RESUMO
The SARS-CoV-2 papain-like protease (PLpro), essential for viral processing and immune response disruption, is a promising target for treating acute infection of SARS-CoV-2. To date, there have been no reports of PLpro inhibitors with both submicromolar potency and animal model efficacy. To address the challenge of PLpro's featureless active site, a noncovalent inhibitor library with over 50 new analogs was developed, targeting the PLpro active site by modulating the BL2-loop and engaging the BL2-groove. Notably, compounds 42 and 10 exhibited strong antiviral effects and were further analyzed pharmacokinetically. 10, in particular, showed a significant lung accumulation, up to 12.9-fold greater than plasma exposure, and was effective in a mouse model of SARS-CoV-2 infection, as well as against several SARS-CoV-2 variants. These findings highlight the potential of 10 as an in vivo chemical probe for studying PLpro inhibition in SARS-CoV-2 infection.
Assuntos
Antivirais , Tratamento Farmacológico da COVID-19 , Proteases Semelhantes à Papaína de Coronavírus , SARS-CoV-2 , Animais , Humanos , Camundongos , Antivirais/farmacologia , Antivirais/química , Antivirais/farmacocinética , Antivirais/síntese química , Domínio Catalítico , Proteases 3C de Coronavírus/antagonistas & inibidores , Proteases 3C de Coronavírus/metabolismo , Proteases Semelhantes à Papaína de Coronavírus/antagonistas & inibidores , Proteases Semelhantes à Papaína de Coronavírus/metabolismo , COVID-19/virologia , Inibidores de Proteases/farmacologia , Inibidores de Proteases/química , Inibidores de Proteases/farmacocinética , Inibidores de Proteases/síntese química , SARS-CoV-2/efeitos dos fármacos , Relação Estrutura-AtividadeRESUMO
The coronavirus disease caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has been spread worldwide for more than 3 years. Although the hospitalization rate and mortality have decreased dramatically due to wide vaccination effort and improved treatment options, the disease is still a global health issue due to constant viral mutations, causing negative impact on social and economic activities. In addition, long COVID and complications arising from COVID-19 weeks after infection have become a concern for public health experts. Therefore, better treatments for COVID-19 are still needed. Herein, we describe a class of macrocyclic peptidomimetic compounds that are potent inhibitors of SARS-Cov-2 3CL protease (3CLpro). Significantly, some of the compounds showed a higher stability against human liver microsomes (HLM t1/2 > 180 min) and may be suitable for oral administration without the need for a pharmacokinetic (PK) boosting agent such as ritonavir.
Assuntos
Antivirais , Proteases 3C de Coronavírus , Compostos Macrocíclicos , SARS-CoV-2 , Proteases 3C de Coronavírus/antagonistas & inibidores , Proteases 3C de Coronavírus/metabolismo , Humanos , SARS-CoV-2/efeitos dos fármacos , Compostos Macrocíclicos/química , Compostos Macrocíclicos/farmacologia , Compostos Macrocíclicos/síntese química , Compostos Macrocíclicos/farmacocinética , Antivirais/farmacologia , Antivirais/química , Antivirais/síntese química , Antivirais/farmacocinética , Microssomos Hepáticos/metabolismo , Peptidomiméticos/farmacologia , Peptidomiméticos/química , Peptidomiméticos/síntese química , Descoberta de Drogas , Tratamento Farmacológico da COVID-19 , Inibidores de Proteases/farmacologia , Inibidores de Proteases/química , Inibidores de Proteases/síntese química , Inibidores de Proteases/farmacocinética , Relação Estrutura-AtividadeRESUMO
The 3CL protease (3CLpro, Mpro) plays a key role in the replication of the SARS-CoV-2 and was validated as therapeutic target by the development and approval of specific antiviral drugs (nirmatrelvir, ensitrelvir), inhibitors of this protease. Moreover, its high conservation within the coronavirus family renders it an attractive therapeutic target for the development of anti-coronavirus compounds with broad spectrum activity to control COVID-19 and future coronavirus diseases. Here we report on the design, synthesis and structure-activity relationships of a new series of small covalent reversible inhibitors of the SARS-CoV-2 3CLpro. As elucidated thanks to the X-Ray structure of some inhibitors with the 3CLpro, the mode of inhibition involves acylation of the thiol of the catalytic cysteine. The synthesis of 60 analogs led to the identification of compound 56 that inhibits the SARS-CoV-2 3CLpro with high potency (IC50 = 70 nM) and displays antiviral activity in cells (EC50 = 3.1 µM). Notably, compound 56 inhibits the 3CLpro of three other human coronaviruses and exhibit a good selectivity against two human cysteine proteases. These results demonstrate the potential of this electrophilic N-acylbenzimidazole series as a basis for further optimization.
