key: cord-0977614-xj0ymkfc
authors: Thapa, Komal; Verma, Nitin; Gurjeet Singh, Thakur; Kaur Grewal, Amarjot; Kanojia, Neha; Rani, Lata
title: COVID-19-Associated Acute Respiratory Distress Syndrome (CARDS): Mechanistic Insights on Therapeutic Intervention and Emerging Trends
date: 2021-11-03
journal: Int Immunopharmacol
DOI: 10.1016/j.intimp.2021.108328
sha: e04f786f59b202fd71a1be9911392e53a463139d
doc_id: 977614
cord_uid: xj0ymkfc

AIMS: The novel Coronavirus disease 2019 (COVID-19) has caused great distress worldwide. Acute respiratory distress syndrome (ARDS) is well familiar but when it happens as part of COVID‐19 it has discrete features which are unmanageable. Numerous pharmacological treatments have been evaluated in clinical trials to control the clinical effects of CARDS, but there is no assurance of their effectiveness. MATERIALS AND METHODS: A systematic review of the literature of the Medline, Scopus, Bentham, PubMed, and EMBASE (Elsevier) databases was examined to understand the novel therapeutic approaches used in COVID-19-Associated Acute Respiratory Distress Syndrome and their outcomes. KEY FINDINGS: Current therapeutic options may not be enough to manage COVID-19-associated ARDS complications in group of patients and therefore, the current review has discussed the pathophysiological mechanism of COVID-19-associated ARDS, potential pharmacological treatment and the emerging molecular drug targets. SIGNIFICANCE: The rationale of this review is to talk about the pathophysiology of CARDS, potential pharmacological treatment and the emerging molecular drug targets. Currently accessible treatment focuses on modulating immune responses, rendering antiviral effects, anti-thrombosis or anti-coagulant effects. It is expected that considerable number of studies conducting globally may help to discover effective therapies to decrease mortality and morbidity occurring due to CARDS. Attention should be also given on molecular drug targets that possibly will help to develop efficient cure for COVID-19-associated ARDS.

The outburst of COVID-19 or SARS-CoV-2 has put terrible impact on global public health [1].

This novel disease was first erupted in December 2019, in China at Wuhan city and presently considered as a deadly disease. According to the current report, worldwide total COVID-19 cases are 179,260,990 with 3,882, 169 deaths [2] . Severity in COVID-19 results from severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infections to viral pneumonia, therefore the manifestation is observed in combination namely viral pneumonia and ARDS. The disease has been said to be spread through zoonotic spill out of β-coronavirus type 2b which is nowadays spreading among humans [3, 4] . Apart from inducing pneumonia and ARDS, corona virus family is likely to transform and contaminate susceptible populations, thereby causing a global threat [5] .

The disease has a distinct pathophysiology and clinical pattern that confuses the efficiency of the presently accessible therapeutics.

A two-peak clinical course is involved in COVID-19 disease headed by an asymptomatic phase (also called as noiseless viral replication) [6, 7] . During first phase, patient undergo with extensive viral replication of about five days after infection [8] and therefore the primary symptoms are generally coughing, fever and dyspnea [9] . Additional common symptoms comprises of headaches, anosmia, diarrhea, fatigue and some neurological symptoms [10] . Viral replication usually collapses about 5-7 days subsequent to the episode of symptoms [11] . After 7-10 days of appearance of symptoms, patients move into second phase and become seriously ill due to strong immune reaction and therefore the urgency to shift patient in ICU becomes high because of ARDS or multi-organ failure [12, 13, 14] . It has been revealed in 25 COVID-19 studies (involving 4881 severe and non-severe cases) [15] that the patients severely affected with COVID-19 were mostly suffering from diabetes and hypertension [15] . Apparently the pervasiveness of CARDS, acute kidney injury or shock was high in patients affected severely with COVID-19 giving rise to mortality rate of about 30%. [15] . Another analysis discovered that higher age, comorbidities involving obesity, diabetes, hypertension, COPD (chronic obstructive lung disease), renal disease, heart disease, immunodeficiency, male gender are risk factors for COVID-19 progression [16] . It has not been reported that COVID-19 may induce maternal or fetal complications in pregnant expression and promote disruption of the immune system and contribute to the development of tissue fibrosis and such impact on ACE2 could be also involved in COVID-19 related fibrosis [32] (Figure2).

SARS-CoV-2 infects airway epithelial cells or immune cells via binding to ACE2 receptors and stimulates release of Damage-associated molecular pattern (DAMP), along with production of inflammatory cytokines. The interaction between epithelial cells and immune cells leads to a broad range of clinical manifestations such as ARDS, pneumonia, cytokine storm and disseminated intravascular coagulation (DIC) [33] (Figure 2 ). The entry of viral cell and its replication leads to widespread endothelial tissue damage that increases the permeability and accumulation of proteinrich fluids in alveolar and interstitial space [33] . During the exudative phase, several changes are observed such as fibrin deposition, hyaline membrane generation, large-scale tissue inflammation, necrosis and apoptosis that cause damage [33] . In addition to these, proliferation of fibroblasts, myofibroblasts, pulmonary vasculopathy may develop pneumonia and lung fibrosis with irreparable destruction may develop due to increased release of cytokines (such as Interleukin (IL)-1β and transforming-growth factor (TGF)-β) [34] . As result lung conformity in CARDS may be reduced or normal [34] whereas, patient self-inflicted lung injury (P-SILI) or potential ventilatorassociated may have augmented pulmonary lesions over time [35] (Figure2).

