While research has identified several inhibitors of the main protease (Mpro) of SARS-CoV-2, a significant portion of these compounds exhibit reduced activity in the presence of reducing agents, raising concerns about their effectiveness in vivo. Furthermore, the conventional biosafety level 3 (BSL-3) for cellular assays using viral particles poses a limitation for the widespread evaluation of Mpro inhibitor efficacy in a cell-based assay. Here, we established a BSL-1 compatible cellular assay to evaluate the in vivo potential of Mpro inhibitors. This assay utilizes mammalian cells expressing a tagged Mpro construct containing N-terminal glutathione S-transferase (GST) and C-terminal hemagglutinin (HA) tags and monitors Mpro autodigestion. Using this method, GC376 and boceprevir effectively inhibited Mpro autodigestion, suggesting their potential in vivo activity. Conversely, carmofur and ebselen did not exhibit significant inhibitory effects in this assay. We further investigated the inhibitory potential of selenoneine on Mpro using this approach. Computational analyses of binding energies suggest that noncovalent interactions play a critical role in facilitating the covalent modification of the C145 residue, leading to Mpro inhibition. Our method is straightforward, cost-effective, and readily applicable in standard laboratories, making it accessible to researchers with varying levels of expertise in infectious diseases.

Boceprevir drug is a ketoamide serine protease inhibitor with a linear peptidomimetic structure that exhibits inhibition activity against 2019-nCoV main protease. This paper reports electronic properties of boceprevir and its molecular docking as well as molecular dynamics simulation analysis with protein receptor. For this, the equilibrium structure of boceprevir has been obtained by DFT at B3LYP and ωB97XD levels with 6-311+G(d,p) basis set in gas and water mediums. HOMO-LUMO and absorption spectrum analysis have been used to evaluate the boceprevir\'s toxicity and photosensitivity, respectively. Molecular docking simulation has been performed to test the binding affinity of boceprevir with 2019-nCoV MPRO; which rendered a variety of desirable binding locations between the ligand and target protein\'s residue positions. The optimum binding location has been considered for molecular dynamics simulation. The findings have been addressed to clarify the boceprevir drug efficacy against the 2019-nCoV MPRO.

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.

Background: Recent international, randomized, placebo-controlled clinical trials (SPRINT-2; RESPOND-2) demonstrated that the triple combination of peginterferon (PEG), ribavirin (RBV) and boceprevir (BOC) was more efficacious than the standard dual therapy of PEG and RBV in treatment of patients chronically infected with genotype 1 hepatitis C virus (HCV) infection. The objective of this study was to evaluate the cost-effectiveness of triple therapy in both treatment-naive and treatment-experienced patients in Hungary. Methods: A Markov model was developed to evaluate the long-term clinical benefits and the costeffectiveness of the triple therapy from the Hungarian payer perspective. Model states were fibrosis (F0-F4, defined using METAVIR fibrosis scores), decompensated cirrhosis (DC), hepatocellular carcinoma (HCC), liver transplantation (LT), and liver-related deaths (LD). Efficacy was estimated from SPRINT-2 and RESPOND-2 studies. Disease progression rates and health state utilities used in the model were obtained from published studies. Estimates of probability of liver transplantation and cost were based on an analysis of the Hungarian Sick Fund database. All cost and benefits were discounted at 5% per year. Results: Compared to dual therapy, triple therapy was projected to increase the life expectancy by 0.98 and 2.42 life years and increase the quality-adjusted life years (QALY) by 0.59 and 1.13 in treatment-naive and treatment-experienced patients, respectively. The corresponding incremental cost-effectiveness ratios were HUF7,747,962 (€26,717) and HUF5,888,240 (€20,304) per QALY. The lifetime incidence of severe liver disease events (DC, HCC, LT, LD) were projected to decrease by 45% and 61% in treatment-naïve and treatment-experienced patients treated with triple therapy groups in comparison with PEG-RBV treatment. Conclusion: The addition of boceprevir to standard therapy for the treatment of patients with genotype 1 chronic HCV infection in Hungary is projected to be cost-effective using a commonly used willingness to pay threshold of HUF 8.46 million (3 times gross domestic product per capita).