Assuntos
Antivirais , Benzimidazóis , Proteases 3C de Coronavírus , SARS-CoV-2 , Proteases 3C de Coronavírus/antagonistas & inibidores , Proteases 3C de Coronavírus/metabolismo , Antivirais/farmacologia , Antivirais/química , Antivirais/síntese química , Relação Estrutura-Atividade , SARS-CoV-2/efeitos dos fármacos , SARS-CoV-2/enzimologia , Humanos , Benzimidazóis/farmacologia , Benzimidazóis/química , Benzimidazóis/síntese química , Cisteína Endopeptidases/metabolismo , Acilação , Cisteína/química , Cisteína/farmacologia , Estrutura Molecular , Relação Dose-Resposta a Droga , Inibidores de Proteases/farmacologia , Inibidores de Proteases/síntese química , Inibidores de Proteases/química , Modelos Moleculares , Desenho de Fármacos , Cristalografia por Raios XRESUMO
Papain-like protease (PLpro) is a promising therapeutic target for its pivotal role in the life cycle of SARS-CoV-2. A series of 1,2,4-oxadiazole derivatives was designed and synthesized via a ring formation strategy based on SARS-CoV-2 PLpro-GRL0617 complex structure. Systematic structure-activity relationship studies revealed that introducing oxadiazole and aryl carboxylic acid moieties to GRL0617 enhanced the enzymatic inhibition activity, affinity, and deubiquitination capacity toward PLpro. 1,2,4-Oxadiazole compounds 13f and 26r, which had PLpro inhibition activity (IC50 = 1.8 and 1.0 µM) and antiviral activity against SARS-CoV-2 (EC50 = 5.4 and 4.3 µM), exhibited good metabolic stability (t1/2 > 93.2 min) and higher plasma exposure (AUC0-t = 17,380.08 and 24,289.76 ng·h/mL) in mice. Especially, compound 26r with moderate oral bioavailability of 39.1% and potent antiviral activity is worthy of further studies in vivo. Our findings provide a new insight for the discovery of antiviral agents targeting PLpro.
Assuntos
Antivirais , Desenho de Fármacos , Oxidiazóis , SARS-CoV-2 , Oxidiazóis/química , Oxidiazóis/farmacologia , Oxidiazóis/síntese química , Oxidiazóis/farmacocinética , Animais , Antivirais/farmacologia , Antivirais/síntese química , Antivirais/química , Antivirais/farmacocinética , Relação Estrutura-Atividade , SARS-CoV-2/efeitos dos fármacos , Camundongos , Humanos , Ácidos Carboxílicos/química , Ácidos Carboxílicos/farmacologia , Ácidos Carboxílicos/síntese química , Simulação de Acoplamento Molecular , Inibidores de Proteases/farmacologia , Inibidores de Proteases/síntese química , Inibidores de Proteases/química , Inibidores de Proteases/farmacocinética , Tratamento Farmacológico da COVID-19 , Proteases Semelhantes à Papaína de Coronavírus/antagonistas & inibidores , Proteases Semelhantes à Papaína de Coronavírus/metabolismoRESUMO
Rhomboid intramembrane serine proteases have been implicated in several pathologies, and emerge as attractive pharmacological target candidates. The most potent and selective rhomboid inhibitors available to date are peptidyl α-ketoamides, but their selectivity for diverse rhomboid proteases and strategies to modulate it in relevant contexts are poorly understood. This gap, together with the lack of suitable in vitro models, hinders ketoamide development for relevant eukaryotic rhomboid enzymes. Here we explore the structure-activity relationship principles of rhomboid inhibiting ketoamides by medicinal chemistry and enzymatic in vitro and in-cell assays with recombinant rhomboid proteases GlpG, human mitochondrial rhomboid PARL and human RHBDL2. We use X-ray crystallography in lipidic cubic phase to understand the binding mode of one of the best ketoamide inhibitors synthesized here containing a branched terminal substituent bound to GlpG. In addition, to extend the interpretation of the co-crystal structure, we use quantum mechanical calculations and quantify the relative importance of interactions along the inhibitor molecule. These combined experimental analyses implicates that more extensive exploration of chemical space at the prime side is unexpectedly powerful for the selectivity of rhomboid inhibiting ketoamides. Together with variations in the peptide sequence at the non-prime side, or its non-peptidic alternatives, this strategy enables targeted tailoring of potent and selective ketoamides towards diverse rhomboid proteases including disease-relevant ones such as PARL and RHBDL2.
