Mechanisms of Lung Injury Induced by SARS-CoV-2 Infection.

The lung is the major target organ of SARS-CoV-2 infection, which causes COVID-19. Here, we outline the multistep mechanisms of lung epithelial and endothelial injury induced by SARS-CoV-2: direct viral infection, chemokine/cytokine-mediated damage, and immune cell-mediated lung injury. Finally, we discuss the recent progress in terms of antiviral therapeutics as well as the development of anti-inflammatory or immunomodulatory therapeutic approaches. This review also provides a systematic overview of the models for studying SARS-CoV-2 infection and discusses how an understanding of mechanisms of lung injury will help identify potential targets for future drug development to mitigate lung injury.

COVID-19 and SARS-CoV-2

SARS-CoV-2 is classified in the Betacoronavirus genus under the Coronaviridae family of viruses (1). It is the seventh known coronavirus to infect people, after 229E, NL63, OC43, HKU1, Middle East Respiratory Syndrome (MERS)-CoV, and the original SARS-CoV (now referred to as SARS-CoV-1) (1). Other coronaviruses that can cause illnesses range from the common cold to more severe diseases, such as SARS and MERS. SARS-CoV-2 shares 79.6% sequence similarity to SARS-CoV-1, which caused an outbreak in 2003, and 96% similarity to a strain of bat coronavirus, implying a potential zoonotic origin. As part of the Riboviria realm in the Orthornavirae kingdom, SARS-CoV-2 contains a positive-sense single-stranded RNA genome and uses RNA-dependent RNA polymerase (RdRp) to replicate its genome (2). SARS-CoV-2 has the second largest known RNA virus genome with over 29,000 basepairs and encodes for 27 proteins, including four structural proteins: spike (S) glycoprotein, matrix, and nucleocapsid, and small envelope proteins and nonstructural proteins . As the COVID-19 pandemic continues to progress, noxious SARS-CoV-2 variants with enhanced transmission rates or increased disease severity have emerged, such as the B.1.351 (beta), P.1 (gamma), B.1.617.2 (delta), and B.1.427 (epsilon) variants, and additional variants will continue to arise. Some variants have shown decreased reactivity to neutralizing antibodies and immune escape, which means that individuals who have been vaccinated or have recovered from a prior COVID-19 infection may still be susceptible to infection by some of the emerging variants of concern.

SARS-CoV-2 utilizes a glycosylated Spike (S) protein for receptor-mediated entry into cells. Many studies have reported the cryo-electron microscopy or crystal structures of the entire SARS-CoV-2 virus (3), SARS-CoV-2 S trimer (4–6), the structure following furin cleavage (7), and RdRp (8), with D614G substitution (9) and angiontensin-converting enzyme 2 (ACE2) binding (10). Additionally, nsp7-nsp8 and nsp12-nsp7-nsp8 polymerase complexes (11, 12), a nsp7-nsp82-nsp12-nsp132-RNA complex (13), complex with a neutralizing antibody (14), and a nanobody-RBD complex (15) have also been reported. Several host factors are known to be involved in SARS-CoV-2 viral entry, including ACE2 (16), neurophilin-1 (17, 18), the two proteinases transmembrane serine protease 2 (TMPRSS2) (16) and cathepsin L (19), and the pro-protein convertase furin (20) among others (21–24). Single cell RNA-seq has been applied to extensively explore the expression of ACE2, which has been detected in a wide variety of cell types, including basal, ciliated, and secretory cells in lung bronchi, alveolar epithelial type 2 (AT2) cells in lung parenchyma, ciliated and secretory cells in the nasal cavity, endothelial cells, basal corneal epithelium, limbal niche, corneal wing cells, limbal superficial cells, superficial conjunctiva, corneal epithelial superficial cells in the cornea, enterocytes in ileum, enterocytes, and goblet cells in intestine, and pancreatic endocrine cells, such as beta cells (25–27). However, overall ACE2 expression in these tissues based on scRNA-seq analyses is low, which might reflect the low detection sensitivity of scRNA-seq platforms, as has been suggested (28).

