ABSTRACT
The COVID-19 pandemic has led to the deaths of millions of people and severe global economic impacts. Small molecule therapeutics have played an important role in the fight against SARS-CoV-2, the virus responsible for COVID-19, but their efficacy has been limited in scope and availability, with many people unable to access their benefits, and better options are needed. EDP-235 is specifically designed to inhibit the SARS-CoV-2 3CLpro, with potent nanomolar activity against all SARS-CoV-2 variants to date, as well as clinically relevant human and zoonotic coronaviruses. EDP-235 maintains potency against variants bearing mutations associated with nirmatrelvir resistance. Additionally, EDP-235 demonstrates a ≥ 500-fold selectivity index against multiple host proteases. In a male Syrian hamster model of COVID-19, EDP-235 suppresses SARS-CoV-2 replication and viral-induced hamster lung pathology. In a female ferret model, EDP-235 inhibits production of SARS-CoV-2 infectious virus and RNA at multiple anatomical sites. Furthermore, SARS-CoV-2 contact transmission does not occur when naïve ferrets are co-housed with infected, EDP-235-treated ferrets. Collectively, these results demonstrate that EDP-235 is a broad-spectrum coronavirus inhibitor with efficacy in animal models of primary infection and transmission.
Subject(s)
Antiviral Agents , COVID-19 , Coronavirus 3C Proteases , SARS-CoV-2 , Virus Replication , Animals , Cricetinae , Female , Humans , Male , Antiviral Agents/pharmacology , Chlorocebus aethiops , Coronavirus 3C Proteases/antagonists & inhibitors , Coronavirus 3C Proteases/metabolism , COVID-19/virology , COVID-19/transmission , COVID-19 Drug Treatment , Disease Models, Animal , Ferrets , Lactams , Leucine , Lung/virology , Lung/drug effects , Lung/pathology , Mesocricetus , Nitriles , Organic Chemicals , Pandemics/prevention & control , Pneumonia, Viral/drug therapy , Pneumonia, Viral/virology , Pneumonia, Viral/transmission , Pneumonia, Viral/prevention & control , Proline , SARS-CoV-2/drug effects , SARS-CoV-2/genetics , SARS-CoV-2/physiology , Vero Cells , Virus Replication/drug effectsABSTRACT
The fungus Talaromyces hainanensis, isolated from the mangrove soil, was characterized as a novel species by morphology observation and phylogenetic analyses. Four new γ-lactam alkaloids talaroilactams A-D (1-4) and two reported compounds harzianic acid (5) and isoharzianic acid (6) were identified from the fungus T. hainanensis WHUF0341, assisted by OSMAC along with molecular networking approaches. Their structures were determined through ECD calculations and spectroscopic analyses. Moreover, the biosynthetic route of 1-4 was also proposed. Compound 1 displayed potent cytotoxicity against HepG2 cell lines, with an IC50 value of 10.75 ± 1.11 µM. In addition, network pharmacology was employed to dissect the probable mechanisms contributing to the antihepatocellular carcinoma effects of compound 1, revealing that cytotoxicity was mainly associated with proteolysis, negative regulation of autophagy, inflammatory response, and the renin-angiotensin system. These results not only expanded the chemical space of natural products from the mangrove associated fungi but also afforded promising lead compounds for developing the antihepatocellular carcinoma agents.
Subject(s)
Alkaloids , Antineoplastic Agents , Lactams , Talaromyces , Talaromyces/chemistry , Talaromyces/metabolism , Humans , Alkaloids/pharmacology , Alkaloids/chemistry , Hep G2 Cells , Antineoplastic Agents/pharmacology , Antineoplastic Agents/chemistry , Lactams/chemistry , Lactams/pharmacology , Phylogeny , Molecular Structure , Wetlands , Rhizophoraceae/microbiology , Rhizophoraceae/chemistryABSTRACT
To the best of our knowledge, enzymes that catalyse intramolecular Diels-Alder ([4+2] cycloaddition) reactions are frequently reported in natural product biosynthesis; however, no native enzymes utilising Lewis acid catalysis have been reported. Verticilactam is a representative member of polycyclic macrolactams, presumably produced by spontaneous cycloaddition. We report that the intramolecular [4+2] cycloadditions can be significantly accelerated by ferredoxins (Fds), a class of small iron-sulphur (Fe-S) proteins. Through iron atom substitution by Lewis acidic gallium (Ga) iron and computational calculations, we confirm that the ubiquitous Fe-S cluster efficiently functions as Lewis acid to accelerate the tandem [4+2] cycloaddition and Michael addition reactions by lowering free energy barriers. Our work highlights Nature's ingenious strategy to generate complex molecule structures using the ubiquitous Fe-S protein. Furthermore, our study sheds light on the future design of Fd as a versatile Lewis acid catalyst for [4+2] cycloaddition reactions.