Assuntos
Amidas , Humanos , Relação Estrutura-Atividade , Estrutura Molecular , Amidas/química , Amidas/farmacologia , Amidas/síntese química , Cristalografia por Raios X , Relação Dose-Resposta a Droga , Inibidores de Proteases/farmacologia , Inibidores de Proteases/química , Inibidores de Proteases/síntese química , Inibidores de Proteases/metabolismo , Modelos MolecularesRESUMO
The global outbreak of the COVID-19 pandemic caused by the SARS-CoV-2 virus had led to profound respiratory health implications. This study focused on designing organoselenium-based inhibitors targeting the SARS-CoV-2 main protease (Mpro). The ligand-binding pathway sampling method based on parallel cascade selection molecular dynamics (LB-PaCS-MD) simulations was employed to elucidate plausible paths and conformations of ebselen, a synthetic organoselenium drug, within the Mpro catalytic site. Ebselen effectively engaged the active site, adopting proximity to H41 and interacting through the benzoisoselenazole ring in a π-π T-shaped arrangement, with an additional π-sulfur interaction with C145. In addition, the ligand-based drug design using the QSAR with GFA-MLR, RF, and ANN models were employed for biological activity prediction. The QSAR-ANN model showed robust statistical performance, with an r2training exceeding 0.98 and an RMSEtest of 0.21, indicating its suitability for predicting biological activities. Integration the ANN model with the LB-PaCS-MD insights enabled the rational design of novel compounds anchored in the ebselen core structure, identifying promising candidates with favorable predicted IC50 values. The designed compounds exhibited suitable drug-like characteristics and adopted an active conformation similar to ebselen, inhibiting Mpro function. These findings represent a synergistic approach merging ligand and structure-based drug design; with the potential to guide experimental synthesis and enzyme assay testing.