Although COVID-19 has been associated with the dysfunction of many vital organs such as the lung, kidney, liver, heart, brain, and gastrointestinal system, the lung is the major target of SARS-CoV-2 and lung failure is the major cause of mortality in COVID-19 patients. SARS-CoV-2 infection of the respiratory epithelium causes severe cough, excessive mucous production, shortness of breath, chest tightness, and wheezing. Severe COVID-19 is marked by pneumonia and the progression to acute respiratory distress syndrome (ARDS) and respiratory failure requiring mechanical ventilation. One study found that 17% of COVID-19 patients developed ARDS and among these 65% rapidly worsened and died from multiple organ failure (29). Older age (>65 years old), diabetes mellitus, and hypertension have been associated with an increased risk of ARDS and respiratory failure (30). In this review, we discuss the pathologic features of lung injury and potential mechanisms, the in vitro and in vivo models to study SARS-CoV-2 infection, and the current status of drug development for COVID-19 (FIGURE 1).

FIGURE 1.

Comparison of different models for COVID-19 studies

SARS-CoV-2 and Lung Injury

Pathologic Features of Lung Epithelium

Autopsy reports of the lungs in COVID-19 patients have found diffuse alveolar damage (DAD) as the predominant lung pathology (31, 32). Histologically, all components of DAD, including injury to the alveolar epithelial cells, hyaline membrane formation, and hyperplasia of type II pneumocytes, have been detected in COVID-19 lung autopsy samples (33). In addition, immunohistochemical studies have shown the formation of interstitial thickening and fibrosis, intra-alveolar hemorrhaging, and denuded alveoli. Fibrin cluster formation and prominent presence of stromal cells in the walls of the thickened alveoli have also been observed (34). Tracheobronchitis was frequently present, which did not appear to depend on intubation or superimposed pneumonia (31).

Pathological Characterization of Lung Endothelium

Endothelial cell damage in the lung may be a leading mediator of COVID-19 severity (35). The alveolar microvasculature shows evidence of edema and thrombosis, with thrombi consisted of fibrin, platelets, and inflammatory cells. In patients who received anticoagulation therapy for thrombosis formation, postmortem analysis still revealed large thrombi, suggesting that antithrombotic treatment alone may not suffice (31). A study comparing autopsy samples of patients who died from COVID-19-associated or influenza-associated respiratory failure reported that the lungs from patients with COVID-19 also showed distinctive vascular features, consisting of severe endothelial injury associated with the presence of intracellular virus and disrupted cell membranes. Histologic analysis of pulmonary vessels in patients with COVID-19 showed widespread thrombosis with microangiopathy and a significantly increased amount of new vessel growth, predominantly through a mechanism of intussusceptive angiogenesis (36). Analysis of small pulmonary blood vessels in autopsy samples from patients who died following a severe COVID-19 infection revealed cytoplasmic swelling, vacuolization, and basement membrane reduplication (31).

In ARDS, injury and death of lung endothelial cells result in the breakdown of the lung endothelial barrier and the formation of protein-rich alveolar edema (37, 38), which in turn leads to hypoxia and respiratory distress. Activation of inflammatory signaling by cytokines such as interleukin-1β can downregulate the endothelial adherens junction protein VE-cadherin (39), which causes a loss of lung endothelial barrier integrity. The breakdown of endothelial adherens junctions in turn amplifies lung inflammation (40) thus creating a vicious cycle of lung endothelial barrier breakdown and inflammation.

Even during homeostasis, the lung endothelium is enriched for the expression of inflammatory and immune regulatory genes when compared to the endothelium of the lung or the heart, which is likely due to the continuous interaction of the lung with environmental pathogens (41). The increased expression levels of inflammatory genes in the lung endothelium even before any inflammatory insult may explain why the lung endothelium may play a critical role in amplifying the inflammatory response during SARS-CoV-2 infection and the pathogenesis of severe COVID-19.

A recent study observed marked downregulation of lung endothelial VE-cadherin and loss of lung endothelial barrier integrity following infection with the live SARS-CoV-2 virus in an animal model (42). Treatment with the IL-1-receptor antagonist anakinra prevented lung vascular leak and reduced overall mortality in the infected mice by 50% (42). There are also reports of noncytokine mediators that induce lung endothelial dysfunction in COVID-19. One study found increased levels of the glycosaminoglycan hyaluronan in critically ill COVID-19 patients and that exposure of lung endothelial cells to hyaluronan also resulted in a reduction of lung endothelial barrier integrity (43). These studies demonstrate the critical role of the lung endothelium in the progression of COVID-19 and the value of developing therapies that can prevent the breakdown of lung endothelial barrier integrity.

