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Protein & Cell Jan 2023The global COVID-19 coronavirus pandemic has infected over 109 million people, leading to over 2 million deaths up to date and still lacking of effective drugs for...
The global COVID-19 coronavirus pandemic has infected over 109 million people, leading to over 2 million deaths up to date and still lacking of effective drugs for patient treatment. Here, we screened about 1.8 million small molecules against the main protease (M) and papain like protease (PL), two major proteases in severe acute respiratory syndrome-coronavirus 2 genome, and identified 1851M inhibitors and 205 PL inhibitors with low nmol/l activity of the best hits. Among these inhibitors, eight small molecules showed dual inhibition effects on both M and PL, exhibiting potential as better candidates for COVID-19 treatment. The best inhibitors of each protease were tested in antiviral assay, with over 40% of M inhibitors and over 20% of PL inhibitors showing high potency in viral inhibition with low cytotoxicity. The X-ray crystal structure of SARS-CoV-2 M in complex with its potent inhibitor 4a was determined at 1.8 Å resolution. Together with docking assays, our results provide a comprehensive resource for future research on anti-SARS-CoV-2 drug development.
Topics: Humans; Antiviral Agents; COVID-19; COVID-19 Drug Treatment; High-Throughput Screening Assays; Molecular Docking Simulation; Protease Inhibitors; SARS-CoV-2; Viral Nonstructural Proteins
PubMed: 36726755
DOI: 10.1093/procel/pwac016 -
Viruses Sep 2023Nirmatrelvir, which targets the SARS-CoV-2 main protease (Mpro), is the first-in-line drug for prevention and treatment of severe COVID-19, and additional Mpro... (Review)
Review
Nirmatrelvir, which targets the SARS-CoV-2 main protease (Mpro), is the first-in-line drug for prevention and treatment of severe COVID-19, and additional Mpro inhibitors are in development. However, the risk of resistance development threatens the future efficacy of such direct-acting antivirals. To gain knowledge on viral correlates of resistance to Mpro inhibitors, we selected resistant SARS-CoV-2 under treatment with the nirmatrelvir-related protease inhibitor boceprevir. SARS-CoV-2 selected during five escape experiments in VeroE6 cells showed cross-resistance to nirmatrelvir with up to 7.3-fold increased half-maximal effective concentration compared to original SARS-CoV-2, determined in concentration-response experiments. Sequence analysis revealed that escape viruses harbored Mpro substitutions L50F and A173V. For reverse genetic studies, these substitutions were introduced into a cell-culture-infectious SARS-CoV-2 clone. Infectivity titration and analysis of genetic stability of cell-culture-derived engineered SARS-CoV-2 mutants showed that L50F rescued the fitness cost conferred by A173V. In the concentration-response experiments, A173V was the main driver of resistance to boceprevir and nirmatrelvir. Structural analysis of Mpro suggested that A173V can cause resistance by making boceprevir and nirmatrelvir binding less favorable. This study contributes to a comprehensive overview of the resistance profile of the first-in-line COVID-19 treatment nirmatrelvir and can thus inform population monitoring and contribute to pandemic preparedness.
Topics: Humans; Protease Inhibitors; Antiviral Agents; SARS-CoV-2; COVID-19 Drug Treatment; COVID-19; Hepatitis C, Chronic; Enzyme Inhibitors; Anti-Infective Agents; Lactams
PubMed: 37766376
DOI: 10.3390/v15091970 -
Marine Drugs Dec 2019Serine proteases play pivotal roles in normal physiology and a spectrum of patho-physiological processes. Accordingly, there is considerable interest in the discovery...
Serine proteases play pivotal roles in normal physiology and a spectrum of patho-physiological processes. Accordingly, there is considerable interest in the discovery and design of potent serine protease inhibitors for therapeutic applications. This led to concerted efforts to discover versatile and robust molecular scaffolds for inhibitor design. This investigation is a bioprospecting study that aims to isolate and identify protease inhibitors from the cnidarian The study isolated two Kunitz-type protease inhibitors with very similar sequences but quite divergent inhibitory potencies when assayed against bovine trypsin, chymostrypsin, and a selection of human sequence-related peptidases. Homology modeling and molecular dynamics simulations of these inhibitors in complex with their targets were carried out and, collectively, these methodologies enabled the definition of a versatile scaffold for inhibitor design. Thermal denaturation studies showed that the inhibitors were remarkably robust. To gain a fine-grained map of the residues responsible for this stability, we conducted in silico alanine scanning and quantified individual residue contributions to the inhibitor's stability. Sequences of these inhibitors were then used to search for Kunitz homologs in an transcriptome library, resulting in the discovery of a further 14 related sequences. Consensus analysis of these variants identified a rich molecular diversity of Kunitz domains and expanded the palette of potential residue substitutions for rational inhibitor design using this domain.
