Antibiotics daptomycin interacts with S protein of SARS-CoV-2 to promote cell invasion of Omicron (B1.1.529) pseudovirus.

The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) pandemic has posed enormous challenges to global public health. The use of antibiotics has greatly increased during the SARS-CoV-2 epidemic owing to the presence of bacterial co-infection and secondary bacterial infections. The antibiotics daptomycin (DAP) is widely used in the treatment of infectious diseases caused by gram-positive bacteria owing to its highly efficient antibacterial activity. It is pivotal to study the antibiotics usage options for patients of coronavirus infectious disease (COVID-19) with pneumonia those need admission to receive antibiotics treatment for bacterial co-infection in managing COVID-19 disease. Herein, we have revealed the interactions of DAP with the S protein of SARS-CoV-2 and the variant Omicron (B1.1.529) using the molecular docking approach and Omicron (B1.1.529) pseudovirus (PsV) mimic invasion. Molecular docking analysis shows that DAP has a certain degree of binding ability to the S protein of SARS-CoV-2 and several derived virus variants, and co-incubation of 1-100 µM DAP with cells promotes the entry of the PsV into human angiotensin-converting enzyme 2 (hACE2)-expressing HEK-293T cells (HEK-293T-hACE2), and this effect is related to the concentration of extracellular calcium ions (Ca2+). The PsV invasion rate in the HEK-293T-hACE2 cells concurrently with DAP incubation was 1.7 times of PsV infection alone. In general, our findings demonstrate that DAP promotes the infection of PsV into cells, which provides certain reference of antibiotics selection and usage optimization for clinicians to treat bacterial coinfection or secondary infection during SARS-CoV-2 infection.

Emerging threat: Antimicrobial resistance proliferation during epidemics - A case study of the SARS-CoV-2 pandemic in South Brazil.

The escalating global concern of antimicrobial resistance poses a significant challenge to public health. This study delved into the occurrence of resistant bacteria and antimicrobial resistance genes in the waters and sediments of urban rivers and correlated this emergence and the heightened use of antimicrobials during the COVID-19 pandemic. Isolating 45 antimicrobial-resistant bacteria across 11 different species, the study identifies prevalent resistance patterns, with ceftriaxone resistance observed in 18 isolates and ciprofloxacin resistance observed in 13 isolates. The detection of extended-spectrum ß-lactamases, carbapenemases, and acquired quinolone resistance genes in all samples underscores the gravity of the situation. Comparison with a pre-pandemic study conducted in the same rivers in 2019 reveals the emergence of previously undetected new resistant species, and the noteworthy presence of new resistant species and alterations in resistance profiles among existing species. Notably, antimicrobial concentrations in rivers increased during the pandemic, contributing significantly to the scenario of antimicrobial resistance observed in these rivers. We underscore the substantial impact of heightened antimicrobial usage during epidemics, such as COVID-19, on resistance in urban rivers. It provides valuable insights into the complex dynamics of antimicrobial resistance in environmental settings and calls for comprehensive approaches to combat this pressing global health issue, safeguarding both public and environmental health.

Discovery of 2-Amide-3-methylester Thiophenes that Target SARS-CoV-2 Mac1 and Repress Coronavirus Replication, Validating Mac1 as an Antiviral Target.

The COVID-19 pandemic caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) virus has made it clear that further development of antiviral therapies will be needed. Here, we describe small-molecule inhibitors for SARS-CoV-2 Mac1, which counters ADP-ribosylation-mediated innate immune responses. Three high-throughput screening hits had the same 2-amide-3-methylester thiophene scaffold. We studied the compound binding mode using X-ray crystallography, allowing us to design analogues. Compound 27 (MDOLL-0229) had an IC50 of 2.1 µM and was selective for CoV Mac1 proteins after profiling for activity against a panel of viral and human proteins. The improved potency allowed testing of its effect on virus replication, and indeed, 27 inhibited replication of both murine hepatitis virus (MHV) prototypes CoV and SARS-CoV-2. Sequencing of a drug-resistant MHV identified mutations in Mac1, further demonstrating the specificity of 27. Compound 27 is the first Mac1-targeted small molecule demonstrated to inhibit coronavirus replication in a cell model.

Efficient in vitro and in vivo transfection of self-amplifying mRNA with linear poly(propylenimine) and poly(ethylenimine-propylenimine) random copolymers as non-viral carriers.