Immune Cells in COVID-19 Lung

Several studies have reported massive immune cell infiltration in the lungs during severe COVID-19, indicative of severe COVID-19 as a manifestation of a hyperinflammatory state in which the maladaptive immune response itself can cause lung issue injury. Diffuse inflammatory infiltrates consisting of interstitial and peribronchial lymphocytes and interalveolar macrophages were reported (44–46). Another autopsy study showed that severely damaged lungs from patients who died early had low interferon-stimulated genes, low viral loads, and abundant infiltrating activated CD8+ T cells and macrophages (47). Recently, a high-parameter imaging mass cytometry targeting the expression of 36 proteins unraveled the disordered structure of the infected and injured lung, alongside the distribution of extensive immune infiltration, at the single cell level. Neutrophil and macrophage infiltration are hallmarks of bacterial pneumonia and COVID-19, respectively (48). Although studies reported the detection of SARS-CoV-2 viral antigens in macrophages, there is lack of evidence of viral propagation in macrophages in vitro culture, suggesting that the presence of SARS-CoV-2 viral antigen in macrophages might be due to phagocytosis instead of active viral infection. Furthermore, lung autopsies of deceased COVID-19 patients have revealed neutrophil infiltration in pulmonary capillaries. Neutrophil extracellular traps, which are indicative of neutrophil activation, have also been described both in patient blood (49–51) and autopsy samples (52), highlighting the importance of hyperactivated neutrophils as a potential therapeutic target in severe COVID-19.

Lung Models to Study SARS-CoV-2 Infection

Although autopsy samples provide an abundance of useful information to understand the lung pathology of COVID-19 patients, most autopsy samples are collected several days/weeks after acute infection, which does not fully recapitulate the pathophysiology during the acute infection. Many models, such as cell lines, primary cells, and organoids for in vitro studies and animal models for in vivo studies, have been developed to discern the viral entry mechanisms, the viral life cycle, the injury mechanisms, and the preclinical evaluation of therapeutic candidates.

Cell Lines and Primary Cells

One of the most commonly used cell lines in virology is the Vero cell line, which was isolated from kidney epithelial cells extracted from an African green monkey in 1962. Vero E6, a subclone of Vero cells, is the standard cell line used to produce viral stocks of SARS-CoV-2 in different laboratories and to perform plaque assays. Additional lung cell lines, such as MRC-5 (human lung fibroblasts), MyDauLu/47.1 (a Daubenton’s bat lung cell line), RhiLu/1.1 (a horseshoe bat lung cell line), Calu-3 (a human lung cancer cell line), and A549 cells (a human lung cancer cell line) have been used to study SARS-CoV-2 infection. For example, Calu-3 cells were first used to demonstrate the SARS-CoV-2 entry factors, including ACE2, the serine protease TMPRSS2, as well as cathepsin B and L (16, 19). A similar study using Calu-3 cells and MRC-5 cells found that the cell entry of SARS-CoV-2 is preactivated by proprotein convertase furin, reducing its dependence on target cell proteases for cell entry (53). The high human (h)ACE2 binding affinity of the RBD, furin preactivation of the spike, and the hidden RBD in the spike protein may enable SARS-CoV-2 to efficiently enter cells while evading immune surveillance. Since A549 cells do not endogenously express the ACE2 receptor at a stable level, A549 cells overexpressing ACE2 have been used to the validate the role of ACE2 as a key entry receptor (54), identify the essential role of heparan sulfate during infection (55), examine the infectibility of different variants (56), perform genome-wide CRISPR-based screens to identify the key factors involved SARS-CoV-2 infection (57), and perform the in vitro evaluation of antibodies (58) and antiviral drugs (59, 60).

Although lung cancer cell lines provide easy and cost-efficient models to study SARS-CoV-2 infection, standard cancer cell lines fail to model the different cell types in lung affected by SARS-CoV-2 infection. In addition, most of these human cancer cell lines carry tumor-associated mutations, such as p53 mutations. p53 has been shown to regulate SARS-CoV-2 replication, which raises concerns for how these cancer cell lines recapitulate and phenocopy the viral biology and life cycle of SARS-CoV-2 in nontransformed cells (61). These cancer cell lines are typically unpolarized and hyperproliferative, which could impact mechanisms of viral entry and replication. Taken together, it seems likely that cells lines have key limitations when used to model SARS-CoV-2 infection.