Topics: Animals; Cattle; Chymotrypsin; Cnidaria; Computer Simulation; Humans; Molecular Dynamics Simulation; Serine Proteases; Serine Proteinase Inhibitors; Trypsin; Trypsin Inhibitors
PubMed: 31842369
DOI: 10.3390/md17120701 -
International Journal of Molecular... Apr 2022The action of proteases can be controlled by several mechanisms, including regulation through gene expression; post-translational modifications, such as glycosylation;... (Review)
Review
The action of proteases can be controlled by several mechanisms, including regulation through gene expression; post-translational modifications, such as glycosylation; zymogen activation; targeting specific compartments, such as lysosomes and mitochondria; and blocking proteolysis using endogenous inhibitors. Protease inhibitors are important molecules to be explored for the control of proteolytic processes in organisms because of their ability to act on several proteases. In this context, plants synthesize numerous proteins that contribute to protection against attacks by microorganisms (fungi and bacteria) and/or invertebrates (insects and nematodes) through the inhibition of proteases in these organisms. These proteins are widely distributed in the plant kingdom, and are present in higher concentrations in legume seeds (compared to other organs and other botanical families), motivating studies on their inhibitory effects in various organisms, including humans. In most cases, the biological roles of these proteins have been assigned based mostly on their in vitro action, as is the case with enzyme inhibitors. This review highlights the structural evolution, function, and wide variety of effects of plant Kunitz protease inhibitors, and their potential for pharmaceutical application based on their interactions with different proteases.
Topics: Endopeptidases; Fungi; Humans; Plants; Protease Inhibitors; Serine Proteases
PubMed: 35563133
DOI: 10.3390/ijms23094742 -
Protein Science : a Publication of the... Aug 2023Antiviral therapeutics to treat SARS-CoV-2 are needed to diminish the morbidity of the ongoing COVID-19 pandemic. A well-precedented drug target is the main viral...
Antiviral therapeutics to treat SARS-CoV-2 are needed to diminish the morbidity of the ongoing COVID-19 pandemic. A well-precedented drug target is the main viral protease (M ), which is targeted by an approved drug and by several investigational drugs. Emerging viral resistance has made new inhibitor chemotypes more pressing. Adopting a structure-based approach, we docked 1.2 billion non-covalent lead-like molecules and a new library of 6.5 million electrophiles against the enzyme structure. From these, 29 non-covalent and 11 covalent inhibitors were identified in 37 series, the most potent having an IC of 29 and 20 μM, respectively. Several series were optimized, resulting in low micromolar inhibitors. Subsequent crystallography confirmed the docking predicted binding modes and may template further optimization. While the new chemotypes may aid further optimization of M inhibitors for SARS-CoV-2, the modest success rate also reveals weaknesses in our approach for challenging targets like M versus other targets where it has been more successful, and versus other structure-based techniques against M itself.
Topics: Humans; COVID-19; SARS-CoV-2; Pandemics; Protease Inhibitors; Molecular Docking Simulation; Viral Nonstructural Proteins; Antiviral Agents
PubMed: 37354015
DOI: 10.1002/pro.4712 -
Antiviral Research Dec 2022FB2001 is a drug candidate that targets the main protease of SARS-CoV-2 via covalently binding to cysteine 145. In this study, we evaluated the inhibitory activities of...