Subject(s)
Biological Products , Cycloaddition Reaction , Iron-Sulfur Proteins , Lewis Acids , Biological Products/metabolism , Biological Products/chemistry , Iron-Sulfur Proteins/metabolism , Iron-Sulfur Proteins/chemistry , Lewis Acids/chemistry , Lewis Acids/metabolism , Catalysis , Iron/chemistry , Iron/metabolism , Lactams/metabolism , Lactams/chemistry , BiocatalysisABSTRACT
Exploring therapeutic options is crucial in the ongoing COVID-19 pandemic caused by SARS-CoV-2. Nirmatrelvir, which is a potent inhibitor that targets the SARS-CoV-2 Mpro, shows promise as an antiviral treatment. Additionally, Ivermectin, which is a broad-spectrum antiparasitic drug, has demonstrated effectiveness against the virus in laboratory settings. However, its clinical implications are still debated. Using computational methods, such as molecular docking and 100 ns molecular dynamics simulations, we investigated how Nirmatrelvir and Ivermectin interacted with SARS-CoV-2 Mpro(A). Calculations using density functional theory were instrumental in elucidating the behavior of isolated molecules, primarily by analyzing the frontier molecular orbitals. Our analysis revealed distinct binding patterns: Nirmatrelvir formed strong interactions with amino acids, like MET49, MET165, HIS41, HIS163, HIS164, PHE140, CYS145, GLU166, and ASN142, showing stable binding, with a root-mean-square deviation (RMSD) of around 2.0 Å. On the other hand, Ivermectin interacted with THR237, THR239, LEU271, LEU272, and LEU287, displaying an RMSD of 1.87 Å, indicating enduring interactions. Both ligands stabilized Mpro(A), with Ivermectin showing stability and persistent interactions despite forming fewer hydrogen bonds. These findings offer detailed insights into how Nirmatrelvir and Ivermectin bind to the SARS-CoV-2 main protease, providing valuable information for potential therapeutic strategies against COVID-19.
Subject(s)
Antiviral Agents , COVID-19 Drug Treatment , Coronavirus 3C Proteases , Ivermectin , Molecular Docking Simulation , Molecular Dynamics Simulation , SARS-CoV-2 , Ivermectin/chemistry , Ivermectin/pharmacology , SARS-CoV-2/drug effects , SARS-CoV-2/enzymology , Coronavirus 3C Proteases/chemistry , Coronavirus 3C Proteases/antagonists & inhibitors , Coronavirus 3C Proteases/metabolism , Humans , Antiviral Agents/chemistry , Antiviral Agents/pharmacology , Protein Binding , Sulfonamides/chemistry , Sulfonamides/pharmacology , Binding Sites , Protease Inhibitors/chemistry , Protease Inhibitors/pharmacology , Lactams , Leucine , Nitriles , ProlineABSTRACT
To facilitate the detection and management of potential clinical antiviral resistance, in vitro selection of drug-resistant severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) against the virus Mpro inhibitor nirmatrelvir (Paxlovid active component) was conducted. Six Mpro mutation patterns containing T304I alone or in combination with T21I, L50F, T135I, S144A, or A173V emerged, with A173V+T304I and T21I+S144A+T304I mutations showing >20-fold resistance each. Biochemical analyses indicated inhibition constant shifts aligned to antiviral results, with S144A and A173V each markedly reducing nirmatrelvir inhibition and Mpro activity. SARS-CoV-2 surveillance revealed that in vitro resistance-associated mutations from our studies and those reported in the literature were rarely detected in the Global Initiative on Sharing All Influenza Data database. In the Paxlovid Evaluation of Protease Inhibition for COVID-19 in High-Risk Patients trial, E166V was the only emergent resistance mutation, observed in three Paxlovid-treated patients, none of whom experienced COVID-19-related hospitalization or death.
Subject(s)
Antiviral Agents , COVID-19 Drug Treatment , Drug Resistance, Viral , Mutation , SARS-CoV-2 , SARS-CoV-2/genetics , SARS-CoV-2/drug effects , Drug Resistance, Viral/genetics , Humans , Antiviral Agents/pharmacology , Antiviral Agents/therapeutic use , COVID-19/virology , COVID-19/genetics , COVID-19/epidemiology , Coronavirus 3C Proteases/genetics , Coronavirus 3C Proteases/antagonists & inhibitors , Lactams , Leucine , Nitriles , ProlineABSTRACT
Nirmatrelvir (PF-07321332), a first-in-class inhibitor of the severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) main protease (Mpro), was developed by Pfizer under intense pressure during the pandemic to treat COVID-19. A weakness of nirmatrelvir is its limited metabolic stability, which led to the development of a combination therapy (paxlovid), involving coadministration of nirmatrelvir with the cytochrome P450 inhibitor ritonavir. However, limitations in tolerability of the ritonavir component reduce the scope of paxlovid. In response to these limitations, researchers at Pfizer have now developed the second-generation Mpro inhibitor PF-07817883 (ibuzatrelvir). Structurally related to nirmatrelvir, including with the presence of a trifluoromethyl group, albeit located differently, ibuzatrelvir manifests enhanced oral bioavailability, so it does not require coadministration with ritonavir. The development of ibuzatrelvir is an important milestone, because it is expected to enhance the treatment of COVID-19 without the drawbacks associated with ritonavir. Given the success of paxlovid in treating COVID-19, it is likely that ibuzatrelvir will be granted approval as an improved drug for treatment of COVID-19 infections, so complementing vaccination efforts and improving pandemic preparedness. The development of nirmatrelvir and ibuzatrelvir dramatically highlights the power of appropriately resourced modern medicinal chemistry to very rapidly enable the development of breakthrough medicines. Consideration of how analogous approaches can be used to develop similarly breakthrough medicines for infectious diseases such as tuberculosis and malaria is worthwhile.