In addition to cell lines, human primary airway epithelial cells, which highly express ACE2 and TMPRSS2, have also been used to study viral infection and host response to SARS-CoV-2 infection. An early report used human primary airway epithelial cells cultured on air-liquid phase to validate the viral infection and cytopathic effect (62). The human primary tracheal airway epithelial cells and small airway epithelial cells were further used to evaluate antiviral drug candidates (63). However, primary lung epithelial cells can be challenging to expand in culture and also exhibit significant patient-to-patient or batch-to-batch variability.

Lung Organoid Models

Organoids are self-organizing three-dimensional tissue cultures that show realistic micro-anatomy. Organoids can be derived from human pluripotent stem cells (hPSCs) or adult stem cells by mimicking human development or organ regeneration, respectively. Compared to cell lines or primary cells, organoids better recapitulate the heterogeneity, cell-cell interaction, and microenvironment of tissues or organs. In addition, the self-replication ability of organoids makes them suitable for large scale preparation required for disease modeling and drug screening.

Both hPSC and adult lung-derived lung organoids, including airway and alveolar organoids, have been used to investigate SARS-CoV-2 tropism, host response, and drug screening. Consistent with autopsy data, AT2 cells are the major cell type infected in alveolar organoids (64–66). In airway organoids, both ciliated cells and club cells have been reported to express ACE2 and are permissive to SARS-CoV-2 infection (66, 67). Single cell RNA-seq was applied in several studies to examine the entry factor expression in alveolar and airway organoids (64, 67), as well as the host response to SARS-CoV-2 infection at the single cell level (68). Several studies using alveolar and airway organoids reported the acute immune response after SARS-CoV-2 infection, such as the increase of chemokines and upregulation of IL-17, TNF (64), or NF-κB pathway (69), which recapitulate the transcriptomic signatures of COVID-19 patient autopsy samples. The infected AT2 cells lose mature cell identities and undergo apoptosis (70). hPSC-derived lung organoids were also used to identify a single nucleotide polymorphism (rs4702) that is located in the 3′-untranslated region of the protease furin, and its impact on SARS-CoV-2 infection (71). Another study using air-liquid interface culture of human airway basal stem cells found that direct cigarette smoke exposure increases SARS-CoV-2 infection and reduces the innate immune response (72). Furthermore, many studies have used alveolar and airway organoids to evaluate antiviral drugs or antibodies (73, 74). Interferons (IFNs) were shown to reduce viral replication in organoid models, suggesting the prophylactic effectiveness of IFN therapy against SARS-CoV-2 (57). Finally, hPSC-derived alveolar organoids were adapted to high throughput platform and screen for Food and Drug Administration (FDA)-approved drugs that block SARS-CoV-2 entry (64). Some drugs identified and validated using alveolar or airway organoid models have been either approved by FDA or are being evaluated in clinical trials. Expansion of the lung organoid model by integrating key cells involved in the pathogenesis of severe COVID-19, such as endothelial cells and immune cells, could further enhance the utility of organoids for modeling the disease and identifying novel therapeutics.

Animal Models to Study SARS-CoV-2 Infection

COVID-19 severity is variable based on the age, sex, gender, and preexisting conditions (75). To fully recapitulate the complex pathophysiology of disease, many animal models are being developed to study SARS-CoV-2 infection and evaluate therapeutic candidates in vivo. Mouse models show a low rate of viral infection by SARS-CoV-2 due to a lack of recognition of the mouse ACE2 receptor by the virus. This is likely due to the histidine at position 353 in mouse ACE2 that is a lysine residue in hACE2 (76). To curtail this problem, transgenic mouse models and human ACE2 knockin mouse models have been developed. The transgenic mice expressing the ACE2 receptor driven by the cytokeratin-18 gene promoter (K18-hACE2 mice) can be infected by SARS-CoV-2 and show key features of COVID-19 pathology such as the infiltration of monocytes, neutrophils, and other immune cells, as well as increased innate immune response pathways, such as NF-κB-dependent IFN signaling (77). Another transgenic mouse model with the endogenous Ace2 promoter-driven hACE2 expression has shown similar histopathological characteristics of COVID-19 (78). SARS-CoV-2 leads to robust replication in lung, trachea, and brain and causes interstitial pneumonia and elevated cytokine levels in aged human ACE2 knockin mice (79). A study using human ACE2 knockin mice showed SARS-CoV-2 spike D614G change enhances viral replication and transmission (80). In addition to modifying mice, several approaches have been used to adapt SARS-CoV-2 to mouse ACEs, including the sequential passaging of SARS-CoV-2 in mouse lung tissue (81) or using reverse genetics to modify the receptor binding domain of the virus (82). However, the mice infected by the adapted virus only developed mild symptoms.