Messenger RNA (mRNA) based vaccines have been introduced worldwide to combat the Covid-19 pandemic. These vaccines consist of non-amplifying mRNA formulated in lipid nanoparticles (LNPs). Consequently, LNPs are considered benchmark non-viral carriers for nucleic acid delivery. However, the formulation and manufacturing of these mRNA-LNP nanoparticles are expensive and time-consuming. Therefore, we used self-amplifying mRNA (saRNA) and synthesized novel polymers as alternative non-viral carrier platform to LNPs, which enable a simple, rapid, one-pot formulation of saRNA-polyplexes. Our novel polymer-based carrier platform consists of randomly concatenated ethylenimine and propylenimine comonomers, resulting in linear, poly(ethylenimine-ran-propylenimine) (L-PEIx-ran-PPIy) copolymers with controllable degrees of polymerization. Here we demonstrate in multiple cell lines, that our saRNA-polyplexes show comparable to higher in vitro saRNA transfection efficiencies and higher cell viabilities compared to formulations with Lipofectamine MessengerMAX™ (LFMM), a commercial, lipid-based carrier considered to be the in vitro gold standard carrier. This is especially true for our in vitro best performing saRNA-polyplexes with N/P 5, which are characterised with a size below 100 nm, a positive zeta potential, a near 100% encapsulation efficiency, a high retention capacity and the ability to protect the saRNA from degradation mediated by RNase A. Furthermore, an ex vivo hemolysis assay with pig red blood cells demonstrated that the saRNA-polyplexes exhibit negligible hemolytic activity. Finally, a bioluminescence-based in vivo study was performed over a 35-day period, and showed that the polymers result in a higher and prolonged bioluminescent signal compared to naked saRNA and L-PEI based polyplexes. Moreover, the polymers show different expression profiles compared to those of LNPs, with one of our new polymers (L-PPI250) demonstrating a higher sustained expression for at least 35 days after injection.

Comparison of effectiveness and safety of molnupiravir versus sotrovimab for COVID-19: A systematic review and meta-analysis.

BACKGROUND AND AIM: This systematic review and meta-analysis aimed to compare the effectiveness and safety of molnupiravir and sotrovimab in the treatment of patients with coronavirus disease 2019 (COVID-19). METHODS: Cochrane Library, Web of Science, PubMed, medRxiv, and Google Scholar were systematically searched to identify relevant evidence up to December 2023. The risk of bias was assessed using the risk of bias in nonrandomized studies of interventions tool. Data were analyzed using Comprehensive Meta-Analysis (CMA). RESULTS: Our search identified and included 13 studies involving 16166 patients. The meta-analysis revealed a significant difference between the molnupiravir and sotrovimab groups in terms of the mortality rate (odds ratio [OR] = 2.07, 95% confidence interval [CI]: 1.16, 3.70). However, no significant difference was observed between the two groups in terms of hospitalization rate (OR = 0.71, 95% CI: 0.47, 1.06), death or hospitalization rate (OR = 1.51, 95% CI: 0.81, 2.83), and intensive care unit admission (OR = 0.59, 95% CI: 0.07, 4.84). In terms of safety, molnupiravir was associated with a higher incidence of adverse events (OR = 1.67, 95% CI: 1.21, 2.30). CONCLUSION: The current findings indicate that sotrovimab may be more effective than molnupiravir in reducing the mortality rate in COVID-19 patients. However, no statistical difference was observed between the two treatments for other effectiveness outcomes. The certainty of evidence for these findings was rated as low or moderate. Further research is required to provide a better comparison of these interventions in treating COVID-19 patients.

Discovery of First-in-Class PROTAC Degraders of SARS-CoV-2 Main Protease.

We have witnessed three coronavirus (CoV) outbreaks in the past two decades, including the COVID-19 pandemic caused by SARS-CoV-2. Main protease (MPro), a highly conserved protease among various CoVs, is essential for viral replication and pathogenesis, making it a prime target for antiviral drug development. Here, we leverage proteolysis targeting chimera (PROTAC) technology to develop a new class of small-molecule antivirals that induce the degradation of SARS-CoV-2 MPro. Among them, MPD2 was demonstrated to effectively reduce MPro protein levels in 293T cells, relying on a time-dependent, CRBN-mediated, and proteasome-driven mechanism. Furthermore, MPD2 exhibited remarkable efficacy in diminishing MPro protein levels in SARS-CoV-2-infected A549-ACE2 cells. MPD2 also displayed potent antiviral activity against various SARS-CoV-2 strains and exhibited enhanced potency against nirmatrelvir-resistant viruses. Overall, this proof-of-concept study highlights the potential of targeted protein degradation of MPro as an innovative approach for developing antivirals that could fight against drug-resistant viral variants.