Assuntos
Antivirais , Proteases 3C de Coronavírus , Desenho de Fármacos , Isoindóis , Aprendizado de Máquina , Simulação de Dinâmica Molecular , Compostos Organosselênicos , Inibidores de Proteases , Relação Quantitativa Estrutura-Atividade , SARS-CoV-2 , SARS-CoV-2/efeitos dos fármacos , SARS-CoV-2/enzimologia , Compostos Organosselênicos/química , Compostos Organosselênicos/farmacologia , Compostos Organosselênicos/síntese química , Isoindóis/química , Isoindóis/farmacologia , Isoindóis/síntese química , Proteases 3C de Coronavírus/antagonistas & inibidores , Proteases 3C de Coronavírus/metabolismo , Inibidores de Proteases/química , Inibidores de Proteases/farmacologia , Inibidores de Proteases/síntese química , Antivirais/farmacologia , Antivirais/química , Antivirais/síntese química , Humanos , Azóis/química , Azóis/farmacologia , Azóis/síntese química , COVID-19/virologia , Domínio CatalíticoRESUMO
The present work elicits a novel approach to combating COVID-19 by synthesizing a series of azo-anchored 3,4-dihydroimidazo[4,5-b]indole derivatives. The envisaged methodology involves the L-proline-catalyzed condensation of para-amino-functionalized azo benzene, indoline-2,3-dione, and ammonium acetate precursors with pertinent aryl aldehyde derivatives under ultrasonic conditions. The structures of synthesized compounds were corroborated through FT-IR, 1H NMR, 13C NMR, and mass analysis data. Molecular docking studies assessed the inhibitory potential of these compounds against the main protease (Mpro) of SARS-CoV-2. Remarkably, in silico investigations revealed significant inhibitory action surpassing standard drugs such as Remdesivir, Paxlovid, Molnupiravir, Chloroquine, Hydroxychloroquine (HCQ), and (N3), an irreversible Michael acceptor inhibitor. Furthermore, the highly active compound was also screened for cytotoxicity activity against HEK-293 cells and exhibited minimal toxicity across a range of concentrations, affirming its favorable safety profile and potential suitability. The pharmacokinetic properties (ADME) of the synthesized compounds have also been deliberated. This study paves the way for in vitro and in vivo testing of these scaffolds in the ongoing battle against SARS-CoV-2.
Assuntos
Antivirais , Tratamento Farmacológico da COVID-19 , Proteases 3C de Coronavírus , Indóis , Simulação de Acoplamento Molecular , Inibidores de Proteases , SARS-CoV-2 , Humanos , Antivirais/farmacologia , Antivirais/química , Antivirais/síntese química , SARS-CoV-2/efeitos dos fármacos , Proteases 3C de Coronavírus/antagonistas & inibidores , Proteases 3C de Coronavírus/metabolismo , Proteases 3C de Coronavírus/química , Indóis/farmacologia , Indóis/química , Indóis/síntese química , Células HEK293 , Inibidores de Proteases/farmacologia , Inibidores de Proteases/química , Inibidores de Proteases/síntese química , Imidazóis/farmacologia , Imidazóis/química , Imidazóis/síntese química , Simulação por Computador , COVID-19/virologia , Compostos Azo/farmacologia , Compostos Azo/química , Compostos Azo/síntese químicaRESUMO
The progressive emergence of SARS-CoV-2 variants has necessitated the urgent exploration of novel therapeutic strategies to combat the COVID-19 pandemic. The SARS-CoV-2 main protease (Mpro) represents an evolutionarily conserved therapeutic target for drug discovery. This study highlights the discovery of meisoindigo (Mei), derived from the traditional Chinese medicine (TCM) Indigo naturalis, as a novel non-covalent and nonpeptidic Mpro inhibitor. Substantial optimizations and structure-activity relationship (SAR) studies, guided by a structure-based drug design approach, led to the identification of several Mei derivatives, including S5-27 and S5-28, exhibiting low micromolar inhibition against SARS-CoV-2 Mpro with high binding affinity. Notably, S5-28 provided significant protection against wild-type SARS-CoV-2 in HeLa-hACE2 cells, with EC50 up to 2.66 µM. Furthermore, it displayed favorable physiochemical properties and remarkable gastrointestinal and metabolic stability, demonstrating its potential as an orally bioavailable drug for anti-COVID-19 therapy. This research presents a promising avenue for the development of new antiviral agents, offering hope in the ongoing battle against COVID-19.