In addition to transgenic or knockin mice, immunodeficient mice transplanted with human lung cells/xenografts have been developed to study the viral permissiveness and evaluate antiviral drugs. NSG mice transplanted with hPSC-derived lung progenitors form lung xenografts several months after transplantation and have been used to evaluate drugs blocking viral entry (64) and inhibiting SARS-CoV-2 replication (83).

The ferret is one of the first animals found to be susceptible to SARS-CoV-2 infection (84), as well as airborne transmission (85, 86). With the use of ferret models, an early study reported the imbalanced host response defined by low levels of type I and III interferons juxtaposed to elevated chemokines and high expression of IL-6 (87). After infection, ferrets also show clinically relevant features of COVID-19 such as epithelial cell necrosis and inflammatory infiltration in the bronchiolar lumina (88). In addition, ferrets have been used to evaluate drug candidates (89), antibodies, and vaccines (90).

Recently, the hamster model is also being used for SARS-CoV-2 studies because SARS-CoV-2 replicates efficiently in the lungs of hamsters. Immunohistochemistry assays demonstrated the presence of viral antigens on days 2 and 5 after inoculation with SARS-CoV-2, followed by rapid viral clearance and pneumocyte hyperplasia at 7 days after inoculation (91). In addition, SARS-CoV-2 infection causes severe lung injury that shares characteristics with SARS-CoV-2-infected human lung, including severe, bilateral and peripherally distributed multilobular ground glass opacities, and regions of lung consolidation (92). Moreover, Syrian hamster models have shown to mimic the age-dependent effects of COVID-19, such as weight loss and a decreased immune response in older hamsters (93). Hamsters have been used to explore the response to different SARS-CoV-2 variants (94, 95) and evaluate drug candidates (96–98), antibodies, or vaccines (99–101).

Finally, nonhuman primate models, such as rhesus macaques, cynomolgus macaques, and African green monkeys, were also used in COVID-19 studies. Studies using rhesus macaques showed high viral loads in the upper and lower respiratory tracts, humoral and cellular immune responses, and pneumonia (102–105), as well as endothelial disruption and vascular thrombosis in lungs (106). Cynomolgus macaques infected with SARS-CoV-2 showed body temperature rises and X-ray radiographic pneumonia without life-threatening clinical signs of disease (107). African green monkey models demonstrate robust SARS-CoV-2 replication and develop pronounced respiratory disease (108), as well as inflammation and coagulopathy in blood and tissues (109). Nonhuman primate models are critical models for the evaluation of drug candidates (110, 111), antibodies (107, 112–115), and vaccines (116–121).

Integrated Model of Lung Injury Mechanisms in COVID-19 Patients

The current studies on autopsy samples, cell lines, primary cells, organoids, and animal models suggest a multistep model of viral lung injury in COVID-19 patients, including direct viral infection and injury of cells, as well as subsequent lung injury induced by lung endothelial barrier breakdown, generation of excessive cytokines or chemokines, and immune cell infiltration resulting in a maladaptive immune response to the viral infection. First, AT2 cells in alveoli, ciliated cells, and club cells in airway are infected by SARS-CoV-2, which leads to decreased mature cell markers and increased cell death. As an acute response to SARS-CoV-2 infection, the infected cells release chemokines/cytokines, which can cause the death of neighboring cells and the breakdown of the lung endothelial barrier by downregulating the endothelial adherens junction protein VE-cadherin. Finally, the chemokines released after acute infection recruit immune cells, such as monocytes, which further differentiate into macrophages that secrete cytokines to induce host cell death (FIGURE 2). Key elements of these interactions were recently modeled by an immunohost in vitro coculture system (122).