Subject(s)
Antiviral Agents , COVID-19 Drug Treatment , SARS-CoV-2 , Humans , SARS-CoV-2/drug effects , Antiviral Agents/therapeutic use , Antiviral Agents/pharmacology , Ritonavir/therapeutic use , Coronavirus 3C Proteases/antagonists & inhibitors , Coronavirus 3C Proteases/metabolism , Indazoles/therapeutic use , Lactams , Leucine , Nitriles , ProlineABSTRACT
BACKGROUND: Clinical trials of treatments for coronavirus disease 2019 (Covid-19) have not shown a significant benefit of postexposure prophylaxis. METHODS: We conducted a phase 2-3 double-blind trial to assess the efficacy and safety of nirmatrelvir-ritonavir in asymptomatic, rapid antigen test-negative adults who had been exposed to a household contact with Covid-19 within 96 hours before randomization. The participants were randomly assigned in a 1:1:1 ratio to receive nirmatrelvir-ritonavir (300 mg of nirmatrelvir and 100 mg of ritonavir) every 12 hours for 5 days or for 10 days or matching placebo for 5 or 10 days. The primary end point was the development of symptomatic SARS-CoV-2 (severe acute respiratory syndrome coronavirus 2) infection, confirmed on reverse-transcriptase-polymerase-chain-reaction (RT-PCR) or rapid antigen testing, through 14 days in participants who had a negative RT-PCR test at baseline. RESULTS: A total of 2736 participants were randomly assigned to a trial group - 921 to the 5-day nirmatrelvir-ritonavir group, 917 to the 10-day nirmatrelvir-ritonavir group, and 898 to the placebo group. Symptomatic, confirmed SARS-CoV-2 infection developed by day 14 in 2.6% of the participants in the 5-day nirmatrelvir-ritonavir group, 2.4% of those in the 10-day nirmatrelvir-ritonavir group, and 3.9% of those in the placebo group. In each nirmatrelvir-ritonavir group, the percentage of participants in whom symptomatic, confirmed SARS-CoV-2 infection developed did not differ significantly from that in the placebo group, with risk reductions relative to placebo of 29.8% (95% confidence interval [CI], -16.7 to 57.8; P = 0.17) in the 5-day nirmatrelvir-ritonavir group and 35.5% (95% CI, -11.5 to 62.7; P = 0.12) in the 10-day nirmatrelvir-ritonavir group. The incidence of adverse events was similar across the trial groups, with dysgeusia being the most frequently reported adverse event (in 5.9% and 6.8% of the participants in the 5-day and 10-day nirmatrelvir-ritonavir groups, respectively, and in 0.7% of those in the placebo group). CONCLUSIONS: In this placebo-controlled trial, postexposure prophylaxis with nirmatrelvir-ritonavir for 5 or 10 days did not significantly reduce the risk of symptomatic SARS-CoV-2 infection. (Funded by Pfizer; ClinicalTrials.gov number, NCT05047601.).
Subject(s)
COVID-19 Drug Treatment , COVID-19 , Post-Exposure Prophylaxis , Ritonavir , SARS-CoV-2 , Humans , Ritonavir/therapeutic use , Ritonavir/adverse effects , Ritonavir/administration & dosage , Double-Blind Method , Male , Female , Adult , Middle Aged , COVID-19/prevention & control , Administration, Oral , Indazoles/adverse effects , Indazoles/therapeutic use , Drug Combinations , Antiviral Agents/therapeutic use , Antiviral Agents/adverse effects , Antiviral Agents/administration & dosage , Indoles/adverse effects , Indoles/therapeutic use , Indoles/administration & dosage , Young Adult , Drug Therapy, Combination , Lactams , Leucine , Nitriles , ProlineABSTRACT
The present study describes a small library of peptides derived from a potent and selective CXCR4 antagonist (3), wherein the native disulfide bond is replaced using a side-chain to tail macrolactamization technique to vary ring size and amino acid composition. The peptides were preliminary assessed for their ability to interfere with the interaction between the receptor and anti-CXCR4 PE-conjugated antibody clone 12G5. Two promising candidates (13 and 17) were identified and further evaluated in a125I-CXCL12 competition binding assay, exhibiting IC50 in the low-nanomolar range. Furthermore, both candidates displayed high selectivity towards CXCR4 with respect to the cognate receptor CXCR7, ability to block CXCL12-dependent cancer cell migration, and receptor internalization, albeit at a higher concentration compared to 3. Molecular modeling studies on 13 and 17 produced a theoretical model that may serve as a guide for future modifications, aiding in the development of analogs with improved affinity. Finally, the study provides valuable insights into developing therapeutic agents targeting CXCR4-mediated processes, demonstrating the adaptability of our lead peptide 3 to alternative cyclization approaches and offering prospects for comprehensive investigations into the receptor region's interaction with its C-terminal region.