Although the inflammatory response in viral pneumonia is beneficial to help local tissues to fight with the infection, but the aggravated inflammatory responses in pneumonia patients may provoke the excessive proinflammatory cytokines release known as "cytokine storm", giving rise to harmful results such as fibrosis, disperse alveolar damage, progressive respiratory failure and multiple organ failure [36] . Cytokine storm is a hyperactive immune condition characterized by T cells proliferation, disturbance in M1/M2 balance. Macrophages consist of two functional subtypes: M1 also known as pro-inflammatory macrophages engaged in developing proinflammatory reactions, chemotaxis, production of free radicals, and M2 is alternatively known as anti-inflammatory macrophages that reduces inflammation via producing growth factors (VEGF, EGF, PDGF) and suppressing effector T cells for repair [37, 26] . Dysregulation in host immune response triggers hyperinflammatory syndrome in COVID-19. The clinical feature of syndrome involves viral-induced hemophagocytic lymphohistiocytosis and macrophage activation with a cytokine storm [38] (Figure1). The cytokine storm usually occurs in viral respiratory infections such as H5N1 influenza, SARS-CoV-1, and SARS-CoV-2. Hyper inflammatory condition is confirmed in extremely pathogenic coronavirus-induced ARDS and also death and has brought major concern in developing therapies dealing with intense immune responses [39] .

Overexpression of IL-6 is considered as a hallmark of cytokine storm as in many severe cases; IL-6 level was found extensively high than in mild cases. The pro-inflammatory cytokine production is facilitated by DAMPS released from infected cells and engagement of PAMPS with Toll-like-receptor (TLR 3, 7/8) [40] . COVID-19 patients admitted in ICU have shown elevated level of tumor-necrosis factors, IL-6 and monocyte chemotactic protein as compared to normal patients [40] . The neutrophil-to-lymphocyte ratio (NLR) is frequently detected high in patients that suggest severity of disease. Amongst several cytokines, one discrete intracellular Prominent IL-6 level in serum has been normally reported in seriously affected COVID-19 patients [42, 43] . Therefore the restorative efficiency of IL-6 antagonists on COVID-19 patients is being evaluated by researchers. These result further approved JAK inhibitors as clinical management strategy for COVID-19. Antiviral response and chemoattractant pathways are regulated by type I interferon (IFN) in COVID-19 development [44] . Therefore, adequate type I IFN response is an essential aspect for fighting with viral infections and impairment in type I IFN responses is related to disease severity. 

Endothelial dysfunction is responsible for COVID-19-associated vascular inflammation and coagulopathy [45] . In a small cohort study involving COVID-19 patients, the augmented levels of 

Thalidomide is an immunomodulatory drug that promotes T cell responses by inhibiting IL-6 and has shown valuable effects in preclinical viral or bacterial induced ARDS [57]. Currently thalidomide is in Phase 2 (ClinicalTrials.gov Identifier: NCT04273581) clinical investigation against SARS-CoV-2. A recombinant IL-1 receptor antagonist known as Anakinra inhibited the physiological action of IL-1a and IL-1b by preventing their attachment with interleukin-1 type receptor in a competitive fashion and is usually employed in rheumatic diseases infection [58] . In a large phase 3 studies, Anakinra did not reduced death rate in patients with septic shock and 

JAK inhibitors are widely used in many inflammation driven diseases such as rheumatoid arthritis, psoriasis and inflammatory bowel diseases [60] . There are numerous US FDA and European Medicine Association approved JAK inhibitors that are baricitinib, ruxolitinib, fedratinib, tofacitinib, upadacitinib, oclacitinib and some are under clinical investigation. Ruxolitinib was the first oral JAK1/2 inhibitor to be approved for neoplastic diseases [61] . Studies have demonstrated that JAK inhibitor has therapeutic implications in condition such as cytokine-driven inflammatory syndromes and sHLH (Haemophagocytic lymphohisticytosis) [62] . Therefore, the data supported the use of JAK inhibitor ruxolitinib in serious COVID-19 cases with dysregulated immune system [63] . Recently a study revealed the efficiency of ruxolitinib in severely affected COVID-19 patients, in which patients treated with ruxolitinib plus standard-of-care (SoC) showed rapid clinical recovery and effectiveness in contrast to control group [64] . Currently clinical trials are underway for investigating ruxolitinib in COVID-19 patients such as one of the trials (ClinicalTrials.gov Identifier: NCT04362137) is being sponsored by Novartis Pharmaceuticals to study the effect of ruxolitinib against COVID-19 associated cytokine storm. Recently Ruxolitinib was evaluated in a clinical study involving 18 COVID-19 patients with progressive acute respiratory distress syndrome [65] . Administration of ruxolitinib ameliorated the course of disease and avoided mechanical ventilation in 89% of treated patients [65] . On the other hand, the rate of COVID-19-associated ARDS patients shifted from NIV to mechanical ventilation with no ruxolitinib treatment was 57% and 27% of them had died [65] .

Interferons emerge to have a multipurpose function in ARDS, amid inconsistent effects based on ARDS etiology and the type of interferon (type I, II or III) [66] . In a phase 2a study, Interferon-β1α (Type I interferon) showed promising results due to its anti-inflammatory, anti-fibrotic and anti-viral effects, but does not showed efficacy against ARDS in phase 3 study [67] . A recent study demonstrated that IFN-γ might facilitate entry of viral SARS-CoV-2 by upregulating ACE2 in the epithelial cells of lungs [68] . However interferon therapy could be a possible therapy for COVID-19. Type I interferons have displayed different inhibitory potencies towards SARS and MERS and have been evaluated in combination with antiviral drugs for these viral infections [68] .

As revealed earlier, a recent phase 2 study of triple therapy comprising lopinavir/ritonavir, ribavirin, and IFN-β1β shown to enhance SARS-CoV-2 infected patient's recovery as compared to lopinavir/ritonavir only [69] . 