FIGURE 2.

Potential mechanisms of lung damage in COVID-19 patients IFNs, interferons; AT2, alveolar epithelial type 2.

Drug Development

During the past year, immense effort has been applied to develop antiviral or anti-inflammation drugs for COVID-19 patients, including both small molecules and antibodies. Although the primary strategy to bring about an end of the COVID-19 pandemic is to increase the vaccination rates. However, vaccine hesitancy or lack of access to vaccines and the emergence of new virus variants necessitate the development of novel targeted COVID-19 therapies.

Antiviral Drugs

Several high-throughput screens using Vero cells, lung cancer cell lines, or alveolar/airway organoids have been developed to screen for anti-SARS-CoV-2 drugs. A high-throughput screen based on hPSC-derived lung organoids identified three entry inhibitors of SARS-CoV-2, including imatinib, mycophenolic acid, and quinacrine dihydrochloride (64). A virtual screening and a fluorogenic protease enzymatic assay based on the main protease of SARS-CoV-2 have been established to screen the protease inhibitors (123). In addition, a Vero E6 cell-based high-throughput drug repurposing screen identified the PIKfyve kinase inhibitor apilimod that antagonizes viral replication (124).

The antiviral compound remdesivir was developed for the treatment of the Ebola virus outbreak in 2017 and has shown some efficacy against SARS-CoV-2 (8). The drug works by inhibiting RNA-dependent RNA polymerase (125). Patients that received the treatment had a shortened recovery time from infection and a reduced mortality rate (126). Remdesivir has undergone robust clinical trials and has been approved by FDA; however, a recent randomized phase 3 trial in hospitalized COVID-19 patients showed no benefit of remdesivir treatment (127), suggesting that the patient target group that might benefit from remdesivir monotherapy may be narrower than previously thought.

Antibodies or Peptides Targeting SARS-CoV-2

Immediately responding to COVID-19 pandemic, huge amounts of effort have been made to develop antibodies against SARS-CoV-2, in particularly, the Spike protein. By targeting the spike protein on the surface of SARS-CoV-2, these antibodies block the ability of the virus to enter host cells and thus limit viral replication in both prophylactic and therapeutic settings as demonstrated in animal models (reviewed in Refs. 128, 129). The early clinical data have shown that some antibodies accelerate the natural decline in viral load, lead to faster viral clearance and reduce incidence of hospitalizations and deaths (130, 131). Three antibody-based therapies, including monotherapy (bamlanivimab) and combination (bamlanivimab and etesevimab) from Eli Lilly and combination therapy (casirivimab and imdevimab) from Regeneron, were authorized under Emergency Use Authorization for treatment of COVID-19 patients (FIGURE 3). Recently, bamlanivimab monotherapy has been revoked by FDA due to the increased frequency of resistant variants. An emerging phase 3 clinical trial reported that bamlanivimab plus etesevimab led to a lower incidence of COVID-19-related hospitalization and death than did placebo and accelerated the decline in the SARS-CoV-2 viral load, among high-risk ambulatory patients (132). Finally, a recent study used an engineered human ACE2 decoy peptide with a much higher affinity for the Spike protein than native ACE2 on the respiratory epithelium, thus preventing SARS-CoV-2 entry into cells (133). This engineered ACE2 decoy peptide is currently being evaluated in preclinical studies using distinct SARS-CoV-2 variants and could provide an alternative to antibody-based therapy.

FIGURE 3.

Food and Drug Administration-approved or Emergency Use Authorization treatment for COVID-19 patients IL-6R, interleukin-6 receptor; GM-CSF, granulocyte macrophage colony-stimulating factor; ACE2, angiotensin-converting enzyme 2; TNFSF14, tumor necrosis factor superfamily member 14; NRP1, neuropilin-1; RdRp, RNA-dependent RNA polymerase; ERGIC, endoplasmic reticulum-Golgi intermediate compartment; nsps, nonstructural proteins.