Assuntos
Antivirais , Tratamento Farmacológico da COVID-19 , Proteases 3C de Coronavírus , SARS-CoV-2 , Humanos , SARS-CoV-2/efeitos dos fármacos , Antivirais/farmacologia , Antivirais/química , Antivirais/síntese química , Relação Estrutura-Atividade , Proteases 3C de Coronavírus/antagonistas & inibidores , Proteases 3C de Coronavírus/metabolismo , Descoberta de Drogas , Administração Oral , Animais , Indóis/química , Indóis/farmacologia , Indóis/síntese química , Células HeLa , COVID-19/virologia , Estrutura Molecular , Ratos , Testes de Sensibilidade Microbiana , Inibidores de Proteases/farmacologia , Inibidores de Proteases/química , Inibidores de Proteases/síntese química , Inibidores de Proteases/uso terapêutico , Simulação de Acoplamento Molecular , Desenho de FármacosRESUMO
Given the crucial role of the main protease (Mpro) in the replication cycle of SARS-CoV-2, this viral cysteine protease constitutes a high-profile drug target. We investigated peptidomimetic azapeptide nitriles as auspicious, irreversibly acting inhibitors of Mpro. Our systematic approach combined an Mpro active-site scanning by combinatorially assembled azanitriles with structure-based design. Encouraged by the bioactive conformation of open-chain inhibitors, we conceptualized the novel chemotype of macrocyclic azanitriles whose binding mode was elucidated by cocrystallization. This strategy provided a favorable entropic contribution to target binding and resulted in the development of the extraordinarily potent Mpro inhibitor 84 with an IC50 value of 3.23 nM and a second-order rate constant of inactivation, kinac/Ki, of 448,000 M-1s-1. The open-chain Mpro inhibitor 58, along with the macrocyclic compounds 83 and 84, a broad-spectrum anticoronaviral agent, demonstrated the highest antiviral activity with EC50 values in the single-digit micromolar range. Our findings are expected to promote the future development of peptidomimetic Mpro inhibitors as anti-SARS-CoV-2 agents.
Assuntos
Antivirais , Proteases 3C de Coronavírus , Nitrilas , SARS-CoV-2 , Antivirais/farmacologia , Antivirais/química , Antivirais/síntese química , SARS-CoV-2/efeitos dos fármacos , Nitrilas/química , Nitrilas/farmacologia , Nitrilas/síntese química , Proteases 3C de Coronavírus/antagonistas & inibidores , Proteases 3C de Coronavírus/metabolismo , Proteases 3C de Coronavírus/química , Relação Estrutura-Atividade , Humanos , Compostos Macrocíclicos/farmacologia , Compostos Macrocíclicos/química , Compostos Macrocíclicos/síntese química , Tratamento Farmacológico da COVID-19 , Descoberta de Drogas , Inibidores de Proteases/farmacologia , Inibidores de Proteases/química , Inibidores de Proteases/síntese química , Peptidomiméticos/farmacologia , Peptidomiméticos/química , Peptidomiméticos/síntese química , Inibidores de Cisteína Proteinase/farmacologia , Inibidores de Cisteína Proteinase/química , Inibidores de Cisteína Proteinase/síntese química , Peptídeos/química , Peptídeos/farmacologia , Peptídeos/síntese químicaRESUMO
Background: The epidemic caused by SARS-CoV-2 swept the world in 2019. The 3C-like protease (3CLpro) of SARS-CoV-2 plays a key role in viral replication, and its inhibition could inhibit viral replication. Materials & methods: The virtual screen based on receptor-ligand pharmacophore models and molecular docking were conducted to obtain the novel scaffolds of the 3CLpro. The molecular dynamics simulation was also carried out. All compounds were synthesized and evaluated in biochemical assays. Results: The compound C2 could inhibit 3CLpro with a 72% inhibitory rate at 10 µM. The covalent docking showed that C2 could form a covalent bond with the Cys145 in 3CLpro. Conclusion: C2 could be a potent lead compound of 3CLpro inhibitors against SARS-CoV-2.
[Box: see text].