In recent years, more than 200 countries of the world have faced a health crisis due to the epidemiological disease of COVID-19 caused by the SARS-CoV-2 virus. It had a huge impact on the world economy and the global health sector. Researchers are studying the design and discovery of drugs that can inhibit SARS-CoV-2. The main protease of SARS-CoV-2 is an attractive target for the study of antiviral drugs against coronavirus diseases. According to the docking results, binding energy for boceprevir, masitinib and rupintrivir with CMP are -10.80, -9.39, and -9.51 kcal/mol respectively. Also, for all investigated systems, van der Waals and electrostatic interactions are quite favorable for binding the drugs to SARS-CoV-2 coronavirus main protease, indicating confirmation of the complex stability.

BACKGROUND: The unavailability of target-specific antiviral drugs for SARS-CoV-2 viral infection kindled the motivation to virtually design derivatives of 6,6-dimethyl-3-azabicyclo[3.1.0]hexane-2-carboxamide as potential antiviral inhibitors against the concerned virus. The molecular docking and molecular dynamic results revealed that the reported derivatives have a potential to act as antiviral drug against SARS-CoV-2. The reported hit compounds can be considered for in vitro and in vivo analyses.
METHODS: Fragment-based drug designing was used to model the derivatives. Furthermore, DFT simulations were carried out using B3LYP/6-311G** basis set. Docking simulations were performed by using a combination of empirical free energy force field with a Lamarckian genetic algorithm under AutoDock 4.2. By the application of AMBER14 force field and SPCE water model, molecular dynamic simulations and MM-PBSA were calculated for 100 ns.

The main protease (Mpro) of the betacoronavirus SARS-CoV-2 is an attractive target for the development of treatments for COVID-19. Structure-based design is a successful approach to discovering new inhibitors of the Mpro. Starting from crystal structures of the Mpro in complexes with the Hepatitis C virus NS3/4A protease inhibitors boceprevir and telaprevir, we optimized the potency of the alpha-ketoamide boceprevir against the Mpro by replacing its P1 cyclobutyl moiety by a γ-lactam as a glutamine surrogate. The resulting compound, MG-78, exhibited an IC50 of 13 nM versus the recombinant Mpro, and similar potency was observed for its P1\' N-methyl derivative MG-131. Crystal structures confirmed the validity of our design concept. In addition to SARS-CoV-2 Mpro inhibition, we also explored the activity of MG-78 against the Mpro of the alphacoronavirus HCoV NL63 and against enterovirus 3C proteases. The activities were good (0.33 µM, HCoV-NL63 Mpro), moderate (1.45 µM, Coxsackievirus 3Cpro), and relatively poor (6.7 µM, enterovirus A71 3Cpro), respectively. The structural basis for the differences in activities was revealed by X-ray crystallo-graphy. We conclude that the modified boceprevir scaffold is suitable for obtaining high-potency inhibitors of the coronavirus Mpros but further optimization would be needed to target enterovirus 3Cpros efficiently.