Anti-Inflammatory Drugs

Since inflammation and a cytokine storm are associated with high mortality rates, many anti-inflammation drugs are being evaluated in COVID-19 context. An immunocardiac model has identified drugs blocking immune cell-mediated host damage and identified tofacitinib, a Janus kinase (JAK) inhibitor, which protects cardiomyocytes from macrophage-induced toxicity (122). As JAK proteins are major signaling molecules for cytokines, JAK inhibition may aid in decreasing the intensity of a cytokine storm that causes ARDS in SARS-CoV-2 infected patients (134). A phase 3 clinical trial suggested that tofacitinib led to a lower risk of death or respiratory failure than placebo among patients hospitalized with COVID-19 pneumonia (135). Ruxolitinib, one of the JAK inhibitors, has previously been reported in a nonrandomized trial to be efficacious in reducing mortality among hospitalized patients with severe COVID-19 (136); however, it failed to meet its primary end point of statistical significance in mortality in a randomized, double-blind, placebo-controlled phase 3 trial in patients on mechanical ventilation with COVID-19-associated ARDS (137). Baricitinib plus remdesivir was superior to remdesivir alone in reducing recovery time and accelerating improvement in clinical status among patients with COVID-19, notably among those receiving high-flow oxygen or noninvasive ventilation (138). The FDA authorized emergency use of baricitinib plus remdesivir for COVID-19 in November 2020.

In addition to the drugs approved or authorized for emergency use by FDA, several drugs now show promising results in clinical trials. For example, lenzilumab, a recombinant monoclonal antibody against the cytokine granulocyte macrophage colony-stimulating factor, was shown to significantly improve ventilator-free survival in hospitalized, hypoxic subjects with COVID-19 pneumonia in a phase 3 randomized, double-blind, placebo-controlled trial (139). CERC-002, a human anti-LIGHT or tumor necrosis factor superfamily member 14 monoclonal antibody, reduced the risk of respiratory failure and death incremental to standard of care including high dose corticosteroids and reduces LIGHT levels in patients with COVID-19 ARDS in a randomized, double-blind, placebo-controlled, multicenter, proof-of-concept clinical trial (140). A randomized, double-blind, placebo-controlled, phase 2 trial showed that patients who received SNG001 (inhaled nebulized interferon beta-1a) had greater odds of improvement and recovered more rapidly from SARS-CoV-2 infection than patients who received placebo (141). Very recently, Merck announced results from an interim analysis of data from 775 patients in the phase III portion of the MOVe-OUT trial, which showing an ∼50% reduction in the risk of hospitalization or death in patients receiving molnupiravir, an oral antiviral candidate that exerts its antiviral action through introduction of copying errors during viral RNA replication (142).

Future Perspective

After ∼18 mo from the beginning of the COVID-19 pandemic, significant progress has been made in terms of developing vaccines and using COVID-19 disease models to identify novel therapies. The treatment landscape is rapidly evolving (Table 1). However, there are still challenges remaining to effective and rapid response to emerging SARS-CoV-2 variants as well as future pandemics. First, we need to continue developing better in vivo and in vitro models that can be used rapidly and efficiently to study coronavirus infections. In this pandemic, organoids have been used as emerging models that allowed for the investigation of host responses and screening for therapeutics. However, most of current organoid models lack vascular structures as well as immune cells, both of which play key roles in disease progression. Second, most newly developed drugs are antibodies, which are expensive and may not adequately neutralize new viral variants. There is a growing demand for cost-efficient pan-antiviral drugs. Last, the pandemic has shown that basic science can indeed be rapidly translated into clinical trials. Currently, more than 6,700 COVID-19-related clinical trials are registered at ClinicalTrials.gov. However, there is a need for better integration of basic science findings with clinical trials to increase the success rate of the clinical trials.

TABLE 1.

Current treatment for COVID-19 patients

Drug	Drug Type	Phase	Proposed Mechanism
Veklury (remdesivir)	Small molecule	Approved	Inhibit RNA-dependent RNA polymerase
Olumiant (baricitinib) + Veklury (remdesivir)	Small molecule	EUA	JAK inhibitor
Actemra (tocilizumab)	Antibody	EUA	Binds to IL-6 receptor
Bamlanivimab+ etesevimab	Antibody	EUA	Binds to the SARS-CoV-2 surface spike protein receptor binding domain
Casirivimab and imdevimab	Antibody	EUA	Bind to different sites on the receptor binding domain of the spike protein of SARS-CoV-2
Sotrovimab	Antibody	EUA	Bind to different sites on the receptor binding domain of the spike protein of SARS-CoV-2

EUA, Emergency Use Authorization.