Design of a SARS-CoV-2 papain-like protease inhibitor with antiviral efficacy in a mouse model.

The emergence of SARS-CoV-2 variants and drug-resistant mutants calls for additional oral antivirals. The SARS-CoV-2 papain-like protease (PLpro) is a promising but challenging drug target. We designed and synthesized 85 noncovalent PLpro inhibitors that bind to a recently discovered ubiquitin binding site and the known BL2 groove pocket near the S4 subsite. Leads inhibited PLpro with the inhibitory constant Ki values from 13.2 to 88.2 nanomolar. The co-crystal structures of PLpro with eight leads revealed their interaction modes. The in vivo lead Jun12682 inhibited SARS-CoV-2 and its variants, including nirmatrelvir-resistant strains with EC50 from 0.44 to 2.02 micromolar. Oral treatment with Jun12682 improved survival and reduced lung viral loads and lesions in a SARS-CoV-2 infection mouse model, suggesting that PLpro inhibitors are promising oral SARS-CoV-2 antiviral candidates.

Discovery of Covalent Lead Compounds Targeting 3CL Protease with a Lateral Interactions Spiking Neural Network.

Covalent drugs exhibit advantages in that noncovalent drugs cannot match, and covalent docking is an important method for screening covalent lead compounds. However, it is difficult for covalent docking to screen covalent compounds on a large scale because covalent docking requires determination of the covalent reaction type of the compound. Here, we propose to use deep learning of a lateral interactions spiking neural network to construct a covalent lead compound screening model to quickly screen covalent lead compounds. We used the 3CL protease (3CL Pro) of SARS-CoV-2 as the screen target and constructed two classification models based on LISNN to predict the covalent binding and inhibitory activity of compounds. The two classification models were trained on the covalent complex data set targeting cysteine (Cys) and the compound inhibitory activity data set targeting 3CL Pro, respected, with good prediction accuracy (ACC > 0.9). We then screened the screening compound library with 6 covalent binding screening models and 12 inhibitory activity screening models. We tested the inhibitory activity of the 32 compounds, and the best compound inhibited SARS-CoV-2 3CL Pro with an IC50 value of 369.5 nM. Further assay implied that dithiothreitol can affect the inhibitory activity of the compound to 3CL Pro, indicating that the compound may covalently bind 3CL Pro. The selectivity test showed that the compound had good target selectivity to 3CL Pro over cathepsin L. These correlation assays can prove the rationality of the covalent lead compound screening model. Finally, covalent docking was performed to demonstrate the binding conformation of the compound with 3CL Pro. The source code can be obtained from the GitHub repository (https://github.com/guzh970630/Screen_Covalent_Compound_by_LISNN).

NaClO Co-selects antibiotic and disinfectant resistance in Klebsiella pneumonia: Implications for the potential risk of extensive disinfectant use during COVID-19 pandemic.

The inappropriate use of antibiotics is widely recognized as the primary driver of bacterial antibiotic resistance. However, less attention has been given to the potential induction of multidrug-resistant bacteria through exposure to disinfectants. In this study, Klebsiella pneumonia, an opportunistic pathogen commonly associated with hospital and community-acquired infection, was experimentally exposed to NaClO at both minimum inhibitory concentration (MIC) and sub-MIC levels over a period of 60 days. The result demonstrated that NaClO exposure led to enhanced resistance of K. pneumonia to both NaClO itself and five antibiotics (erythromycin, polymyxin B, gentamicin, tetracycline, and ciprofloxacin). Concurrently, the evolved resistant strains exhibited fitness costs, as evidenced by decreased growth rates. Whole population sequencing revealed that both concentrations of NaClO exposure caused genetic mutations in the genome of K. pneumonia. Some of these mutations were known to be associated with antibiotic resistance, while others had not previously been identified as such. In addition, 11 identified mutations were located in the virulence factors, demonstrating that NaClO exposure may also impact the pathogenicity of K. pneumoniae. Overall, this study highlights the potential for the widespread use of NaClO-containing disinfectants during the COVID-19 pandemic to contribute to the emergence of antibiotic-resistant bacteria. ENVIRONMENTAL IMPLICATION: Considering the potential hazardous effects of disinfectant residues on environment, organisms and biodiversity, the sharp rise in use of disinfectants during COVID-19 pandemic has been considered highly likely to cause worldwide secondary disasters in ecosystems and human health. This study demonstrated that NaClO exposure enhanced the resistance of K. pneumonia to both NaClO and five antibiotics (erythromycin, polymyxin B, gentamicin, tetracycline, and ciprofloxacin), highlighting the widespread use of NaClO-containing disinfectants during the COVID-19 pandemic may increase the emergence of antibiotic-resistant bacteria in the environment.