The novel coronavirus disease (COVID-19) has spread throughout the globe, affecting millions of people. The World Health Organization (WHO) has declared this infectious disease a pandemic. At present, several clinical trials are going on to identify possible drugs for treating this infection. SARS-CoV-2 Mpro is one of the most critical drug targets for the blockage of viral replication. The aim of this study was to identify potential natural anthraquinones that could bind to the active site of SARS-CoV-2 main protease and stop the viral replication. Blind molecular docking studies of 13 anthraquinones and one control drug (Boceprevir) with SARS-CoV-2 Mpro were carried out using the SwissDOCK server, and alterporriol-Q that showed the highest binding affinity towards Mpro were subjected to molecular dynamics simulation studies. This study indicated that several antiviral anthraquinones could prove to be effective inhibitors for SARS-CoV-2 Mpro of COVID-19 as they bind near the active site having the catalytic dyad, HIS41 and CYS145 through non-covalent forces. The anthraquinones showed less inhibitory potential as compared to the FDA-approved drug, boceprevir. Among the anthraquinones studied, alterporriol-Q was found to be the most potent inhibitor of SARS-CoV-2 Mpro. Further, MD simulation studies for Mpro- alterporriol-Q system suggested that alterporriol-Q does not alter the structure of Mpro to a significant extent. Considering the impact of COVID-19, identification of alternate compounds like alterporriol-Q that could inhibit the viral infection will help in accelerating the process of drug discovery.
UNASSIGNED: The online version contains supplementary material available at 10.1007/s11756-021-01004-4.

Because of the scale of the novel coronavirus (COVID-19) pandemic and the swift transmission of this highly contagious respiratory virus, repurposing existing drugs has become an urgent treatment approach. The objective of our study is to unravel the binding mechanism of the Food and Drug Administration (FDA)-approved dexamethasone (Dex) and boceprevir (Boc) drugs with selected COVID-19 protein targets SARS-CoV-2 spike protein C-terminal domain (spike-CTD), main protease (Mpro), and interleukin-6 (IL-6). Another objective is to analyze the effects of binding Dex and Boc drugs on the interactions of viral spike protein to human angiotensin-converting enzyme 2 (hACE2). Molecular docking and one-microsecond-long molecular dynamics simulations of each of the six protein-drug complexes along with steered molecular dynamics (SMD) and umbrella sampling (US) methods have revealed the binding mode interactions and the physicochemical stability of the three targeted proteins with two drugs. Results have shown that both drugs bind strongly with the three protein targets through hydrogen bonding and hydrophobic interactions. A major finding from this study is how the binding of the drugs with viral spike protein affects its interactions at the binding interface with hACE2 protein. Simulations of drug-bound spike-CTD with hACE2 show that due to the presence of a drug at the binding interface of spike-CTD, hACE2 is being blocked from making putative interactions with viral protein at such interface. These important findings regarding the binding affinity and stability of the two FDA-approved drugs with the main targets of COVID-19 along with the effect of drugs on hACE2 interactions would contribute to COVID-19 drug discovery and development.
UNASSIGNED: The online version contains supplementary material available at 10.1007/s11696-021-01843-0.

Remdesivir (GS-5734), a drug initially developed to treat hepatitis C and Ebola virus disease, was the first approved treatment for severe coronavirus disease 2019 (COVID-19). However, apart from remdesivir, there is a paucity of other specific anti-viral agents against SARS-CoV-2 infection. In 2017, researchers had documented the anti-coronavirus potential of remdesivir in animal models. At the same time, trials performed during two Ebola outbreaks in Africa showed that the drug was safe. Although vaccines against SARS-CoV-2 infection have emerged at an enormously high speed, equivalent results from efforts towards the development of anti-viral drugs, which could have played a truly life-saving role in the current stage of the pandemic, have been stagnating. In this review, we will focus on the current treatment options for COVID-19 which mainly consist of repurposed agents or treatments conferring passive immunity (convalescent plasma or monoclonal antibodies). Additionally, potential specific anti-viral therapies under development will be reviewed, such as the decoy miniprotein CTC-445.2d, protease inhibitors, mainly against the Main protein Mpro, nucleoside analogs, such as molnupiravir and compounds blocking the replication transcription complex proteins, such as zotatifin and plitidepsin. These anti-viral agents seem to be very promising but still require meticulous clinical trial testing in order to establish their efficacy and safety. The continuous emergence of viral variants may pose a real challenge to the scientific community towards that end. In this context, the advent of nanobodies together with the potential administration of a combination of anti-viral drugs could serve as useful tools in the armamentarium against COVID-19.