Subject(s)
Disulfides , Peptides , Receptors, CXCR4 , Receptors, CXCR4/antagonists & inhibitors , Receptors, CXCR4/metabolism , Humans , Binding Sites/drug effects , Peptides/chemistry , Peptides/pharmacology , Peptides/chemical synthesis , Disulfides/chemistry , Disulfides/pharmacology , Structure-Activity Relationship , Molecular Structure , Dose-Response Relationship, Drug , Lactams/chemistry , Lactams/pharmacology , Lactams/chemical synthesis , Cell Movement/drug effects , Models, Molecular , Cell Line, TumorABSTRACT
Since influenza virus RNA polymerase subunit PAN is a dinuclear Mn2+ dependent endonuclease, metal-binding pharmacophores (MBPs) with Mn2+ coordination has been elucidated as a promising strategy to develop PAN inhibitors for influenza treatment. However, few attentions have been paid to the relationship between the optimal arrangement of the donor atoms in MBPs and anti-influenza A virus (IAV) efficacy. Given that, the privileged hydroxypyridinones fusing a seven-membered lactam ring with diverse side chains, chiral centers or cyclic systems were designed and synthesized. A structure-activity relationship study resulted in a hit compound 16l (IC50 = 2.868 ± 0.063 µM against IAV polymerase), the seven-membered lactam ring of which was fused a pyrrolidine ring. Further optimization of the hydrophobic binding groups on 16l afforded a lead compound (R, S)-16s, which exhibited a 64-fold more potent inhibitory activity (IC50 = 0.045 ± 0.002 µM) toward IAV polymerase. Moreover, (R, S)-16s demonstrated a potent anti-IAV efficacy (EC50 = 0.134 ± 0.093 µM) and weak cytotoxicity (CC50 = 15.35 µM), indicating the high selectivity of (R, S)-16s. Although the lead compound (R, S)-16s exhibited a little weaker activity than baloxavir, these findings illustrated the utility of a metal coordination-based strategy in generating novel MBPs with potent anti-influenza activity.
Subject(s)
Antiviral Agents , Drug Design , Endonucleases , Influenza A virus , Lactams , Pyridones , Antiviral Agents/pharmacology , Antiviral Agents/chemistry , Antiviral Agents/chemical synthesis , Lactams/chemistry , Lactams/pharmacology , Lactams/chemical synthesis , Structure-Activity Relationship , Endonucleases/antagonists & inhibitors , Endonucleases/metabolism , Pyridones/pharmacology , Pyridones/chemistry , Pyridones/chemical synthesis , Influenza A virus/drug effects , Molecular Structure , Enzyme Inhibitors/pharmacology , Enzyme Inhibitors/chemistry , Enzyme Inhibitors/chemical synthesis , Dose-Response Relationship, Drug , Humans , Microbial Sensitivity Tests , Dogs , Madin Darby Canine Kidney Cells , AnimalsABSTRACT
Lorlatinib is a pharmaceutical ALK kinase inhibitor used to treat ALK driven non-small cell lung cancers. This paper analyses the intersection of past published data on the physiological consequences of two unrelated drugs from general medical practice-itraconazole and cilostazol-with the pathophysiology of ALK positive non-small cell lung cancer. A conclusion from that data analysis is that adding itraconazole and cilostazol may make lorlatinib more effective. Itraconazole, although marketed worldwide as a generic antifungal drug, also inhibits Hedgehog signaling, Wnt signaling, hepatic CYP3A4, and the p-gp efflux pump. Cilostazol, marketed worldwide as a generic thrombosis preventative drug, acts by inhibiting phosphodiesterase 3, and, by so doing, lowers platelets' adhesion, thereby partially depriving malignant cells of the many tumor trophic growth factors supplied by platelets. Itraconazole may enhance lorlatinib effectiveness by (i) reducing or stopping a Hedgehog-ALK amplifying feedback loop, by (ii) increasing lorlatinib's brain levels by p-gp inhibition, and by (iii) inhibiting growth drive from Wnt signaling. Cilostazol, surprisingly, carries minimal bleeding risk, lower than that of aspirin. Risk/benefit assessment of the combination of metastatic ALK positive lung cancer being a low-survival disease with the predicted safety of itraconazole-cilostazol augmentation of lorlatinib favors a trial of this drug trio in ALK positive lung cancer.