Primary Antioxidant Power and Mpro SARS-CoV-2 Non-Covalent Inhibition Capabilities of Miquelianin.

The antioxidant power of quercetin-3-O-glucuronide (miquelianin) has been studied, at the density functional level of theory, in both lipid-like and aqueous environments. In the aqueous phase, the computed pKa equilibria allowed the identification of the neutral and charged species present in solution that can react with the ⋅OOH radical. The Hydrogen Atom Transfer (HAT), Single Electron Transfer (SET) and Radical Adduct Formation (RAF) mechanisms were considered, and the individual, total and fraction corrected rate constants were obtained. Potential non-covalent inhibition of Mpro from SARS-CoV-2 by miquelianin has been also evaluated.

Reavaliação do nirmatrelvir/ritonavir para o tratamento de pacientes com COVID-19 não hospitalizados e com alto risco de doença grave / Revalidation of nirmatrelvir/ritonavir for the treatment of patients with COVID-19 who are not hospitalized and with a high risk of serious illness

INTRODUÇÃO: Pacientes com fatores de risco como idade avançada, imunodepressão, obesidade e doenças cardiovasculares têm risco aumentado de internação, intubação e morte. De acordo com dados brasileiros, o risco de morte por covid-19 aumenta com o número de fatores de risco que o paciente apresenta, sendo igual a 17% em pacientes com 2 fatores de risco e 76% na presença de 8 fatores de risco. Além disso, mesmo aqueles pacientes que sobrevivem a uma internação em terapia intensiva frequentemente enfrentam sequelas e representam alto custo para o sistema público. O medicamento nirmatrelvir associado ao ritonavir têm o objetivo de prevenir internações, complicações e morte. Ele é indicado para pacientes com Covid-19 leve a moderada, não hospitalizados, até 5 dias do início dos sintomas. Apesar dos avanços da vacinação no Brasil, evidências sobre a falha vacinal em idosos e imunodeprimidos destacam a importância da disponibilidade de alternativas terapêuticas para essas populações. O presente relatório teve por objetivo avaliar evidências sobre a efetividade do tratamento em pacientes vacinados com alto risco de agravamento da doença. PERGUNTA: O medicamento nirmatrelvir/ritonavir é eficaz, seguro e custo-efetivo para pacientes com covid19 leve a moderada não hospitalizados vacinados que apresentam alto risco de agravamento da doença? EVIDÊNCIAS CLÍNICAS: Resultados obtidos a partir de estudos observacionais de mundo real confirmaram os resultados do ensaio clínico do medicamento nirmatrelvir/ritonavir, demonstrando que o tratamento de pacientes de grupos de risco é capaz de reduzir o risco de desfechos desfavoráveis como internação e óbito entre cerca de 50% e 70%, inclusive entre pacientes previamente vacinados. ANÁLISE DE IMPACTO ORÇAMENTÁRIO: A análise de impacto orçamentário do relatório de recomendação do nirmatrelvir/ritonavir foi atualizada considerando-se o cenário atual da pandemia no Brasil. De acordo com a nova análise, o uso do nirmatrelvir/ritonavir por pacientes com idade ≥ 65 anos e imunossuprimidos com idade ≥ 18 anos, resultaria em uma economia de recursos de R$ 408.957.111,38 em 5 anos. Ressalta-se, no entanto, que devido à dinâmica de difícil previsão da pandemia, este montante está sujeito à incerteza. Considerando-se a análise realizada anteriormente no relatório de recomendação, pode-se concluir que o montante economizado se reduz proporcionalmente à redução do número de casos da doença na população alvo. CONSIDERAÇÕES FINAIS: De acordo com as evidências atualmente disponíveis, o uso do nirmatrelvir/ritonavir é efetivo e seguro para pacientes com covid-19 leve a moderada não hospitalizados vacinados que apresentam alto risco de agravamento da doença. O impacto orçamentário está sujeito a incertezas já que o número de casos da doença no horizonte temporal da análise é de difícil previsão. RECOMENDAÇÃO PRELIMINAR DA CONITEC: Diante do exposto, os membros do Comitê de Medicamentos da Conitec, em sua 16ª Reunião Extraordinária, realizada no dia 1º de novembro de 2023, deliberaram que a matéria fosse disponibilizada em Consulta Pública com recomendação preliminar favorável à incorporação no SUS do nirmatrelvir/ritonavir para o tratamento de pacientes com Covid-19 não hospitalizados com idade a partir de 65 anos ou pacientes imunossuprimidos a partir de 18 anos de idade. Os membros do Comitê concordaram na manutenção da indicação de uso, não havendo ampliação do público-alvo, justificada pela restrição orçamentária, considerando que há incremento de custo da aquisição da tecnologia, ainda que haja economia de recursos ao serem evitadas internações e óbitos. CONSULTA PÚBLICA: Das nove contribuições recebidas, cinco contribuições foram de cunho técnico-científico e quatro contribuições de experiência ou opinião. Todas as contribuições concordaram com a recomendação preliminar da Conitec de incorporar o nirmatrelvir/ritonavir. Duas contribuições técnico-científicassugeriram ampliação da população elegível ao tratamento com o medicamento com a inclusão de indicação para pacientes com taxa de filtração glomerular menor que 30 ml/min/1,73m2 e de pacientes adultos com asma grave independentemente da faixa etária. Uma contribuição técnico-científica enviada pela empresa fabricante do medicamento expressou sua concordância com e solicitou a inclusão de informaçõea adicionais no relatório. As contribuições de experiência ou opinião ressaltaram a eficácia e segurança do medicamento para a população alvo. RECOMEDAÇÃO FINAL DA CONITEC: Diante do exposto, os membros do Comitê de Medicamentos, presentes na 126ª Reunião Ordinária da Conitec, realizada no dia 01 de fevereiro de 2024, deliberaram, por unanimidade, após reavaliação, manter a incorporação do nirmatrelvir/ritonavir, no SUS, para o tratamento da Covid-19 nos seguintes grupos de pacientes com sintomas leves a moderados, que não requerem oxigênio suplementar, independentemente do status vacinal: a) imunocomprometidos com idade ≥ 18 anos; b) com idade ≥ 65 anos. Foi assinado o registro de deliberação nº 874/2024. DECISÃO: manter a incorporação, no âmbito do Sistema Único de Saúde - SUS, de nirmatrelvir/ritonavir para o tratamento da Covid-19 para pacientes com sintomas leves a moderados, que não requerem oxigênio suplementar, independentemente do status vacinal e com idade igual ou superior a 65 anos ou imunocomprometidos com idade igual ou superior a 18 anos, publicada no Diário Oficial da União nº 46, seção 1, página 54, em 07 de março de 2024.

