key: cord-0912601-h3lzl1un authors: Banerjee, Aditi; Czinn, Steven J.; Reiter, Russel J.; Blanchard, Thomas G. title: Crosstalk between endoplasmic reticulum stress and anti-viral activities: A novel therapeutic target for COVID-19 date: 2020-05-23 journal: Life Sci DOI: 10.1016/j.lfs.2020.117842 sha: ff502c1833ce7652641132d10251370440456a25 doc_id: 912601 cord_uid: h3lzl1un The outbreak of COVID-19 caused by 2019–nCov/SARS-CoV-2 has become a pandemic with an urgent need for understanding the mechanisms and identifying a treatment. Viral infections including SARS-CoV are associated with increased levels of reactive oxygen species, disturbances of Ca(++) caused by unfolded protein response (UPR) mediated by endoplasmic reticulum (ER) stress and is due to the exploitation of virus's own protein i.e., viroporins into the host cells. Several clinical trials are on-going including testing Remdesivir (anti-viral), Chloroquine and Hydroxychloroquine derivatives (anti-malarial drugs) etc. Unfortunately, each drug has specific limitations. Herein, we review the viral protein involvement to activate ER stress transducers (IRE-1, PERK, ATF-6) and their downstream signals; and evaluate combination therapies for COVID-19 mediated ER stress alterations. Melatonin is an immunoregulator, anti-pyretic, antioxidant, anti-inflammatory and ER stress modulator during viral infections. It enhances protective mechanisms for respiratory tract disorders. Andrographolide, isolated from Andrographis paniculata, has versatile biological activities including immunomodulation and determining SARS-CoV-2 binding site. Considering the properties of both compounds in terms of anti-inflammatory, antioxidant, anti-pyrogenic, anti-viral and ER stress modulation and computational approaches revealing andrographolide docks with the SARS-CoV2 binding site, we predict that this combination therapy may have potential utility against COVID-19. Viral diseases continue to emerge and represent a serious issue to public health [1], Coronavirus is part of a family of enveloped viruses with positive sense non-segmented single-stranded RNA genomes. There are two human α-corona viruses, HCoV-229E and HCoV-NL63 and two β-corona viruses, HCoV-OC43 and HCoV-HKU1. Among these, HCoV-NL63 and HCoV-HKU1 have been identified as SARS-CoV and account for the recent outbreaks. These viruses are endemic in the human populations, causing 15-30% of respiratory tract infections each year [2] . In December 2019, a novel strain of the 2019-nCov/SARS-CoV-2, a β-coronavirus, emerged in Wuhan, Hubei province, China. This etiologic agent of this new lung disease, COVID-19 caused by SARS-CoV-2, poses a global health emergency affecting millions of lives worldwide [3, 4] . Recent studies demonstrated the crystal structure of CR3022, a neutralizing antibody isolated from a convalescent SARS patient. This antibody targets a highly conserved epitope, distal from the receptor-binding site, that enables cross-reactive binding between SARS-CoV-2 and SARS-CoV [5] . On March 11, the COVID-19 outbreak was characterized as a pandemic by the WHO [6] . As of April 2020, this pandemic has taken the lives of ~250 k people and infected 3.5 M individuals worldwide. In the US, ~1.2 M laboratory confirmed infectious were reported including more than 65 k deaths. There is an urgent need for the development of an effective mechanism to treat and prevent 2019--nCov/SARS-CoV-2 outbreaks. In this review article, we provide an indication of future research in order to understand the molecular mechanism related to COVID-19 and possible drug targets to regulate the impact of this viral infection. According to the Centers for Disease Control and Prevention (CDC), people with COVID-19 have had a wide range of symptoms ranging from mild to severe illness and which may appear within 2-14 days after exposure to the virus. The high risk of fatality due to COVID-19 is a consequence of age-associated conditions, such as cardiovascular, pulmonary, and diabetic disorders as well as immune-compromised conditions [7, 8] . From the perspective of cell biology, COVID-19 can be divided into three phases that correspond to different clinical stages of the disease. The stages are J o u r n a l P r e -p r o o f 3 as follows: i. Asymptomatic state (initial 1-2 days of infection). In this stage the inhaled virus binds