Remdesivir is a broad-spectrum anti-viral drug that has been formerly explored as an anti-Ebola 

Lopinavir/ritonavir is HIV protease inhibitors usually employed in combination therapies [86] . An open-labeled study involving 199 severely COVID-19 patients unluckily didn't display any clinical improvement; however mortality rate was less in treated group [87] . In a recently phase 2 study the combined effect of lopinavir/ritonavir with ribavirin and IFN-β1β was studied in mildmodest COVID-19 patients and revealed that the combination reduced viral shedding and hospital resides in comparison to lopinavir/ritonavir alone [88] .

Umifenovir is an anti-viral drug approved for use in influenza which alters viral contact with ACE2, and recently showed enhanced viral clearance in a retrospective study involving 50 COVID-19 patients as compared to lopinavir/ritonavir [89] . More studies are currently undergoing for investigating the safety and efficiency of Umifenovir in COVID-19 patients.

Chloroquine and hydroxychloroquine are antimalarial drugs that acts by preventing virus from fusing with the host cells via inhibiting ACE2 receptor present on the membrane of host cells and reduces release of pro-inflammatory cytokine [90, 91] . In preclinical models, Chloroquine has shown to inhibit lung injury caused by influenza A H5N1 virus [92] and SARS-CoV-2 infection in vitro [93] . It has been newly revealed that combination of hydroxychloroquine with azithromycin condensed viral load in 20 SARS-CoV-2 infected patients [94] . On the other hand, potential adverse effects such as cardiotoxicity have been informed with the use of combination of azithromycin with chloroquine and hydroxychloroquine in COVID-19 patients [95] . There are other clinical investigation in progress using Nitazoxanide (NTZ) either alone or in combination with other drugs (Ivermectin or Hydroxychloroquine) to treat COVID-19 patients (ClinicalTrials.gov Identifier: NCT04360356, NCT04361318). Nitazoxanide, an antiparasitosis has shown great potential for repurposing to treat a variety of viral infections including SARS-CoV and MERS-CoV by targeting both host and viral components [96] and is currently ongoing clinical trial to evaluate its efficacy in patients with COVID-19 (ClinicalTrials.gov Identifier: NCT04552483). Ivermectin is also an antiparasitic drug is widely used to treat worm infections and scabies. Numerous observational and randomized studies have evaluated Ivermectin for the treatment against COVID-19 infection and the results concluded that Ivermectin demonstrated a strong therapeutic efficacy against COVID-19 by reducing death rates [97] and currently it is undergoing clinical trial for in COVID-19 high risk patients (I-TECH), ClinicalTrials.gov Identifier: NCT04920942.

Thrombosis, both microvascular and macrovascular, is a well-known feature in multiple organs with fatal cases of COVID-19. Thrombosis may thus contribute to renal failure, hepatic injury and respiratory failure in COVID-19 patients [98] . Thrombotic stroke has been recently reported in 

Dysfunctioning in coagulation process is one of the pathological complications in severe COVID-19 patients which results in pulmonary microvascular thrombosis and deep venous thrombosis [99] . Therefore anticoagulant therapy such as heparin has been suggested by some expert group as treatment regimens for these complications in severe COVID-19 patients, but its effectiveness

have not yet proven. Heparin is currently undergoing in clinical "REMAP-CAP study (ClinicalTrials.gov Identifier: NCT02735707)". Study such as "CHARTER study" on nebulized heparin is also undergoing] (Figure 2 ).

Alteration in coagulation process and reduced fibrinolysis are the attributes of COVID-19 that leads to microvascular thrombosis of the lung vessels which are linked to ACP (Acute cor pulmonale) and ARDS [100] . Currently rtPA was evaluated in 67 year old patient affected with ACP with elevated levels of C-reactive protein, high ferritin values and dead alveolar space [100] .

The effect was that rtPA reduced all these parameters, improved oxygenation and reduced ventilatory ratio. The rtPA effects may have been more significant if the patients wouldn't have had certain comorbidities involving obesity and hypertension. In severely affected COVID-19associated ARDS, micro-thrombosis is possibly observed as a unique phenotype characterized by high levels of D-dimers, an enlarged portion of dead space and hypercapnia. Recent clinical data have suggested that rtPA can be a potential therapy in COVID-19-associated ARDS [101] .

Currently nebulized rtPA for ARDS due to COVID-19 is under phase 2 clinical evaluation (ClinicalTrials.gov Identifier NCT04356833).

One of the hopeful treatment strategies in COVID-19 patients is plasma therapy. It has been demonstrated that serum obtained from SARS-CoV-1 patients provided protection from SARS-CoV-2 infection. This strategy possibly will be more successful if prophylactically employed [102] [103] [104] . In critically ill H1N1 patients, convalescent plasma treatment decreased viral load and transience [105] and the similar effect was observed in seriously ill SARS-CoV-2 patients [106] .

Treatment of convalescent plasma was well tolerated in a trial involving 5000 COVID-19 patients (ClinicalTrials.gov Identifier: NCT04338360) [107] . There are other trials ongoing (ClinicalTrials.gov Identifier: NCT04356534, NCT04372979) for the evaluation of effectiveness and safety of anti-SARS-CoV ( Figure 2 ).