Subject(s)
Aminopyridines , Cilostazol , Drug Resistance, Neoplasm , Itraconazole , Humans , Itraconazole/pharmacology , Itraconazole/therapeutic use , Cilostazol/pharmacology , Cilostazol/therapeutic use , Drug Resistance, Neoplasm/drug effects , Aminopyridines/pharmacology , Aminopyridines/therapeutic use , Pyrazoles/pharmacology , Pyrazoles/therapeutic use , Lung Neoplasms/drug therapy , Lung Neoplasms/pathology , Carcinoma, Non-Small-Cell Lung/drug therapy , Carcinoma, Non-Small-Cell Lung/pathology , Drug Repositioning , Lactams/pharmacology , Lactams/therapeutic use , Anaplastic Lymphoma Kinase/metabolism , Anaplastic Lymphoma Kinase/antagonists & inhibitors , Anaplastic Lymphoma Kinase/genetics , Protein Kinase Inhibitors/therapeutic use , Protein Kinase Inhibitors/pharmacologyABSTRACT
Solute Carrier Family 3, Member 2 (SLC3A2 or 4F2hc) is a multifunctional glycoprotein that mediates integrin-dependent signaling, acts as a trafficking chaperone for amino acid transporters, and is involved in polyamine transportation. We identified SLC3A2 as a potential Anaplastic Lymphoma Kinase (ALK) interacting partner in a BioID-proximity labeling screen in neuroblastoma (NB) cells. In this work we show that endogenous SLC3A2 and ALK interact in NB cells and that this SLC3A2:ALK interaction was abrogated upon treatment with the ALK inhibitor lorlatinib. We show here that loss of ALK activity leads to decreased SLC3A2 expression and reduced SLC3A2 protein stability in a panel of NB cell lines, while stimulation of ALK with ALKAL2 ligand resulted in increased SLC3A2 protein levels. We further identified MARCH11, an E3 ligase, as a regulator of SLC3A2 ubiquitination downstream of ALK. Further, knockdown of SLC3A2 resulted in inhibition of NB cell growth. To investigate the therapeutic potential of SLC3A2 targeting, we performed monotreatment of NB cells with AMXT-1501 (a polyamine transport inhibitor), which showed only moderate effects in NB cells. In contrast, a combination lorlatinib/AMXT-1501 treatment resulted in synergistic inhibition of cell growth in ALK-driven NB cell lines. Taken together, our results identify a novel role for the ALK receptor tyrosine kinase (RTK), working in concert with the MARCH11 E3 ligase, in regulating SLC3A2 protein stability and function in NB cells. The synergistic effect of combined ALK and polyamine transport inhibition shows that ALK/MARCH11/SLC3A2 regulation of amino acid transport is important for oncogenic growth and survival in NB cells.
Subject(s)
Anaplastic Lymphoma Kinase , Cell Proliferation , Fusion Regulatory Protein 1, Heavy Chain , Neuroblastoma , Signal Transduction , Humans , Anaplastic Lymphoma Kinase/metabolism , Anaplastic Lymphoma Kinase/antagonists & inhibitors , Anaplastic Lymphoma Kinase/genetics , Neuroblastoma/pathology , Neuroblastoma/metabolism , Fusion Regulatory Protein 1, Heavy Chain/metabolism , Fusion Regulatory Protein 1, Heavy Chain/genetics , Cell Line, Tumor , Cell Proliferation/drug effects , Signal Transduction/drug effects , Ubiquitin-Protein Ligases/metabolism , Ubiquitin-Protein Ligases/genetics , Lactams/pharmacology , Aminopyridines/pharmacology , Ubiquitination/drug effects , Pyrazoles/pharmacologyABSTRACT
BACKGROUND: In randomized controlled trials, Nirmatrelvir/ritonavir (NMV/r) and Molnupiravir (MPV) reduced the risk of severe/fatal COVID-19 disease. Real-world data are limited, particularly studies directly comparing the two agents. METHODS: Using the VA National COVID-19 database, we identified previously uninfected, non-hospitalized individuals with COVID-19 with ≥1 risk factor for disease progression who were prescribed either NMV/r or MPV within 3 days of a positive test. We used inverse probability of treatment weights (IPTW) to account for providers' preferences for a specific treatment. Absolute risk difference (ARD) with 95% confidence intervals were determined for those treated with NMV/r vs. MPV. The primary outcome was hospitalization or death within 30 days of treatment prescription using the IPTW approach. Analyses were repeated using propensity-score matched groups. RESULTS: Between January 1 and November 30, 2022, 9,180 individuals were eligible for inclusion (6,592 prescribed NMV/r; 2,454 prescribed MPV). The ARD for hospitalization/death for NMV/r vs MPV was -0.25 (95% CI -0.79 to 0.28). There was no statistically significant difference in ARD among strata by age, race, comorbidities, or symptoms at baseline. Kaplan-Meier curves did not demonstrate a difference between the two groups (p-value = 0.6). Analysis of the propensity-score matched cohort yielded similar results (ARD for NMV/r vs. MPV -0.9, 95% CI -2.02 to 0.23). Additional analyses showed no difference for development of severe/critical/fatal disease by treatment group. CONCLUSION: We found no significant difference in short term risk of hospitalization or death among at-risk individuals with COVID-19 treated with either NMV/r or MPV.