to the receptor of angiotensin converting enzyme 2; (ACE2) on epithelial cells in the nasal cavity and replicates. ii) Upper airway and conducting airway response (next few days). The virus propagates and migrates down the respiratory tract along the conducting airways, and a more robust innate immune response is triggered. For about 80% of the infected patients, the disease will be mild and mostly restricted to the upper and conducting airways. iii) Hypoxia, ground glass infiltrates and progression to ARDS (Acute Respiratory Distress Syndrome). Unfortunately, about 20% of the infected patients will progress to stage 3 disease and will develop pulmonary infiltrates and some of these will develop severe disease. The pathological results of SARS and COVID-19 are diffuse alveolar damage with fibrin rich hyaline membranes and a few multinucleated giant cells [9, 10] In eukaryotic cells, one of the largest organelles, the endoplasmic reticulum (ER) is the site of synthesis and folding of membrane, secretory proteins, lipids, sterols, and storage of free calcium [11] . Alterations of protein folding in the ER due to physiological stress such as disturbances in redox, Ca ++ levels, glycosylation or other environmental elements cause accumulation of misfolded proteins leading to ER stress. The increased levels of reactive oxygen species (ROS) triggered by ER stress activate not only proinflammatory signals but also inflammasome formation, suggesting that ER stress exerts immunogenic effects [12] and can be activated by excessive lipids or proinflammatory cytokines [13] . As a result, a series of signal transduction cascades or an unfolded protein response (UPR) occurs. The hallmark of the UPR is the expression of ER-resident chaperones, such as immunoglobulin heavy chain binding protein (BiP/GRP78) and glucose-regulated protein 94 (GRP94). In addition, PERK, IRE-1, and ATF-6 serve as proximal sensors which regulate components that upregulate the capacity of the ER to fold newly synthesized proteins and degrade misfolded/unfolded proteins [14] . In addition, UPR is associated with several major cellular activities J o u r n a l P r e -p r o o f including apoptosis, angiogenesis, autophagy, the mitogen-activated protein (MAP) kinase pathways, innate immunity, and pro-inflammatory response. Accumulating evidence suggests that ER stress and sustained UPR signaling are major contributors to the pathogenesis of several diseases, including inflammatory disorders and viral infections [15] and can increase the severity of these events [16] . Viruses may interact with the host UPR to maintain an environment favorable for establishment of persistent infection [17] . The mechanism is the imbalance of calcium concentration by the expression of viroporins, small virally encoded hydrophobic proteins that oligomerize in the membrane of host cells. This leads to the formation of hydrophilic pores, and consequent depletion of ER membrane due to the release of virions [18] which cause ER stress in the host cells by generating large amounts of unfolded or misfolded proteins [19] . It is well documented that the replication of corona virus occurs in the cytoplasm and is strongly associated with ER and its transducers. In brief, cells infected with SARS-CoV or cells overexpressing the SARS-CoVS2 subunit showed increased levels of GRP94 (ER stress associated gene) and GRP78 gene expression. Like GRP, a significant phosphorylation of PKR and PERK has been observed in SARS-CoV infected cells [20] . Several well-documented studies have shown that melatonin has a protective role in infections induced by encephalitis virus due to its activity in the central nervous system, associated with its capability to regulate immune function [33] . Another study also confirmed its protective mechanism in bronchiolitis, a severe inflammatory lower respiratory tract disorder mediated by RSV (Respiratory syncytial virus) infection [34] . It is suggested that respiratory disorders induced by many other human pathogens may result from an exuberant generation of reactive oxygen species by inflammatory cells in response to infection [35] . A recent review article documented melatogenergic pathway's role in viral infections, emphasizing influenza and COVID-19 infections. Therefore, melatonin has the potential to be a therapeutic target of COVID-19 