The human monoclonal antibodies, Tocilizumab and sarilumab have shown therapeutic effects in patients suffering from Still's disease intricated with SIRS and ARDS [108] . A new retrospective study involving 21 COVID-19patients suggested that treatment with tocilizumab improved lung oxygenation with improved CT lung opacity and decreased white cell counts [109] . Presently several phases 2/3 trials are evaluating sarilumab or tocilizumab for COVID-19 patients, with probable outcomes. The available pharmacotherapies have been summarized in the table below (Table 1) . [109]

The vigorous activation of innate immune system is a characteristic of COVID-19 severity that triggers intense release of proinflammatory cytokine and chemokines and IL-6 which is a special prognostic of COVID-19 death [110] . High extent of interleukins such as IL-1β and IL-6 were identified in autopsy tissues of SARS-CoV patients. The fatality causing due to inflammatory response has led to the development of immunoregulators to treat CARDS. Activation of immune system during viral infections is highly correlated with NLRP3 (Nod-like receptor family, pyrin domain-containing 3) inflammasome [111] [112] [113] . There are numerous literatures linking NLRP3

inflammasome and cytokine storm in the pathogenesis of COVID-19 patients [114] . The viral (SARS-CoV 3a) protein triggers the activation of NLP3 inflammasome in the lipopolysaccharideprimed macrophages and stimulates IL-1β secretion which leads to K + efflux and generation of mitochondrial reactive oxygen species [115] . NLRP3 comprises of inactive procaspase-1 and an adapter element apoptosis-associated speck-like protein containing a CARD (caspase activation and recruitment domain). Studies have reported that various internal and external stimuli or viral RNA activates NLRP3 inflammasome through pores formation and lysosomal degradation leading to pyroptosis and inflammation (associated cell death) [116] . NLP3 after activation converts an enzyme procaspase-1 into an active effector protease caspase-1 which then promotes maturation of pro-inflammatory cytokines for example IL-pro-IL-1β (pro-interleukin 1β) into its active form IL-1β [117] . These further stimulates other downstream inflammatory mediators such as inflammatory potential in SARS-CoVs infection and emerges out to be a considerable target whose inhibition may reduce COVID-19 induced lung tissue inflammation [122] . Efforts are being made to discover the potential role of NLRP3 inflammasome in various inflammatory diseases. Some naturally derived NLRP3 inflammasome inhibitor, Parthenolide (sesquiterpene lactone found in feverfew plant) in addition to synthetic compound Bay 11-7082 and related vinyl sulfone compounds have shown to exert inhibitory effect on NLRP3 inflammasome [123] . Noticeably, Bay 11-7082 and Parthenolide inhibited NLRP3 inflammasome and NF-κB inflammatory pathways and reduced lung inflammation and improved survival in SARS-CoV-infected animals.

Pirfenidone also suppressed apoptosis and oxidative stress in a mouse model of LPS-induced acute lung Injury (ALI), where treatment with Pirfenidone decreased caspase activation and reduced inflammatory release of IL-1β and TGF-β [124] . Another recently available NLRP3 inflammasome inhibitor is Tetracycline that shown to reduce mortality, by reducing neutrophil infiltration, vascular leakage in murine LPS ALI model. It also reduced release of proinflammatory cytokine and caspase activation [125] . Currently, Pirfenidone is being investigated in clinical phase 3 (ClinicalTrials.gov Identifier: NCT04282902) for SARS-CoV-2 treatment.

Heme oxygenase, a ubiquitous enzyme has currently attained a lot of interest because of its multiple therapeutic effects in most disease conditions. There exist three isoforms of Heme oxygenase 1; HO-1, HO-2 and HO-3, HO-3 is the splice variant of HO-2 [126] . Out of these isoforms, HO-1 is also recognized as heat shock protein-32 encoded HMOX1 gene and its transcription is induced by various stimuli such as radiations, infections, toxins and injuries such as acute lung injury and lung I/R injury [127] . The physiological function of HO-1 is to promote heme oxidation to carbon monoxide (CO), ferrous iron and biliverdin (BV) [128] . BV then gets converted to bilirubin (BR) via enzyme biliverdin reductase. This mechanism makes BR more electrophilic and relatively increases BR affinity for Keap1-Nrf2 that stimulates induction of Nrf2dependant antioxidant gene [129] . At present, no pre-clinical or clinical data is available to validate the beneficial role of modulating HO-1 in COVID-19, but administration of heme or induction of HO-1 could be beneficial in fighting SARS-CoV-2 infection by degrading these end-products [130] .

HO-1 performs an important role in cell survival as it provides protection against inflammation, oxidative stress and removes degraded proteins as revealed in vitro and in vivo models of acute lung injury and inflammation [131] . The upregulation of HO-1 in response to inflammation and oxidative stress has been discovered in numerous cells such as neutrophils, monocytes, basophils, vascular smooth muscle cells, endothelial cells and macrophages. In different lung disease models,

HO-1 has displayed its anti-inflammatory action [132] . Higher expression of HO-1 was found in mononuclear cells of inflammatory lesions of carrageenan-induced lung inflammation model in contrast to peripheral mononuclear cells [133] . The enhanced HO-1 expression resulted in reduced inflammation, whereas pretreatment with HO-1 inhibitor stimulated inflammation and HO-1 inducer decreased inflammation. Further HO-1 also reduced HMGB1 release from macrophages and production of pro-inflammatory cytokine in LPS model via CO generation [134, 135] . In another study, CO Supplementation and HO-1 induction in-vivo LPS induced septic shock model reduced HMGB, IL-β and TNF-α level in plasma. CO has also displayed anti-apoptotic and antiinflammatory effects in vivo and in vitro models of lung ischemia/reperfusion via modulation of P38-MAPK pathway [136] . In another in-vivo model of LPS induced acute lung injury, treatment with BV reduced inflammation in bronchoalveolar epithelial cells and alveolus along with reduced pulmonary edema, by decreasing the expression of transcription factor NF-κB which is responsible for LPS-stimulated inflammation and cytokine production [137] . Recent facts discovered that HO- administration of curcumin for about 6 months dose 1 to 4 g/day had shown to increase cholesterol levels and the opposite effect was observed in short-term treatment. Therefore, while targeting HO-1 system, hormetic response should be measured first [142, 143] .