Oral Simnotrelvir for Adult Patients with Mild-to-Moderate Covid-19.

BACKGROUND: Simnotrelvir is an oral 3-chymotrypsin-like protease inhibitor that has been found to have in vitro activity against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and potential efficacy in a phase 1B trial. METHODS: In this phase 2-3, double-blind, randomized, placebo-controlled trial, we assigned patients who had mild-to-moderate coronavirus disease 2019 (Covid-19) and onset of symptoms within the past 3 days in a 1:1 ratio to receive 750 mg of simnotrelvir plus 100 mg of ritonavir or placebo twice daily for 5 days. The primary efficacy end point was the time to sustained resolution of symptoms, defined as the absence of 11 Covid-19-related symptoms for 2 consecutive days. Safety and changes in viral load were also assessed. RESULTS: A total of 1208 patients were enrolled at 35 sites in China; 603 were assigned to receive simnotrelvir and 605 to receive placebo. Among patients in the modified intention-to-treat population who received the first dose of trial drug or placebo within 72 hours after symptom onset, the time to sustained resolution of Covid-19 symptoms was significantly shorter in the simnotrelvir group than in the placebo group (180.1 hours [95% confidence interval {CI}, 162.1 to 201.6] vs. 216.0 hours [95% CI, 203.4 to 228.1]; median difference, -35.8 hours [95% CI, -60.1 to -12.4]; P = 0.006 by Peto-Prentice test). On day 5, the decrease in viral load from baseline was greater in the simnotrelvir group than in the placebo group (mean difference [±SE], -1.51±0.14 log10 copies per milliliter; 95% CI, -1.79 to -1.24). The incidence of adverse events during treatment was higher in the simnotrelvir group than in the placebo group (29.0% vs. 21.6%). Most adverse events were mild or moderate. CONCLUSIONS: Early administration of simnotrelvir plus ritonavir shortened the time to the resolution of symptoms among adult patients with Covid-19, without evident safety concerns. (Funded by Jiangsu Simcere Pharmaceutical; ClinicalTrials.gov number, NCT05506176.).

iDVEIP: A computer-aided approach for the prediction of viral entry inhibitory peptides.

