cells, which triggers pro-inflammatory cytokine and chemokine release. The resulting flood of chemicals includes TNF, IL-1β, IL-2, and IL-6, and the chemokines CXCL8 (IL-8) and CCL2, all linked to the development of ARDS.
One strategy to treat or prevent ARDS in these patients, therefore, would be chemokine blockage, which is not, however, sufficient to prevent neutrophil recruitment completely.
Again, the viral protein causes IL-8 secretion from the lung epithelium, while alveolar type I cells release epithelial membrane protein 2 (EMP2) that regulates neutrophil migration in ARDS. Anti-EMP2 antibodies could, therefore reduce neutrophil infiltration.
Formyl peptide antagonists
They are activated by mitochondrial formyl peptides as well, the latter also being high in ARDS. Formyl peptide receptor 1 antagonists have been tested and found to be promising in treating ARDS in COVID-19.
The role of NET inhibitors
NETs are sticky nets of chromatin extruded by neutrophils decorated with the components of various neutrophil granules, and viscous mucus is richer in NETs. The virus stimulates healthy neutrophils to form NETs. NETs are elevated in COVID-19-positive individuals and cause microthrombi. Thus, drugs that maintain sputum viscosity were preferable, and mucolytics were avoided in these patients. NET inhibitors also reduce NET formation and macrophage M1 markers in mice with acute lung injury.
In ARDS, macrophage phagocytosis of NETs is lower than normal, while macrophage efferocytosis is promoted by the AMPK pathway. This offers another route for intervention in ARDS.
Neutrophil elastase inhibitors
Several drugs that affect neutrophil function are used in lung disease, and these, along with other neutrophil-targeting drugs, could be used to inhibit ARDS. These include neutrophil elastase inhibitors like sivelestat, since this enzyme, as part of the serine protease family, is known to cause ARDS and to exacerbate asthma and COPD. Another potential therapy is the use of cathepsin C or DPP1, which activates serine proteases like neutrophil elastase.
N-acetylcysteine (NAC), which is used in respiratory disease and skin disease for its mucolytic and antioxidant activity, is also a respiratory burst inhibitor and has been found to reduce active lung injury. There is some evidence of the benefit of NAC in COVID-19-associated ARDS.
Drugs that increase cAMP/cGMP levels
Phosphodiesterases (PDEs) are enzymes that break down cAMP and cGMP, the molecules that act as the second messenger. These belong to the class of enzymes that metabolize the intracellular second messenger cyclic adenosine monophosphate (cAMP) and cyclic guanosine monophosphate (cGMP). In particular, cAMP-specific PDE4 is present on a wide variety of immune cells and plays a role in neutrophil-induced lung inflammation. PDE4 inhibitors are thus another treatment option to be explored.
Drugs that increase cAMP levels like dipyridamole and pentoxifylline, a platelet inhibitor-cum-immunomodulator, are among other possible therapies.
Other potential strategies
DNase inhibitors such as dornase alfa could help lyse sputum; disulfiram (a gasdermin D inhibitor), may prevent NETosis; and a variety of chemokine/cytokine inhibitors such as the IL-6R antagonist tocilizumab, and the IL-1R inhibitor anakinra, are undergoing clinical trials for their value in COVID-19.
Finally, corticosteroids are already recommended for seriously ill patients with COVID-19 on ventilators or receiving oxygen supplementation because of their anti-inflammatory effects. However, they can promote neutrophilic inflammation. More studies are required to explore their use.
Older immunomodulators such as dapsone and colchicine are also being tested for their inhibitory effect on pro-inflammatory cytokines. Hydroxychloroquine and azithromycin were strongly recommended by many in the popular media at first, primarily due to their promising in vitro activity, but have fallen out of favor after several trials failed to show any benefit.
Thus, the study draws attention to the potential for antineutrophilic drugs to ameliorate the severity of ARDS in COVID-19 and thus improve the outcome by preventing excessive neutrophil activation.