Various thrombosis and platelet aggregation [147] . It has been confirmed that HO-1 induction with the treatment of activated protein C in a mouse model of venous thrombosis reduced IL-6 production.

Therefore all these studies propose that HO-1 induction via HO-1 inducer or heme degradation end-products (CO, BV, and BR) show anti-thrombotic effects by reducing endothelial injury, inflammatory responses and levels of pro-coagulant factors such as von Willebrand factor, tissue factor and PAI, thus HO-1 may be a proficient approach in preventing COVID-19 linked deaths as inflammation stimulated-coagulopathy is the most disturbing consequences of COVID-19 [148] .

One of the key mediators of platelet activation and aggregation is Proteinase-activated receptor 1 (PAR1), a G-Protein coupled receptor (GPCR) and also an instigator of the coagulation cascade.

Activation of PAR1 is stimulated by a serine protease known as Thrombin or by PAR1 agonist via cleaving its amino terminal and revealing a ligand which further self stimulates the receptor [149] .

The expression of PAR1 is found in all cell types related to the pathobiology COVID-19 involving pneumocytes, platelets, endothelial cells and fibroblasts. Thrombin produced from prothrombin is the part of coagulation process and the main constituent for coagulation process is the tissue factor.

disease is induced by hyper inflammation [150] . Therefore elevated production of thrombin and tissue factor activates PAR1 in patients with severe acute lung injury or COVID-19-associated ARDS. PAR1 also regulates endothelial function besides its action on platelets [150, 151] .

Thrombin is protective at low concentration but at high concentration it promotes endothelial disruption or dysfunction. PAR1 displays pathological phenotype in epithelial and alveolar cells of the lung such as release of inflammatory factors and apoptosis. PAR1 is also responsible in promoting lung fibrosis by stimulating transformation of fibroblasts into myofibroblasts and increasing secretions of extracellular matrix proteins [152] . The pathophysiology of COVID-19 has put forward PAR1 as a potential target in COVID-19. Study on mouse model have suggested that PAR1 inhibition reduced inflammation and enhanced immune response to viral infection in host, whereas a study demonstrated PAR1 having protective role in viral infection [153] . The contradictory effects may be due to variation in viral load in animals undertaken in these studies.

However the most of the studies (in vivo) keep up the idea that PAR1 plays pathological role in inflammation and infectious diseases and its inhibition may have beneficial effects against viral respiratory infections [154] . Therefore vorapaxar or atopaxar are PAR1 inhibitors that can be employed in trials, however their preclinical validation on COVID-19 models prior to clinical trials is needed as vorapaxar has major side effects such as fatal bleeding events, but atopaxar has fewer bleeding events as well as shorter half-life than vorapaxar [155] . Knowing these risks, carefulness is needed while evaluating the potential of PAR1 inhibition as a means of treatment in COVID-19

patients. Further preclinical studies should assess the outcomes of PAR1 antagonists on fibroblasts, alveolar epithelial cells and endothelial cells in animal models of COVID-19 [156, 157] . Also the effect of PARP1antagonists in COVID-19 associated thrombosis need to be replicated in animal models. Efforts are being made for developing animal models for COVID-19 such as rhesus monkeys, ferrets and other organisms [157] . Studies on animal models may further help to describe the mechanism of protective effect of PAR1 inhibition. Currently, data regarding the effect of PAR1 on platelets and other cell types in mouse models of Acute respiratory distress syndrome (ARDS) and Acute lung injury (ALI) are available that can be further extended in COVID-19 relevant animal models.

Inhibition of Akt with therapeutic agents have been accounted to inhibit the expressions of angiotensin-converting enzyme 2 (ACE2) receptor which aids viral entrance into lung cells, therefore targeting Akt for COVID-19 seems to be feasible option as the use of angiotensinconverting enzyme (ACE) inhibitors aggravates COVID-19-associated ARDS [158] [159] [160] . In numerous disease states, PI3K/Akt pathway has shown to promote inflammation, whereas Akt1 gene deletion in mice following myocardial ischemia recovered cardiac function and reduced inflammation [161] . Also it has been revealed that Akt inhibition suppresses inflammation but inhibition of PTEN may promote inflammation via activation of Akt in regulatory T cells (Tregs) [162] . Adoptive relocation of Tregs also exhibited to restrain fibro proliferation and improved lung injury resolution in experimental animal model disclosing that rising Tregs number in ARDS lungs could be a novel approach to treat COVID-19 patients at complex points and it has been recently discovered that Tregs number can be increased in ARDS lung via Akt inhibition as demonstrated in mouse model of bacterial endo-toxin induced lung injury, promoting recovery and injury resolution [163] . ACE2 has a pathological role in lung inflammation and pulmonary arterial hypertension. Akt inhibitors for example MK2206 and triciribine are Akt have been accounted to improve the pathophysiological outcome of ACE2 in hepatic steatosis [164] . But, a connection needs to be established between ACE2 activation and Akt pathway in COVID-19 patients. Virus expressions are mainly found in peripheral immune cells, macrophages of CNS and microglia [165] . P2X7Rs on activation allows entry of Na + /Ca 2+ and exit of K + . The complications associated with acute respiratory distress syndrome (ARDS) such as pneumonia is the main reason of mortality and morbidity in Covid-19 patient and there is no efficient treatment until now [166] . In ARDS patients, high ATP levels were found in bronchoalveolar lavage fluid (BALF) and also in mice induced with LPS-induced acute lung injury (ALI) [167] . P2X7Rs are regarded as the main drivers of inflammation. The receptors on Macrophages/microglia contains patterns of recognition responsible for recognizing pathogen-associated molecular patterns (PAMPs) such as lipopolysaccharide (LPS) or danger-associated molecular patterns (DAMPs), such as ATP [168] .

