Nafamostat Mesylate Monotherapy in Patients with Moderate COVID-19: a Single-Center, Retrospective Study.

Coronavirus disease (COVID-19) has spread dramatically worldwide. Nafamostat mesylate inhibits intracellular entry of the novel severe acute respiratory syndrome coronavirus 2 and is believed to have therapeutic potential for treating patients with COVID-19. In this study, patients with moderate COVID-19 who were admitted to our hospital were retrospectively analyzed. Thirty-one patients received monotherapy with nafamostat mesylate, and 33 patients were treated conservatively. Nafamostat mesylate was administered with continuous intravenous infusion for an average of 4.5 days. Compared with the conservative treatment, nafamostat mesylate did not improve outcomes or laboratory data 5 days after admission. In addition, no significant differences in laboratory data 5 days after admission and outcomes in high-risk patients were observed. The incidence of hyperkalemia was significantly higher in the nafamostat mesylate group; however, none of the patients required additional treatment. In conclusion, monotherapy with nafamostat mesylate did not improve clinical outcomes in patients with moderate COVID-19. This study did not examine the therapeutic potential of combining nafamostat mesylate with other antiviral agents, and further investigation is required. Because of the high incidence of hyperkalemia, regular laboratory monitoring is required during the use of nafamostat mesylate.

Nafamostat Mesylate for Treatment of COVID-19 in Hospitalised Patients: A Structured, Narrative Review.

The search for clinically effective antivirals against the severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) is ongoing. Repurposing of drugs licensed for non-coronavirus disease 2019 (COVID-19) indications has been extensively investigated in laboratory models and in clinical studies with mixed results. Nafamostat mesylate (nafamostat) is a drug licensed in Japan and Korea for indications including acute pancreatitis and disseminated intravascular coagulation. It is available only for continuous intravenous infusion. In vitro human lung cell line studies with nafamostat demonstrate high antiviral potency against SARS-CoV-2 (half maximal inhibitory concentration [IC50] of 0.0022 µM [compared to remdesivir 1.3 µM]), ostensibly via inhibition of the cellular enzyme transmembrane protease serine 2 (TMPRSS2) preventing viral entry into human cells. In addition, the established antithrombotic activity is hypothesised to be advantageous given thrombosis-associated sequelae of COVID-19. Clinical reports to date are limited, but indicate a potential benefit of nafamostat in patients with moderate to severe COVID-19. In this review, we will explore the pre-clinical, pharmacokinetic and clinical outcome data presently available for nafamostat as a treatment for COVID-19. The recruitment to ongoing clinical trials is a priority to provide more robust data on the safety and efficacy of nafamostat as a treatment for COVID-19.

A SARS-CoV-2 cytopathicity dataset generated by high-content screening of a large drug repurposing collection.

SARS-CoV-2 is a novel coronavirus responsible for the COVID-19 pandemic, in which acute respiratory infections are associated with high socio-economic burden. We applied high-content screening to a well-defined collection of 5632 compounds including 3488 that have undergone previous clinical investigations across 600 indications. The compounds were screened by microscopy for their ability to inhibit SARS-CoV-2 cytopathicity in the human epithelial colorectal adenocarcinoma cell line, Caco-2. The primary screen identified 258 hits that inhibited cytopathicity by more than 75%, most of which were not previously known to be active against SARS-CoV-2 in vitro. These compounds were tested in an eight-point dose response screen using the same image-based cytopathicity readout. For the 67 most active molecules, cytotoxicity data were generated to confirm activity against SARS-CoV-2. We verified the ability of known inhibitors camostat, nafamostat, lopinavir, mefloquine, papaverine and cetylpyridinium to reduce the cytopathic effects of SARS-CoV-2, providing confidence in the validity of the assay. The high-content screening data are suitable for reanalysis across numerous drug classes and indications and may yield additional insights into SARS-CoV-2 mechanisms and potential therapeutic strategies.

COVID-19-associated coagulopathy and disseminated intravascular coagulation.

The pathology of coronavirus disease 2019 (COVID-19) is exacerbated by the progression of thrombosis, and disseminated intravascular coagulation (DIC), and cytokine storms. The most frequently reported coagulation/fibrinolytic abnormality in COVID-19 is the increase in D-dimer, and its relationship with prognosis has been discussed. However, limits exist to the utility of evaluation by D-dimer alone. In addition, since the coagulation/fibrinolytic condition sometimes fluctuates within a short period of time, regular examinations in recognition of the significance of the examination are desirable. The pathophysiology of disseminated intravascular coagulation (DIC) associated with COVID-19 is very different from that of septic DIC, and both thrombotic and hemorrhagic pathologies should be noted. COVID-19 thrombosis includes macro- and microthrombosis, with diagnosis of the latter depending on markers of coagulation and fibrinolysis. Treatment of COVID-19 is classified into antiviral treatment, cytokine storm treatment, and thrombosis treatment. Rather than providing uniform treatment, the treatment method most suitable for the severity and stage should be selected. Combination therapy with heparin and nafamostat is expected to develop in the future. Fibrinolytic therapy and adsorption therapy require further study.