Assuntos
Antivirais , Proteases 3C de Coronavírus , Desenho de Fármacos , Simulação de Acoplamento Molecular , Simulação de Dinâmica Molecular , SARS-CoV-2 , Proteases 3C de Coronavírus/antagonistas & inibidores , Proteases 3C de Coronavírus/metabolismo , SARS-CoV-2/efeitos dos fármacos , Antivirais/farmacologia , Antivirais/síntese química , Antivirais/química , Humanos , Inibidores de Proteases/farmacologia , Inibidores de Proteases/síntese química , Inibidores de Proteases/química , Tratamento Farmacológico da COVID-19 , Relação Estrutura-AtividadeRESUMO
Emerging RNA viruses, including SARS-CoV-2, continue to be a major threat. Cell entry of SARS-CoV-2 particles via the endosomal pathway involves cysteine cathepsins. Due to ubiquitous expression, cathepsin L (CatL) is considered a promising drug target in the context of different viral and lysosome-related diseases. We characterized the anti-SARS-CoV-2 activity of a set of carbonyl- and succinyl epoxide-based inhibitors, which were previously identified as inhibitors of cathepsins or related cysteine proteases. Calpain inhibitor XII, MG-101, and CatL inhibitor IV possess antiviral activity in the very low nanomolar EC50 range in Vero E6 cells and inhibit CatL in the picomolar Ki range. We show a relevant off-target effect of CatL inhibition by the coronavirus main protease α-ketoamide inhibitor 13b. Crystal structures of CatL in complex with 14 compounds at resolutions better than 2 Å present a solid basis for structure-guided understanding and optimization of CatL inhibitors toward protease drug development.
Assuntos
Antivirais , Catepsina L , SARS-CoV-2 , Catepsina L/antagonistas & inibidores , Catepsina L/metabolismo , Antivirais/farmacologia , Antivirais/química , Antivirais/síntese química , Animais , Chlorocebus aethiops , Células Vero , SARS-CoV-2/efeitos dos fármacos , Humanos , Relação Estrutura-Atividade , Inibidores de Cisteína Proteinase/farmacologia , Inibidores de Cisteína Proteinase/química , Inibidores de Cisteína Proteinase/síntese química , Cristalografia por Raios X , Inibidores de Proteases/farmacologia , Inibidores de Proteases/química , Inibidores de Proteases/síntese química , Inibidores de Proteases/metabolismo , Modelos MolecularesRESUMO
El vaso constrictor, angiotensina II y el péptido natriurético auricular (PNA) tienen funciones opuestas en el desarrollo de la insuficiencia cardíaca congestiva realizada en un modelo de insuficiencia cardíaca (ICC) en ratas. Este modelo de insuficiencia cardíaaca se caracteriza por una retención de sal a pesar del aumento de la concentración plasmática de PNA, indicando la ausencia de respuestas renales al PNA. S21402 (RB105), N-2S, 3R-(2-mercaptomethyl-1-oxo-3-phenyl butyl)L-alanine]}, es un inhibidor mixto que inhibe la enzima convertidora de angiotensina (ECA; Ki=4,2ñ0,5nM) y la endopeptidasa neutra (NEP;Ki=1,7ñ0,3nM). La inhibición de la NEP potencializa el PNA endógeno, y la inhibición de la ECA bloquea la producción de angiotensina II. La inhibición de las dos peptidasas permite una potencialización más completa de la bradicinina (BK). La inducción de la insuficiencia cardíaca en ratas por ligadura de las coronarias produjo el aumento de las concentraciones plasmáticas basales de PNA, de cGMP y de la renina, las cuales están relacionadas con el tamaño del infarto (moderados y severos). El PNA, el cGMP y la BK urinarias aumentan en las ratas con ICC en relación al tamaño del infarto. El S21402 (RB105) (25 mg.kg-1 i.v.) indujo un descenso de la presión arterial y un aumento de la natriuresis en función de la severidad del infarto. El compuesto S21402 (RB105) indujo un aumento de la renina plasmática a pesar del nivel basal elevado de la renina en los animales con ICC. El inhibidor mixto no produjo aumento detectable del PNA plasmático. Sin embargo, el cGMP plasmático fue aumentado por el compuesto S21402 (RB105) en función de la severidad del infarto. Las excreciones urinarias del PNA, del cGMP y de la BK fueron también aumentadas por el inhibidor mixto en función de la severidad del infarto. Independientemente del tamaño del infarto, el S21402 (RB105) es capaz de inducir natriuresis en un modelo caracterizado por la desensibilidad a la acción del PNA. La inhibición de ambas peptidasas (MEP,ECA) bloquea también la producción de angiotensina II. Esta combinación de efectos renales y vasculares podría tener un interés terapéutico de la insuficiencia cardíaca congestiva