The activation of macrophages promotes cytokine release via two processes firstly by LPS mediated stimulation of toll-like receptor 4 and secondly ATP-induced P2X7Rs activation which further encourages NLRP3 induced caspase-1 activation and later IL-1β secretion [169] . As a result NLRP3 can be activated by SARS-CoV-2 viral proteins and P2X7 receptor. Activation of P2X7 receptor enhances chemokine and cytokine release such as IL-6, IL-8, TNF-α, CCL2, CCL3

and pro-fibrotic factors for example TGF-β. In P2X7 deficient mice model of silica-induced lung fibrosis or models of bleomycin, reduced inflammation was observed [170] [171] [172] . Thus, it suggests that P2X7 receptor deletion or antagonists might be of benefit for less severe COVID-19 patients.

activated syndrome (MAS) and causes long term cytokine release common of this syndrome and of COVID-19 as well [173] . Besides, a key feature of ARDS is the wide pulmonary edema due to elevated level of VEGF and VEGF. The P2X7 receptor may induce in vivo VEGF release and neoangiogenesis, thereby known as a potent inducer of VEGF release and blockade of P2X7 receptors inhibited VEGF-mediated increase in vascular permeability [174] . Thus, targeting P2X7 receptor may be beneficial to fight early phase in ARDS. Further, thromboembolic complications common amongst critically ill COVID-19 patients can be reduced by targeting P2X7 receptors as activation of these receptors leads to enormous release of tissue factor [175] . In mice model of ARDS, LPS was applied intratrachealy which induced lung inflammation similar to human ARDS [176] . LPS inhalation induces ALI, since LPS primarily targets plasma membrane pattern recognition receptors (PRRs), whereas SARS-CoV-2 targets intracellular receptors and P2X7 receptor blockade substantially condensed cytokine levels, infiltration of inflammatory cells and lung damage in ALI [177, 178] . Deletion of P2X7 receptor also decreased alveolar macrophage and [182] . The therapeutic effects on blockade of P2X7 receptor has never been evaluated in uncontrolled hyperinflammation condition as in Covid-19. Therefore, P2XR might be a promising strategy and antagonists for P2X7 receptor could be beneficial in patient with COVID-19-associated ARDS with severe pneumonia.

The interaction of viral protein also known as spike (S) protein with ACE-2 is catalyzed by a serine protease enzyme known as TMPRSSS-2 which further cleaves spike protein into S1 and S2 and then facilitates the interaction of these proteins with ACE2 receptor on host cell surface. S1 helps viral spike protein to bind with ACE-2, whereas S2 helps viral RNA to fuse in host cell membrane [183] . The expression of TMPRSSS-2 is highly found in nasal epithelial in goblet, ciliated cells of nasal epithelium and other parts of lung tissue [184] . Therefore blocking the activity of TMPRSS-2 might be a possible therapy for SARS-CoV-2. The general structure of this serine protease is illustrated by N-terminal domain and C-terminal catalytic domain [185] . The active site of the enzyme possesses extremely conserved nature of amino acid residues such as Ser441, His296,

Gln438, Trp461 and Lys432 [185] . Studies have demonstrated that serine protease inhibitors can inhibit SARS-CoV-2 in the host cell. Camostat mesilate, a serine protease inhibitor reduced 65% mortality in mouse infected with SARS-Co. This clinically approved serine protease inhibitors is currently under clinical investigation (NCT04321096) for its effect against SARS-CoV-2 [186] .

Therefore TMPRSS2 could be considered as a helpful tool for pharmacological research on SARS-CoV-2. There are other potential drug apart from camostat that can be repurposed to evaluate their in vitro activity against SARS-CoV-2, such as nafamostat and 4-(2-aminoethyl) benzenesulfonyl

fluoride. In addition, mucolytic agents have been also proposed as TMPRSS2 inhibitor for COVID-19 therapy [187] . Further, a new therapeutic option is to use experimental and computational methods or estrogen and androgen-related compounds such as Enzatulamide, Genistein and Estradiol for transcriptional inhibition of TMPRSS2 [188] . TMPRSS2 expression can be altered by androgens and estrogens, therefore a study has suggested that inhibition of androgen pathways and activation of estrogen pathways may be a new strategy to manage symptoms in COVID-19 patients [189] .

At present, SARS-CoV-2 is causing rigorous illness all across the world sustaining transmission from human-to-human and has emerged as a serious danger to public health. Understanding of pathophysiological mechanism of COVID-19-associated ARDS (CARDS) has led to repurpose some approved drugs against SARS-CoV-2 infection such as Immunomodulators, anti-viral drugs, anti-thrombosis drugs and immunotherapy. Currently more pharmacological agents are undergoing clinical evaluation and their reports are underway. Apart from currently available pharmacological treatment for CARDS, attention should be also given on development of molecular drug targets and evaluation for their efficacy in preclinical and clinical platform that may help to manage the complications associated with CARDS. Despite of advanced scientific development, no permanent cure for diabetes, hypertension and cardiac disease exists. Also immune diseases such as HIV are also difficult to treat and therefore attention should be given on new strategies to maintain immune balance [190] . Zinc supplement in combination with anti-viral drug against viral infection is a dual-edged sword. Long-term Zinc treatment suppresses immune