COVID-19-associated coagulopathy and disseminated intravascular coagulation.

The pathology of coronavirus disease 2019 (COVID-19) is exacerbated by the progression of thrombosis, and disseminated intravascular coagulation (DIC), and cytokine storms. The most frequently reported coagulation/fibrinolytic abnormality in COVID-19 is the increase in D-dimer, and its relationship with prognosis has been discussed. However, limits exist to the utility of evaluation by D-dimer alone. In addition, since the coagulation/fibrinolytic condition sometimes fluctuates within a short period of time, regular examinations in recognition of the significance of the examination are desirable. The pathophysiology of disseminated intravascular coagulation (DIC) associated with COVID-19 is very different from that of septic DIC, and both thrombotic and hemorrhagic pathologies should be noted. COVID-19 thrombosis includes macro- and microthrombosis, with diagnosis of the latter depending on markers of coagulation and fibrinolysis. Treatment of COVID-19 is classified into antiviral treatment, cytokine storm treatment, and thrombosis treatment. Rather than providing uniform treatment, the treatment method most suitable for the severity and stage should be selected. Combination therapy with heparin and nafamostat is expected to develop in the future. Fibrinolytic therapy and adsorption therapy require further study.

Unravelling high-affinity binding compounds towards transmembrane protease serine 2 enzyme in treating SARS-CoV-2 infection using molecular modelling and docking studies.

The coronavirus disease-19 (COVID-19) outbreak that is caused by a highly contagious severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) has become a zoonotic pandemic, with approximately 24.5 million positive cases and 8.3 lakhs deaths globally. The lack of effective drugs or vaccine provoked the research for drug candidates that can disrupt the spread and progression of the virus. The identification of drug molecules through experimental studies is time-consuming and expensive, so there is a need for developing alternative strategies like in silico approaches which can yield better outcomes in less time. Herein, we selected transmembrane protease serine 2 (TMPRSS2) enzyme, a potential pharmacological target against SARS-CoV-2, involved in the spread and pathogenesis of the virus. Since 3D structure is not available for this protein, the present study aims at homology modelling and validation of TMPRSS2 using Swiss-model server. Validation of the modelled TMPRSS2 using various online tools confirmed model accuracy, topology and stereochemical plausibility. The catalytic triad consisting of Serine-441, Histidine-296 and Aspartic acid-345 was identified as active binding site of TMPRSS2 using existing ligands. Molecular docking studies of various drugs and phytochemicals against the modelled TMPRSS2 were performed using camostat as a standard drug. The results revealed eight potential drug candidates, namely nafamostat, meloxicam, ganodermanontriol, columbin, myricetin, proanthocyanidin A2, jatrorrhizine and baicalein, which were further studied for ADME/T properties. In conclusion, the study unravelled eight high affinity binding compounds, which may serve as potent antagonists against TMPRSS2 to impact COVID-19 drug therapy.

Molecular Insights into Human Transmembrane Protease Serine-2 (TMPS2) Inhibitors against SARS-CoV2: Homology Modelling, Molecular Dynamics, and Docking Studies.

The severe acute respiratory syndrome coronavirus 2 (SARS-CoV2), which caused novel corona virus disease-2019 (COVID-19) pandemic, necessitated a global demand for studies related to genes and enzymes of SARS-CoV2. SARS-CoV2 infection depends on the host cell Angiotensin-Converting Enzyme-2 (ACE2) and Transmembrane Serine Protease-2 (TMPRSS2), where the virus uses ACE2 for entry and TMPRSS2 for S protein priming. The TMPRSS2 gene encodes a Transmembrane Protease Serine-2 protein (TMPS2) that belongs to the serine protease family. There is no crystal structure available for TMPS2, therefore, a homology model was required to establish a putative 3D structure for the enzyme. A homology model was constructed using SWISS-MODEL and evaluations were performed through Ramachandran plots, Verify 3D and Protein Statistical Analysis (ProSA). Molecular dynamics simulations were employed to investigate the stability of the constructed model. Docking of TMPS2 inhibitors, camostat, nafamostat, gabexate, and sivelestat, using Molecular Operating Environment (MOE) software, into the constructed model was performed and the protein-ligand complexes were subjected to MD simulations and computational binding affinity calculations. These in silico studies determined the tertiary structure of TMPS2 amino acid sequence and predicted how ligands bind to the model, which is important for drug development for the prevention and treatment of COVID-19.