With the notable surge in therapeutic peptide development, various peptides have emerged as potential agents against virus-induced diseases. Viral entry inhibitory peptides (VEIPs), a subset of antiviral peptides (AVPs), offer a promising avenue as entry inhibitors (EIs) with distinct advantages over chemical counterparts. Despite this, a comprehensive analytical platform for characterizing these peptides and their effectiveness in blocking viral entry remains lacking. In this study, we introduce a groundbreaking in silico approach that leverages bioinformatics analysis and machine learning to characterize and identify novel VEIPs. Cross-validation results demonstrate the efficacy of a model combining sequence-based features in predicting VEIPs with high accuracy, validated through independent testing. Additionally, an EI type model has been developed to distinguish peptides specifically acting as Eis from AVPs with alternative activities. Notably, we present iDVEIP, a web-based tool accessible at http://mer.hc.mmh.org.tw/iDVEIP/, designed for automatic analysis and prediction of VEIPs. Emphasizing its capabilities, the tool facilitates comprehensive analyses of peptide characteristics, providing detailed amino acid composition data for each prediction. Furthermore, we showcase the tool's utility in identifying EIs against severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2).

In-silico evaluation of natural alkaloids against the main protease and spike glycoprotein as potential therapeutic agents for SARS-CoV-2.

Severe Acute Respiratory Syndrome Corona Virus (SARS-CoV-2) is the causative agent of COVID-19 pandemic, which has resulted in global fatalities since late December 2019. Alkaloids play a significant role in drug design for various antiviral diseases, which makes them viable candidates for treating COVID-19. To identify potential antiviral agents, 102 known alkaloids were subjected to docking studies against the two key targets of SARS-CoV-2, namely the spike glycoprotein and main protease. The spike glycoprotein is vital for mediating viral entry into host cells, and main protease plays a crucial role in viral replication; therefore, they serve as compelling targets for therapeutic intervention in combating the disease. From the selection of alkaloids, the top 6 dual inhibitory compounds, namely liensinine, neferine, isoliensinine, fangchinoline, emetine, and acrimarine F, emerged as lead compounds with favorable docked scores. Interestingly, most of them shared the bisbenzylisoquinoline alkaloid framework and belong to Nelumbo nucifera, commonly known as the lotus plant. Docking analysis was conducted by considering the key active site residues of the selected proteins. The stability of the top three ligands with the receptor proteins was further validated through dynamic simulation analysis. The leads underwent ADMET profiling, bioactivity score analysis, and evaluation of drug-likeness and physicochemical properties. Neferine demonstrated a particularly strong affinity for binding, with a docking score of -7.5025 kcal/mol for main protease and -10.0245 kcal/mol for spike glycoprotein, and therefore a strong interaction with both target proteins. Of the lead alkaloids, emetine and fangchinoline demonstrated the lowest toxicity and high LD50 values. These top alkaloids, may support the body's defense and reduce the symptoms by their numerous biological potentials, even though some properties naturally point to their direct antiviral nature. These findings demonstrate the promising anti-COVID-19 properties of the six selected alkaloids, making them potential candidates for drug design. This study will be beneficial in effective drug discovery and design against COVID-19 with negligible side effects.

AC-73 and Syrosingopine Inhibit SARS-CoV-2 Entry into Megakaryocytes by Targeting CD147 and MCT4.

Coagulation disorders are described in COVID-19 and long COVID patients. In particular, SARS-CoV-2 infection in megakaryocytes, which are precursors of platelets involved in thrombotic events in COVID-19, long COVID and, in rare cases, in vaccinated individuals, requires further investigation, particularly with the emergence of new SARS-CoV-2 variants. CD147, involved in the regulation of inflammation and required to fight virus infection, can facilitate SARS-CoV-2 entry into megakaryocytes. MCT4, a co-binding protein of CD147 and a key player in the glycolytic metabolism, could also play a role in SARS-CoV-2 infection. Here, we investigated the susceptibility of megakaryocytes to SARS-CoV-2 infection via CD147 and MCT4. We performed infection of Dami cells and human CD34+ hematopoietic progenitor cells induced to megakaryocytic differentiation with SARS-CoV-2 pseudovirus in the presence of AC-73 and syrosingopine, respective inhibitors of CD147 and MCT4 and inducers of autophagy, a process essential in megakaryocyte differentiation. Both AC-73 and syrosingopine enhance autophagy during differentiation but only AC-73 enhances megakaryocytic maturation. Importantly, we found that AC-73 or syrosingopine significantly inhibits SARS-CoV-2 infection of megakaryocytes. Altogether, our data indicate AC-73 and syrosingopine as inhibitors of SARS-CoV-2 infection via CD147 and MCT4 that can be used to prevent SARS-CoV-2 binding and entry into megakaryocytes, which are precursors of platelets involved in COVID-19-associated coagulopathy.