system. An in vitro study revealed that surplus Zinc supplementation can decrease IFN-γ expression in younger subject whereas in elderly persons increased interferon-alpha (IFN-α) production by leukocytes. However award-risk ratio is in favor of Zinc supplementation in COVID-19 [191, 192] . There are less preclinical and clinical data on this aspect and results from currently ongoing trials employing Zinc in COVID-19 will throw light on the efficacy against viral infections in vivo. During viral infection, patients suffer from vitamin C deficiency due to high metabolic consumption which reduces the immunity of patient to fight COVID-19 associated detrimental effects so vitamin C supplementation is included in the treatment protocol of patients with viral infection. A recent study evaluated the role and effectiveness of vitamin C in counteracting the cytokines storm in cases with COVID-19 [193] . Vitamin D supplementation also have shown to provide protection to respiratory tract by killing enveloped viruses and preserving tight junctions. It also reduces cytokine storm by reducing pro-inflammatory cytokines production patients and the emerging molecular drug targets that possibly will help to develop effectual treatment for COVID-19-associated ARDS patients. 

Janus kinase inhibitors: A therapeutic strategy for cancer and autoimmune diseases

Use of the JAK inhibitor ruxolitinib in the treatment of hemophagocytic lymphohistiocytosis

The potential of JAK/STAT pathway inhibition by ruxolitinib in the treatment of COVID-19

Ruxolitinib in treatment of severe coronavirus disease 2019 (COVID-19): A multicenter, single-blind, randomized controlled trial

Ruxolitinib rapidly reduces acute respiratory distress syndrome in COVID-19disease. Analysis of data collection from RESPIRE protocol

Tramadol-induced seizurogenic effect: a possible role of opioid-dependent histamine (H1) receptor activation-linked mechanism

Emerging pharmacological therapies for ARDS: COVID-19and beyond

SARS-CoV-2 receptor ACE2 is an interferon-stimulated gene in human airway epithelial cells and is detected in specific cell subsets across tissues

Hydroxychloroquine and azithromycin as a treatment of COVID-19: results of an open-label non-randomized clinical trial

Combination of Hydroxychloroquine Plus Azithromycin As Potential Treatment for COVID-19Patients: Safety Profile, Drug Interactions, and Management of Toxicity

Therapeutic Potential of Nitazoxanide: An Appropriate Choice for Repurposing versus SARS-CoV-2

Ivermectin for prevention and treatment of COVID-19 infection: a systematic review, meta-analysis and trial sequential analysis to inform clinical guidelines

Venous and arterial thromboembolic complications in COVID-19 patients admitted to an academic hospital in

Recombinant tissue plasminogen activator treatment for COVID-19associated ARDS and acute cor pulmonale

Plasma tissue plasminogen activator and plasminogen activator inhibitor-1 in hospitalized COVID-19patients

High levels of plasminogen activator inhibitor-1, tissue plasminogen activator and fibrinogen in patients with severe COVID-19. MedRxiv

Convalescent Blood: Current Perspective on the Efficacy of a Legacy Approach in COVID-19Treatment

Convalescent plasma therapy: a promising coronavirus disease 2019 treatment strategy

Treatment for emerging viruses: Convalescent plasma and COVID-19

Convalescent plasma treatment reduced mortality in patients with severe pandemic influenza A (H1N1) 2009 virus infection

Treatment of 5 critically ill patients with COVID-19with convalescent plasma

Expanded access to convalescent plasma for the treatment of patients with COVID-19

Role of IL-6 inhibitor in treatment of COVID-19-related cytokine release syndrome

Pharmacological management of COVID-19patients with ARDS (CARDS): A narrative review

SARS-CoV-2 infection: The role of cytokines in COVID-19disease

Targeting the NLRP3 inflammasome in severe COVID-19

Novel coronavirus-induced NLRP3 inflammasome activation: a potential drug target in the treatment of COVID-19

Severe COVID-19: NLRP3 inflammasome dysregulated

Severe acute respiratory syndrome coronavirus viroporin 3a activates the NLRP3 inflammasome

The NLRP3 inflammasome and COVID-19: Activation, pathogenesis and therapeutic strategies

Differential splicing of the apoptosis-associated speck like protein containing a caspase recruitment domain (ASC) regulates inflammasomes

Inflammasome activation and regulation: toward a better understanding of complex mechanisms

The deregulated immune reaction and cytokines release storm (CRS) in COVID-19disease

Insights into SARS-CoV-2 genome, structure, evolution, pathogenesis and therapies: Structural genomics approach

Identification and characterisation of small molecule inhibitors of feline coronavirus replication

Novel Coronavirus-Induced NLRP3 Inflammasome Activation: A Potential Drug Target in the Treatment of COVID-19

Anti-inflammatory compounds parthenolide and Bay 11-7082 are direct inhibitors of the inflammasome

Pirfenidone ameliorates lipopolysaccharide-induced pulmonary inflammation and fibrosis by blocking NLRP3 inflammasome activation

Tetracycline alleviates acute lung injury by inhibition of NLRP3 inflammasome

Haem oxygenase (HO): an overlooked enzyme of plant metabolism and defence

Targeting heme oxygenase-1 and carbon monoxide for therapeutic modulation of inflammation

Heme oxygenase-1: its therapeutic roles in inflammatory diseases

Role of Nrf2/HO-1 system in development, oxidative stress response and diseases: an evolutionarily conserved mechanism

Therapeutic potential of heme oxygenase-1/carbon monoxide in lung disease

Significance of Heme and Heme Degradation in the Pathogenesis of Acute Lung and Inflammatory Disorders

Heme oxygenase-1 modulation: A potential therapeutic target for COVID-19and associated complications

Heme oxygenase-1 (HO-1) cytoprotective pathway: A potential treatment strategy against coronavirus disease 2019 (COVID-19)-induced cytokine storm syndrome

Heme-oxygenase-1 induction and carbon monoxide-releasing molecule inhibit lipopolysaccharide (LPS)-induced high-mobility group box 1 release in vitro and improve survival of mice in LPS-and cecal ligation and puncture-induced sepsis model in vivo

Carbon monoxide rescues ischemic lungs by interrupting MAPK-driven expression of early growth response 1 gene and its downstream target genes

Heme Oxygenase Dependent Bilirubin Generation in Vascular Cells: A Role in Preventing Endothelial Dysfunction in Local Tissue

The standard of care of patients with ARDS: ventilatory settings and rescue therapies for refractory hypoxemia

Can bilirubin nanomedicine become a hope for the management of COVID-19?