Subject(s)
Antiviral Agents , COVID-19 Drug Treatment , COVID-19 , Cytidine , Disease Progression , Hospitalization , Hydroxylamines , Leucine , Ritonavir , SARS-CoV-2 , Humans , Male , Female , Ritonavir/therapeutic use , Middle Aged , Hydroxylamines/therapeutic use , Cytidine/analogs & derivatives , Cytidine/therapeutic use , COVID-19/mortality , COVID-19/epidemiology , Antiviral Agents/therapeutic use , Leucine/analogs & derivatives , Leucine/therapeutic use , Aged , SARS-CoV-2/isolation & purification , Proline/analogs & derivatives , Proline/therapeutic use , Indoles/therapeutic use , Adult , Pandemics , Risk Factors , Coronavirus Infections/drug therapy , Coronavirus Infections/mortality , Pneumonia, Viral/drug therapy , Pneumonia, Viral/mortality , Pneumonia, Viral/virology , Betacoronavirus , Lactams , NitrilesABSTRACT
The family of human-infecting coronaviruses (HCoVs) poses a serious threat to global health and includes several highly pathogenic strains that cause severe respiratory illnesses. It is essential that we develop effective broad-spectrum anti-HCoV agents to prepare for future outbreaks. In this study, we used PROteolysis TArgeting Chimera (PROTAC) technology focused on degradation of the HCoV main protease (Mpro), a conserved enzyme essential for viral replication and pathogenicity. By adapting the Mpro inhibitor GC376, we produced two novel PROTACs, P2 and P3, which showed relatively broad-spectrum activity against the human-infecting CoVs HCoV-229E, HCoV-OC43, and SARS-CoV-2. The concentrations of these PROTACs that reduced virus replication by 50 % ranged from 0.71 to 4.6 µM, and neither showed cytotoxicity at 100 µM. Furthermore, mechanistic binding studies demonstrated that P2 and P3 effectively targeted HCoV-229E, HCoV-OC43, and SARS-CoV-2 by degrading Mpro within cells in vitro. This study highlights the potential of PROTAC technology in the development of broad-spectrum anti-HCoVs agents, presenting a novel approach for dealing with future viral outbreaks, particularly those stemming from CoVs.
Subject(s)
Antiviral Agents , SARS-CoV-2 , Humans , Antiviral Agents/pharmacology , Antiviral Agents/chemistry , Antiviral Agents/chemical synthesis , SARS-CoV-2/drug effects , SARS-CoV-2/enzymology , Proteolysis/drug effects , Coronavirus 229E, Human/drug effects , Coronavirus OC43, Human/drug effects , Virus Replication/drug effects , Molecular Structure , Dose-Response Relationship, Drug , Microbial Sensitivity Tests , Structure-Activity Relationship , Drug Development , Lactams , Leucine/analogs & derivatives , Sulfonic AcidsABSTRACT
INTRODUCTION: Anaplastic lymphoma kinase (ALK) tyrosine kinase inhibitors (TKIs) are standard first- and second-line treatment for advanced ALK+ non-small cell lung cancer (NSCLC). We evaluated outcomes in patients with ALK+ NSCLC receiving third-line ALK TKI versus non-ALK-directed therapy. METHODS: Flatiron Health OncoEMR data were extracted for patients with ALK+ NSCLC initiating first-line ALK TKI between January 2015 and March 2022 followed by second-line ALK TKI and third-line ALK TKI (group A) or non-TKI therapy (group B). Time-to-treatment discontinuation (TTD) and overall survival (OS) were analyzed using multivariate modelling. RESULTS: Among patients receiving third-line ALK TKI (A, n = 85) or non-TKI therapy (B, n = 43), most received first-line crizotinib (A/B: 64%/60%) and second-line alectinib (36%/30%), ceritinib (24%/19%), or lorlatinib (15%/30%). Common third-line treatments were lorlatinib/alectinib (41%/33%) in A and immunotherapy, chemotherapy, or chemotherapy + immunotherapy (30%/28%/21%) in B. Group A versus B had longer TTD of first-line treatment (hazard ratio [HR] 0.62, 95% confidence interval [CI] 0.41-0.93; p = 0.020) and second-line treatment (HR 0.50, 95% CI 0.33-0.75; p < 0.001) and longer OS from start of first-line treatment (HR 0.32, 95% CI 0.19-0.54; p < 0.001) and second-line treatment (HR 0.40, 95% CI 0.24-0.66; p < 0.001). For third-line treatment, median TTD (A/B) was 6.2/2.4 months (HR 0.61, 95% CI 0.37-1.00; p = 0.049) and OS was 17.6/6.5 months (HR 0.57, 95% CI 0.33-0.98; p = 0.042). CONCLUSIONS: Patients receiving third-line non-ALK-directed therapy had suboptimal outcomes on prior TKIs. Patients with longer duration of prior ALK TKI treatment appeared to benefit from third-line ALK TKIs.