Discovery of 3-oxo-1,2,3,4-tetrahydropyrido[1,2-a]pyrazin derivatives as SARS-CoV-2 main protease inhibitors through virtual screening and biological evaluation.

The COVID-19 caused by SARS-CoV-2 has led to a global pandemic that continues to impact societies and economies worldwide. The main protease (Mpro) plays a crucial role in SARS-CoV-2 replication and is an attractive target for anti-SARS-CoV-2 drug discovery. Herein, we report a series of 3-oxo-1,2,3,4-tetrahydropyrido[1,2-a]pyrazin derivatives as non-peptidomimetic inhibitors targeting SARS-CoV-2 Mpro through structure-based virtual screening and biological evaluation. Further similarity search and structure-activity relationship study led to the identification of compound M56-S2 with the enzymatic IC50 value of 4.0 µM. Moreover, the molecular simulation and predicted ADMET properties, indicated that non-peptidomimetic inhibitor M56-S2 might serve as a useful starting point for the further discovery of highly potent inhibitors targeting SARS-CoV-2 Mpro.

Development, Histological, and Ultrastructural Evaluation of Sheep Hyperimmune Serum Therapy for COVID-19 / Desarrollo, Evaluación Histológica y Ultraestructural de la Terapia con Suero Hiperinmune de Oveja para COVID-19

SUMMARY: In response to the threat posed by new variants of SARS-CoV-2 and the urgent need for effective treatments in the absence of vaccines, the aim of this study was to develop a rapid and cost-effective hyperimmune serum (HS) derived from sheep and assess its efficacy. The utilization of a halal-certified, easily maintained in certain geographic regions, easy-to-handle animal such as sheep could provide a viable alternative to the expensive option of horses. Sheep were immunized with a whole inactivated SARS-CoV- 2 antigen to produce HS, which was evaluated for neutralizing potency using the PRNT50 assay. K18-hACE2 transgenic mice (n=35) were divided into three groups: control, SARS-CoV-2 exposure through inhalation, and SARS-CoV-2 exposed mice treated with HS. HS efficacy was assessed through serum proinflammatory cytokine levels, qRT-PCR analysis, histopathological examination of lungs and hearts, and transmission electron microscopy. Purified HS exhibited significant neutralizing activity (1/24,576). The SARS-CoV-2+HS group showed lower levels of TNF-α, IL-10, and IL-6 (P<0.01) and relatively lower levels of MCP-1 compared to the SARS-CoV-2 group. HS prevented death, reduced viral RNA levels in the lungs and hearts, protected against severe interstitial pneumonia, preserved lung tissue integrity, and prevented myocyte damage, while the SARS-CoV-2 group exhibited viral presence in the lungs. This study successfully developed a sheep-derived HS against the entire SARS-CoV-2 virus, resulting in a significant reduction in infection severity, inflammation, and systemic cytokine production. The findings hold promise for treating severe COVID-19 cases, including emerging viral variants, and immunocompromised patients.