FB2001 is a drug candidate that targets the main protease of SARS-CoV-2 via covalently binding to cysteine 145. In this study, we evaluated the inhibitory activities of FB2001 against several SARS-CoV-2 variants in vitro and in vivo (in mice), and we also evaluated the histopathological analysis and immunostaining of FB2001 on lung and brain which have been rarely reported. The results showed that FB2001 exhibited potent antiviral efficacy against several current SARS-CoV-2 variants in Vero E6 cells, namely, B.1.1.7 (Alpha): EC = 0.39 ± 0.01 μM, EC = 0.75 ± 0.01 μM; B.1.351 (Beta): EC = 0.28 ± 0.11 μM, EC = 0.57 ± 0.21 μM; B.1.617.2 (Delta): EC = 0.27 ± 0.05 μM, EC = 0.81 ± 0.20 μM; B.1.1.529 (Omicron): EC = 0.26 ± 0.06 μM and EC = 0.042 ± 0.007 μM (in the presence of a P-glycoprotein inhibitor). FB2001 remained potent against SARS-CoV-2 replication in the presence of high concentrations of human serum, which indicating that human serum had no significant effect on the in vitro inhibitory activity. Additionally, this inhibitor exhibited an additive effect against SARS-CoV-2 when combined with Remdesivir. Furthermore, FB2001 significantly reduced the SARS-CoV-2 copy numbers and titers in the lungs and brains in vivo, and alleviated the pathological symptoms. In addition, FB2001 could alleviated local bleeding, erythrocyte overflow, edema, and inflammatory cell infiltration in brain tissue, and inhibitors reducing viral titers and improving inflammation in the brain have been rarely reported. A physiologically based pharmacokinetic model was established and verified to predict the FB2001 concentration in human lungs. When FB2001 was administered at 200 mg twice a day for 5 days, the observed C in plasma and predicted C of lung total concentration were 0.163 and 2.5 μg/mL, which were approximately 9 and 132-fold higher than the EC of 0.019 μg/mL (0.042 μM) against Omicron variant. Taken together, our study suggests that FB2001 is a promising therapeutic agent in COVID-19 treatment and can be combined with remdesivir to achieve improved clinical outcomes. Owing to its good safety and tolerability in healthy human (NCT05197179 and NCT04766931), FB2001 has been approved for Phase II/III clinical trial (NCT05445934).
Topics: Animals; Humans; Mice; Antiviral Agents; Betacoronavirus; Coronavirus Infections; Pandemics; Pneumonia, Viral; Protease Inhibitors; SARS-CoV-2; Clinical Trials, Phase II as Topic; Clinical Trials, Phase III as Topic; COVID-19 Drug Treatment
PubMed: 36354082
DOI: 10.1016/j.antiviral.2022.105450 -
Paediatric Anaesthesia Oct 2022The protease inhibitor, ritonavir, is a strong inhibitor of CYP 3A. The drug is used for management of the human immunovirus and is currently part of an oral antiviral... (Review)
Review
The protease inhibitor, ritonavir, is a strong inhibitor of CYP 3A. The drug is used for management of the human immunovirus and is currently part of an oral antiviral drug combination (nirmatrelvir-ritonavir) for the early treatment of SARS-2 COVID-19-positive patients aged 12 years and over who have recognized comorbidities. The CYP 3A enzyme system is responsible for clearance of numerous drugs used in anesthesia (e.g., alfentanil, fentanyl, methadone, rocuronium, bupivacaine, midazolam, ketamine). Ritonavir will have an impact on drug clearances that are dependent on ritonavir concentration, anesthesia drug intrinsic hepatic clearance, metabolic pathways, concentration-response relationship, and route of administration. Drugs with a steep concentration-response relationship (ketamine, midazolam, rocuronium) are mostly affected because small changes in concentration have major changes in effect response. An increase in midazolam concentration is observed after oral administration because CYP 3A in the gastrointestinal wall is inhibited, causing a large increase in relative bioavailability. Fentanyl infusion may be associated with a modest increase in plasma concentration and effect, but the large between subject variability of pharmacokinetic and pharmacodynamic concentration changes suggests it will have little impact on an individual patient, especially when used with adverse effect monitoring. It has been proposed that drugs that have no or only a small metabolic pathway involving the CYP 3A enzyme be used during anesthesia, for example, propofol, atracurium, remifentanil, and the volatile agents. That anesthesia approach denies children of drugs with considerable value. It is better that the inhibitory changes in clearance of these drugs are understood so that rational drug choices can be made to tailor drug use to the individual patient. Altered drug dose, anticipation of duration of effect, timing of administration, use of reversal agents and perioperative monitoring would better behoove children undergoing anesthesia.
Topics: Alfentanil; Anesthesia; Antiviral Agents; Child; Cytochrome P-450 CYP3A; Drug Interactions; Enzyme Inhibitors; Humans; Ketamine; Midazolam; Protease Inhibitors; Ritonavir; Rocuronium; COVID-19 Drug Treatment
PubMed: 35842922
DOI: 10.1111/pan.14529 -
Computers in Biology and Medicine Mar 2022Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the causative agent of coronavirus disease 2019 (COVID-19), has adversely affected global health since its...
Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the causative agent of coronavirus disease 2019 (COVID-19), has adversely affected global health since its emergence in 2019. The lack of effective treatments prompted worldwide efforts to immediately develop therapeutic strategies against COVID-19. The main protease (M) of SARS-CoV-2 plays a crucial role in viral replication, and therefore it serves as an attractive target for COVID-19-specific drug development. Due to the richness and diversity of insect protease inhibitors, we docked SARS-CoV-2 M onto 25 publicly accessible insect-derived protease inhibitors using the ClusPro server, and the regions with high inhibitory potentials against M were used to design peptides. Interactions of these inhibitory peptides with M were further assessed by two directed docking programs, AutoDock and Haddock. AutoDock analysis predicted the highest binding energy (-9.39 kcal/mol) and the lowest inhibition constant (130 nM) for the peptide 1KJ0-7 derived from SGCI (Schistocerca gregaria chymotrypsin inhibitor). On the other hand, Haddock analysis resulted in the discovery of a different peptide designated 2ERW-9 from infestin, a serine protease inhibitor of Triatoma infestans, with the best docking score (-131), binding energy (-11.7 kcal/mol), and dissociation constant (2.6E-09 M) for M. Furthermore, using molecular dynamic simulations, 1KJ0-7 and 2ERW-9 were demonstrated to form stable complexes with M. The peptides also showed suitable drug-likeness properties compared to commercially available drugs based on Lipinski's rule. Our findings present two peptides with possible protease inhibitor activities against M and further demonstrate the potential of insect-derived peptides and computer-aided methods for drug discovery.
Topics: Animals; COVID-19; Coronavirus 3C Proteases; Humans; Insecta; Molecular Docking Simulation; Molecular Dynamics Simulation; Protease Inhibitors; SARS-CoV-2
PubMed: 35051855
DOI: 10.1016/j.compbiomed.2022.105228 -
Plant Foods For Human Nutrition... Mar 2022Plant-based diets are a great source of protease inhibitors (PIs). Two of the most well-known families of PIs are Bowman-Birk inhibitors (BBI) and Kunitz-type inhibitors... (Review)
Review
Plant-based diets are a great source of protease inhibitors (PIs). Two of the most well-known families of PIs are Bowman-Birk inhibitors (BBI) and Kunitz-type inhibitors (KTI). The first group acts mainly on trypsin, chymotrypsin, and elastase; the second is on serine, cysteine, and aspartic proteases. PIs can retard or inhibit the catalytic action of enzymes; therefore, they are considered non-nutritional compounds; nevertheless, animal studies and cell line experiments showed promising results of PIs in treating human illnesses such as obesity, cardiovascular diseases, autoimmune diseases, inflammatory processes, and different types of cancer (gastric, colorectal, breast, and lung cancer). Anticarcinogenic activity's proposed mechanisms of action comprise several inhibitory effects at different molecular levels, i.e., transcription, post-transcription, translation, post-translation, and secretion of cancer cells. This work reviews the potential therapeutic applications of PIs as anticarcinogenic and anti-inflammatory agents in human diseases and the mechanisms by which they exert these effects.
Topics: Animals; Aspartic Acid Proteases; Protease Inhibitors; Trypsin; Trypsin Inhibitor, Bowman-Birk Soybean; Trypsin Inhibitors
PubMed: 35000105
DOI: 10.1007/s11130-022-00949-4 -
Journal of Medicinal Chemistry May 2022The coronavirus disease (COVID-19) pandemic has highlighted the ability of scientists to quickly react to the immense challenge presented to the world. The orally...
The coronavirus disease (COVID-19) pandemic has highlighted the ability of scientists to quickly react to the immense challenge presented to the world. The orally available 3CL protease inhibitor S-217622 is currently progressing through clinical trials and its discovery via structure-based drug design, screening and optimization by Shionogi and Hokkaido University is presented here.
Topics: Antiviral Agents; Coronavirus 3C Proteases; Humans; Protease Inhibitors; SARS-CoV-2; COVID-19 Drug Treatment
PubMed: 35507419
DOI: 10.1021/acs.jmedchem.2c00624