Subject(s)
Anaplastic Lymphoma Kinase , Carcinoma, Non-Small-Cell Lung , Crizotinib , Lung Neoplasms , Piperidines , Protein Kinase Inhibitors , Humans , Carcinoma, Non-Small-Cell Lung/drug therapy , Lung Neoplasms/drug therapy , Anaplastic Lymphoma Kinase/antagonists & inhibitors , Female , Male , Middle Aged , Protein Kinase Inhibitors/therapeutic use , Aged , Piperidines/therapeutic use , Adult , Crizotinib/therapeutic use , Aminopyridines/therapeutic use , Lactams/therapeutic use , Pyrimidines/therapeutic use , Treatment Outcome , Sulfones/therapeutic use , Carbazoles/therapeutic use , Pyrazoles/therapeutic use , Retrospective Studies , Antineoplastic Agents/therapeutic useABSTRACT
OBJECTIVES: This study aimed to determine the association of early use of oral antiviral drugs (including nirmatrelvir-ritonavir and molnupiravir) with the risk of post COVID-19 condition (PCC) and compare the possible efficacy of nirmatrelvir-ritonavir and molnupiravir. METHODS: PubMed, Web of Science, Embase, Cochrane, MedRxiv, and Psycinfo were searched from inception until November 1, 2023. We included studies that assessed the effect of oral antiviral drugs on the incidence of PCC. Pairwise and network meta-analyses were conducted using a random-effects model. Risk ratios (RRs) for oral antiviral drugs were calculated with a confidence interval (CI). RESULTS: Nine observational studies containing 866,066 patients were included. Nirmatrelvir-ritonavir and molnupiravir were evaluated in eight and two studies respectively, with both drugs evaluated in one study. Pair-wise meta-analysis showed that early oral antiviral drugs reduced PCC risk (RR 0.77, 95% CI 0.68-0.88). Network meta-analysis showed that nirmatrelvir-ritonavir may perform better than molnupiravir (surface under the cumulative ranking curve: 95.5% vs. 31.6%) at reducing PCC risk. CONCLUSIONS: Early use of oral antiviral drugs may potentially protect against developing PCC in non-hospitalized patients with COVID-19. These findings support the standardized administration of oral antiviral drugs in patients during the acute phase of COVID-19 according to the guidelines.
Subject(s)
Antiviral Agents , COVID-19 Drug Treatment , COVID-19 , Network Meta-Analysis , Ritonavir , SARS-CoV-2 , Humans , Antiviral Agents/therapeutic use , Antiviral Agents/administration & dosage , Ritonavir/therapeutic use , Ritonavir/administration & dosage , Administration, Oral , COVID-19/epidemiology , Drug Combinations , Hydroxylamines/therapeutic use , Hydroxylamines/administration & dosage , Post-Acute COVID-19 Syndrome , Lactams , Cytidine/analogs & derivatives , Nitriles , Proline , LeucineABSTRACT
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.
Subject(s)
Antiviral Agents , Azoles , Coronavirus 3C Proteases , Isoindoles , Organoselenium Compounds , Proline , SARS-CoV-2 , Antiviral Agents/pharmacology , Antiviral Agents/chemistry , Humans , SARS-CoV-2/drug effects , SARS-CoV-2/enzymology , Organoselenium Compounds/pharmacology , Organoselenium Compounds/chemistry , Isoindoles/pharmacology , Coronavirus 3C Proteases/antagonists & inhibitors , Coronavirus 3C Proteases/metabolism , Azoles/pharmacology , Azoles/chemistry , Proline/analogs & derivatives , Proline/pharmacology , Proline/chemistry , Molecular Docking Simulation , Protease Inhibitors/pharmacology , Protease Inhibitors/chemistry , COVID-19 Drug Treatment , COVID-19/virology , HEK293 Cells , Lactams , Leucine/analogs & derivatives , Sulfonic AcidsABSTRACT
In a pivotal trial (EPIC-HR), a 5-day course of oral ritonavir-boosted nirmatrelvir, given early during symptomatic SARS-CoV-2 infection (within three days of symptoms onset), decreased hospitalization and death by 89.1% and nasal viral load by 0.87 log relative to placebo in high-risk individuals. Yet, nirmatrelvir/ritonavir failed as post-exposure prophylaxis in a trial, and frequent viral rebound has been observed in subsequent cohorts. We develop a mathematical model capturing viral-immune dynamics and nirmatrelvir pharmacokinetics that recapitulates viral loads from this and another clinical trial (PLATCOV). Our results suggest that nirmatrelvir's in vivo potency is significantly lower than in vitro assays predict. According to our model, a maximally potent agent would reduce the viral load by approximately 3.5 logs relative to placebo at 5 days. The model identifies that earlier initiation and shorter treatment duration are key predictors of post-treatment rebound. Extension of treatment to 10 days for Omicron variant infection in vaccinated individuals, rather than increasing dose or dosing frequency, is predicted to lower the incidence of viral rebound significantly.