En respuesta a la amenaza que suponen las nuevas variantes del SARS-CoV-2 y la urgente necesidad de tratamientos eficaces en ausencia de vacunas, el objetivo de este estudio fue desarrollar un suero hiperinmune (HS) rápido y rentable derivado de ovejas. y evaluar su eficacia. La utilización de un animal con certificación halal, de fácil mantenimiento en determinadas regiones geográficas y de fácil manejo, como las ovejas, podría proporcionar una alternativa viable a la costosa opción de los caballos. Las ovejas fueron inmunizadas con un antígeno de SARS-CoV-2 completamente inactivado para producir HS, cuya potencia neutralizante se evaluó mediante el ensayo PRNT50. Los ratones transgénicos K18-hACE2 (n = 35) se dividieron en tres grupos: control, exposición al SARS-CoV-2 mediante inhalación y ratones expuestos al SARS-CoV-2 tratados con HS. La eficacia de HS se evaluó mediante niveles de citoquinas proinflamatorias en suero, análisis qRT-PCR, examen histopatológico de pulmones y corazones y microscopía electrónica de transmisión. El HS purificado exhibió una actividad neutralizante significativa (1/24,576). El grupo SARS-CoV-2+HS mostró niveles más bajos de TNF-α, IL-10 e IL-6 (P<0,01) y niveles relativamente más bajos de MCP-1 en comparación con el grupo SARS-CoV-2. HS evitó la muerte, redujo los niveles de ARN viral en los pulmones y el corazón, protegió contra la neumonía intersticial grave, preservó la integridad del tejido pulmonar y evitó el daño de los miocitos, mientras que el grupo SARS-CoV-2 exhibió presencia viral en los pulmones. Este estudio desarrolló con éxito un HS derivado de ovejas contra todo el virus SARS-CoV-2, lo que resultó en una reducción significativa de la gravedad de la infección, la inflamación y la producción sistémica de citocinas. Los hallazgos son prometedores para el tratamiento de casos graves de COVID- 19, incluidas las variantes virales emergentes y los pacientes inmunocomprometidos.

Covalent Inhibitors from Saudi Medicinal Plants Target RNA-Dependent RNA Polymerase (RdRp) of SARS-CoV-2.

COVID-19, a disease caused by SARS-CoV-2, has caused a huge loss of human life, and the number of deaths is still continuing. Despite the lack of repurposed drugs and vaccines, the search for potential small molecules to inhibit SARS-CoV-2 is in demand. Hence, we relied on the drug-like characters of ten phytochemicals (compounds 1-10) that were previously isolated and purified by our research team from Saudi medicinal plants. We computationally evaluated the inhibition of RNA-dependent RNA polymerase (RdRp) by compounds 1-10. Non-covalent (reversible) docking of compounds 1-10 with RdRp led to the formation of a hydrogen bond with template primer nucleotides (A and U) and key amino acid residues (ASP623, LYS545, ARG555, ASN691, SER682, and ARG553) in its active pocket. Covalent (irreversible) docking revealed that compounds 7, 8, and 9 exhibited their irreversible nature of binding with CYS813, a crucial amino acid in the palm domain of RdRP. Molecular dynamic (MD) simulation analysis by RMSD, RMSF, and Rg parameters affirmed that RdRP complexes with compounds 7, 8, and 9 were stable and showed less deviation. Our data provide novel information on compounds 7, 8, and 9 that demonstrated their non-nucleoside and irreversible interaction capabilities to inhibit RdRp and shed new scaffolds as antivirals against SARS-CoV-2.

In silico studies of anti-oxidative and hot temperament-based phytochemicals as natural inhibitors of SARS-CoV-2 Mpro.

Main protease (Mpro) of SARS-CoV-2 is considered one of the key targets due to its role in viral replication. The use of traditional phytochemicals is an important part of complementary/alternative medicine, which also accompany the concept of temperament, where it has been shown that hot medicines cure cold and cold medicines cure hot, with cold and hot pattern being associated with oxidative and anti-oxidative properties in medicine, respectively. Molecular docking in this study has demonstrated that a number of anti-oxidative and hot temperament-based phytochemicals have high binding affinities to SARS-CoV-2 Mpro, both in the monomeric and dimeric deposited states of the protein. The highest ranking phytochemicals identified in this study included savinin, betulinic acid and curcumin. Complexes of savinin, betulinic acid, curcumin as well as Nirmatrelvir (the only approved inhibitor, used for comparison) bound to SARS-CoV-2 Mpro were further subjected to molecular dynamics simulations. Subsequently, RMSD, RMSF, Rg, number of hydrogen bonds, binding free energies and residue contributions (using MM-PBSA) and buried surface area (BSA), were analysed. The computational results suggested high binding affinities of savinin, betulinic acid and curcumin to both the monomeric and dimeric deposited states of Mpro, while highlighting the lower binding energy of betulinic acid in comparison with savinin and curcumin and even Nirmatrelvir, leading to a greater stability of the betulinic acid-SARS-CoV-2 Mpro complex. Overall, based on the increasing mutation rate in the spike protein and the fact that the SARS-CoV-2 Mpro remains highly conserved, this study provides an insight into the use of phytochemicals against COVID-19 and other coronavirus diseases.