infection due to its anti-inflammation, anti-oxidation, and immune enhancing properties [36] . J o u r n a l P r e -p r o o f 6 Melatonin modulates ER stress and activates UPR response during viral infections. Due to its antioxidant properties, it regulates ER stress and controls autophagic and apoptotic processes. An earlier study showed that melatonin reduces macrophage inflammation by controlling the ER stress associated signaling pathways [37] . During RHDV (rabbit hemorrhagic disease virus) infection, melatonin induced a decrease in the autophagy associated with this infection and inhibited RHDV RNA replication. The molecular mechanism involved an interplay of RHDV-induced autophagy with oxidative stress, ER stress and apoptosis [38] . A recent study illustrated that melatonin treatment attenuated viral myocarditis via sustaining cardiomyocyte viability, repressing mitochondrial dysfunction and inhibiting ER stress [39] . The rationale for using melatonin in viral diseases is supported by its capability to modulate UPR during viral infection due to its immune enhancing actions, anti-inflammatory and antioxidant properties. Andrographolide is a lactone (bicyclic diterpenoid) derived from Andrographis paniculata [40] . Like melatonin, it has several biological activities including anti-carcinogenic [41] [42] [43] [44] , anti-inflammatory [45, 46] , immunomodulator [47] , antioxidant [48] [49] [50] , antipyrogenic [51] and anti-viral properties [52] [53] [54] [55] [56] [57] . Andrographolide induces ER stress leading to cancer cell death due to apoptosis through the induction of ROS [48] , which can inhibit virus-induced carcinogenesis. Additional inhibitory effects of andrographolide include that of cell migration, invasion, matrix metalloproteinase expression, antiangiogenesis, autophagy, and dysregulation of signaling pathway has been reported for inflammatory disorders including cancer [41, 50, 58, 59] . Upregulation of CTLs and NK cell activity has been found after andrographolide treatment [47] which demonstrates its [51, 56, 57, [61] [62] [63] [64] [65] [66] [67] [68] [69] [70] [71] [72] . Recent studies showed that andrographolide is a potential inhibitor of the main protease of SARS-CoV-2 through in silico studies, such as molecular docking, target analysis, toxicity prediction and ADME prediction (absorption, distribution, metabolism, and excretion) [57] . The molecular mechanisms of the antiviral properties of andrographolide are as follows: 1). Enhanced H1N1 virus-I, induced cell death through the inhibition of viral-induced activation of the retinoic acid-inducible gene I (RIG-I)-like receptors (RLRs) signaling pathway [53] and diminished lung virus titer through its immune-modulatory activity [51] 2). Alteration of ER stress mediated UPR pathway on virus replication pathway [55, 73] . 3) Induction of heme oxygenase 1 (HO-1) expression [74, 75] . 4) Involvement of multiple pathway including NFkβ and JAK-STAT. Hydroxychloroquine derivatives are being used in emergency cases; however, they are not suitable for patients with conditions such as diabetes, hypertension and cardiac issues [81] . Social isolation is currently the best way to manage the spread of COVID-19 in the absence of an effective treatment. It is revealed that Remdesivir, a drug thought to be one of the best prospects for treating COVID-19, has severe side effects, leading to its discontinuation in trial. Therefore, a novel combination therapy drug with immunomodulators might be a promising therapeutic approach for COVID-19. available at a low cost. However, previous reports of adverse reactions to andrographolide in a phase I clinical trial [71] suggests that a combination therapy of andrographolide and melatonin could unveil a potentially useful treatment for COVID- 19 . Melatonin has been shown to protect against the toxicity of a variety of drugs and toxins. This may increase the efficacy of the combined therapy. We wish to thank the Department of Pediatrics at the University of Maryland School of Medicine for support. We acknowledge Vivekjyoti Banerjee for creating the corona virus graphic using Blender software. A.B wrote the original draft, T.G.B., S. J. C. and R. J. R reviewed, and edited the manuscript. The authors declare that they have no competing interests. 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