Dynamic changes in fibrinogen and D-dimer levels in COVID-19 patients on nafamostat mesylate.

Critical illnesses associated with coronavirus disease 2019 (COVID-19) are attributable to a hypercoagulable status. There is limited knowledge regarding the dynamic changes in coagulation factors among COVID-19 patients on nafamostat mesylate, a potential therapeutic anticoagulant for COVID-19. First, we retrospectively conducted a cluster analysis based on clinical characteristics on admission to identify latent subgroups among fifteen patients with COVID-19 on nafamostat mesylate at the University of Tokyo Hospital, Japan, between April 6 and May 31, 2020. Next, we delineated the characteristics of all patients as well as COVID-19-patient subgroups and compared dynamic changes in coagulation factors among each subgroup. The subsequent dynamic changes in fibrinogen and D-dimer levels were presented graphically. All COVID-19 patients　were classified into three subgroups: clusters A, B, and C, representing low, intermediate, and high risk of poor outcomes, respectively. All patients were alive 30 days from symptom onset. No patient in cluster A required mechanical ventilation; however, all patients in cluster C required mechanical ventilation, and half of them were treated with venovenous extracorporeal membrane oxygenation. All patients in cluster A maintained low D-dimer levels, but some critical patients in clusters B and C showed dynamic changes in fibrinogen and D-dimer levels. Although the potential of nafamostat mesylate needs to be evaluated in randomized clinical trials, admission characteristics of patients with COVID-19 could predict subsequent coagulopathy.

High levels of anti-SSA/Ro antibodies in COVID-19 patients with severe respiratory failure: a case-based review : High levels of anti-SSA/Ro antibodies in COVID-19.

We treated two patients with severe respiratory failure due to coronavirus disease 2019 (COVID-19). Case 1 was a 73-year-old woman, and Case 2 was a 65-year-old-man. Neither of them had a history of autoimmune disease. Chest computed tomography scans before the antiviral therapy showed bilateral multiple patchy ground-glass opacities (GGO) consistent with COVID-19 pneumonia. The GGO regressed over the course of the antiviral treatment; however, new non-segmental patchy consolidations emerged, which resembled those of interstitial lung disease (ILD), specifically collagen vascular disease-associated ILD. We tested the patients' sera for autoantibodies and discovered that both patients had high anti-SSA/Ro antibody titers. In Case 1, the patient recovered with antiviral therapy alone. However, in Case 2, the patient did not improve with antiviral therapy alone but responded well to corticosteroid therapy (methylprednisolone) and made a full recovery. The relationship between some immunological responses and COVID-19 pneumonia exacerbation has been discussed previously; our discovery of the elevation of anti-SSA/Ro antibodies suggests a contribution from autoimmunity functions of the immune system. Although it is unclear whether the elevation of anti-SSA/Ro antibodies was a cause or an outcome of aggravated COVID-19 pneumonia, we hypothesize that both patients developed aggravated the COVID-19 pneumonia due to an autoimmune response. In COVID-19 lung injury, there may be a presence of autoimmunity factors in addition to the known effects of cytokine storms. In patients with COVID-19, a high level of anti-SSA/Ro52 antibodies may be a surrogate marker of pneumonia severity and poor prognosis.

Clinical improvement in a patient with severe coronavirus disease 2019 after administration of hydroxychloroquine and continuous hemodiafiltlation with nafamostat mesylate.

The number of people infected with severe acute respiratory syndrome coronavirus 2 is increasing globally, and some patients have a fatal clinical course. In light of this situation, the World Health Organization (WHO) declared coronavirus disease 2019 (COVID-19) a pandemic on March 11, 2020. While clinical studies and basic research on a treatment for COVID-19 are ongoing around the world, no treatment has yet been proven to be effective. Several clinical studies have demonstrated the efficacy of chloroquine phosphate and nafamostat mesylate with COVID-19. Here, we report the case of a Japanese patient with COVID-19 with severe respiratory failure who improved following the administration of hydroxychloroquine and continuous hemodiafiltlation with nafamostat mesylate. Hence, hydroxychloroquine with nafamostat mesylate might be a treatment option for severe COVID-19.