The role of cytokines during the pathogenesis of ventilator-associated and ventilator-induced lung injury

333 Seventh Avenue

Endothelial cell signaling and ventilatorinduced lung injury: molecular mechanisms, genomic analyses, and therapeutic targets

Significance of Heme and Heme Degradation in the Pathogenesis of Acute Lung and Inflammatory Disorders

Curcumin, hormesis and the nervous system

Curcumin effects on blood lipid profile in a 6-month human study

Targeting heme oxygenase-1 in vascular disease

Paradoxical rescue from ischemic lung injury by inhaled carbon monoxide driven by derepression of fibrinolysis

Targeting the heme-heme oxygenase system to prevent severe complications following COVID-19infections

Heme oxygenase-1 and carbon monoxide in the heart: the balancing act between danger signaling and prosurvival

Activated protein C accelerates venous thrombus resolution through heme oxygenase-1 induction

Protease-activated receptors (PARs): mechanisms of action and potential therapeutic modulators in PAR-driven inflammatory diseases

Proteinase-activated receptor 1: A target for repurposing in the treatment of COVID-19?

Protease-activated receptor 1 as a potential therapeutic target for COVID-19

Targeting neutrophils to treat acute respiratory distress syndrome in Coronavirus disease

PAR1 contributes to influenza A virus pathogenicity in mice

Infection and inflammation and the coagulation system

Novel anti-platelet agents: focus on thrombin receptor antagonists

Inflammation and thrombosis in COVID-19pathophysiology: proteinase-activated and purinergic receptors as drivers and candidate therapeutic targets

Structure, function and pathophysiology of protease activated receptors

Is targeting Akt a viable option to treat advanced-stage COVID-19patients?

A review on possible mechanistic insights of Nitazoxanide for repurposing in COVID-19

SARS-CoV-2 infectivity and neurological targets in the brain

Differential effects of Akt1 signaling on short-versus long-term consequences of myocardial infarction and reperfusion injury

The PTEN pathway in Tregs functions as a critical driver of the immunosuppressive tumor microenvironment and tolerance to apoptotic cells

Delayed Akt suppression in the lipopolysaccharide-induced acute lung injury promotes resolution that is associated with enhanced effector regulatory T cells

Angiotensin-converting enzyme 2/angiotensin-(1-7)/Mas axis activates Akt signaling to ameliorate hepatic steatosis

P2X7 Receptors Amplify CNS Damage in Neurodegenerative Diseases

COVID-19: Consider cytokine storm syndromes and immunosuppression

Extracellular ATP is a danger signal activating P2X7 receptor in a LPS mediated inflammation (ARDS/ALI)

Extracellular ATP is a danger signal activating P2X7 receptor in lung inflammation and fibrosis

The inflammasome: a caspase-1-activation platform that regulates immune responses and disease pathogenesis

Emerging role of IL-6 and NLRP3 inflammasome as potential therapeutic targets to combat COVID-19: role of lncRNAs in cytokine storm modulation

New Insights into Pathomechanisms and Treatment Possibilities for Lung Silicosis

Natural killer cell dysfunction and its role in COVID-19

Vascular endothelial growth factor (VEGF) in acute lung injury (ALI) and acute respiratory distress syndrome (ARDS): paradox or paradigm?

Thromboembolic complications in critically ill COVID-19patients are associated with impaired fibrinolysis

Animal models of acute lung injury

A rationale for targeting the P2X7 receptor in coronavirus disease 19

The cytokine storm in COVID-19: An overview of the involvement of the chemokine/chemokine-receptor system

Lipopolysaccharide induces alveolar macrophage necrosis via CD14 and the P2X7 receptor leading to interleukin-1α release

A potential therapeutic role for P2X7 receptor (P2X7R) antagonists in the treatment of inflammatory diseases

Can anti-inflammatory strategies light up the dim depression pipeline?

Role of proteolytic enzymes in the COVID-19infection and promising therapeutic approaches

SARS-CoV-2 entry factors are highly expressed in nasal epithelial cells together with innate immune genes

Gene of the month: TMPRSS2 (transmembrane serine protease 2)

Camostat mesylate against SARS-CoV-2 and COVID-19-Rationale, dosing and safety

TMPRSS2: A potential target for treatment of influenza virus and coronavirus infections

In-silico approaches to detect inhibitors of the human severe acute respiratory syndrome coronavirus envelope protein ion channel

SARS-CoV-2 and the possible connection to ERs, ACE2, and RAGE: Focus on susceptibility factors

Targeting host cell proteases as a potential treatment strategy to limit the spread of SARS-CoV-2 in the respiratory tract

Vitamin C-An adjunctive therapy for respiratory infection, sepsis and COVID-19

Zinc and COVID-19: basis of current clinical trials

Immune-boosting role of vitamins D, C, E, zinc, selenium and omega-3 fatty acids: Could they help against COVID-19?

The role of vitamin C in pneumonia and COVID-19