Subject(s)
Antiviral Agents , COVID-19 Drug Treatment , COVID-19 , Ritonavir , SARS-CoV-2 , Viral Load , Humans , SARS-CoV-2/drug effects , Ritonavir/therapeutic use , Ritonavir/administration & dosage , COVID-19/prevention & control , COVID-19/virology , COVID-19/immunology , Viral Load/drug effects , Antiviral Agents/administration & dosage , Antiviral Agents/therapeutic use , Antiviral Agents/pharmacology , Indazoles/pharmacology , Models, Theoretical , Post-Exposure Prophylaxis/methods , Lactams , Leucine , Nitriles , ProlineABSTRACT
Nirmatrelvir, a pivotal component of the oral antiviral Paxlovid for COVID-19, targets the SARS-CoV-2 main protease (Mpro) as a covalent inhibitor. Here, we employed combined computational methods to explore how the prevalent Omicron variant mutation P132H, alone and in combination with A173V (P132H-A173V), affects nirmatrelvir's efficacy. Our findings suggest that P132H enhances the noncovalent binding affinity of Mpro for nirmatrelvir, whereas P132H-A173V diminishes it. Although both mutants catalyze the rate-limiting step more efficiently than the wild-type (WT) Mpro, P132H slows the overall rate of covalent bond formation, whereas P132H-A173V accelerates it. Comprehensive analysis of noncovalent and covalent contributions to the overall binding free energy of the covalent complex suggests that P132H likely enhances Mpro sensitivity to nirmatrelvir, while P132H-A173V may confer resistance. Per-residue decompositions of the binding and activation free energies pinpoint key residues that significantly affect the binding affinity and reaction rates, revealing how the mutations modulate these effects. The mutation-induced conformational perturbations alter drug-protein local contact intensities and the electrostatic preorganization of the protein, affecting noncovalent binding affinity and the stability of key reaction states, respectively. Our findings inform the mechanisms of nirmatrelvir resistance and sensitivity, facilitating improved drug design and the detection of resistant strains.
Subject(s)
Antiviral Agents , Coronavirus 3C Proteases , Mutation , SARS-CoV-2 , SARS-CoV-2/enzymology , SARS-CoV-2/drug effects , SARS-CoV-2/genetics , Coronavirus 3C Proteases/antagonists & inhibitors , Coronavirus 3C Proteases/metabolism , Coronavirus 3C Proteases/chemistry , Coronavirus 3C Proteases/genetics , Antiviral Agents/pharmacology , Antiviral Agents/chemistry , Humans , COVID-19 Drug Treatment , Molecular Dynamics Simulation , Protease Inhibitors/pharmacology , Protease Inhibitors/chemistry , Protease Inhibitors/metabolism , Leucine/chemistry , Thermodynamics , Sulfonamides/pharmacology , Sulfonamides/chemistry , Sulfonamides/metabolism , Protein Binding , Succinates/chemistry , Succinates/pharmacology , Succinates/metabolism , Lactams , Nitriles , ProlineABSTRACT
PURPOSE: The study of cell-free DNA (cfDNA) enables sequential analysis of tumor cell-specific genetic alterations in patients with neuroblastoma. EXPERIMENTAL DESIGN: Eighteen patients with relapsing neuroblastoma having received lorlatinib, a third-generation ALK inhibitor, were identified (SACHA national registry and/or in the institution). cfDNA was analyzed at relapse for nine patients and sequentially for five patients (blood/bone marrow plasma) by performing whole-genome sequencing library construction followed by ALK-targeted ddPCR of the hotspot mutations [F1174L, R1275Q, and I1170N; variant allele fraction (VAF) detection limit 0.1%] and whole-exome sequencing (WES) to evaluate disease burden and clonal evolution, following comparison with tumor/germline WES. RESULTS: Overall response rate to lorlatinib was 33% (CI, 13%-59%), with response observed in 6/10 cases without versus 0/8 cases with MYCN amplification (MNA). ALK VAFs correlated with the overall clinical disease status, with a VAF < 0.1% in clinical remission, versus higher VAFs (>30%) at progression. Importantly, sequential ALK ddPCR detected relapse earlier than clinical imaging. cfDNA WES revealed new SNVs, not seen in the primary tumor, in all instances of disease progression after lorlatinib treatment, indicating clonal evolution, including alterations in genes linked to tumor aggressivity (TP53) or novel targets (EGFR). Gene pathway analysis revealed an enrichment for genes targeting cell differentiation in emerging clones, and cell adhesion in persistent clones. Evidence of clonal hematopoiesis could be observed in follow-up samples. CONCLUSIONS: We demonstrate the clinical utility of combining ALK cfDNA ddPCR for disease monitoring and cfDNA WES for the study of clonal evolution and resistance mechanisms in patients with neuroblastoma receiving ALK-targeted therapy.
Subject(s)
Anaplastic Lymphoma Kinase , Cell-Free Nucleic Acids , Clonal Evolution , Mutation , Neuroblastoma , Humans , Neuroblastoma/genetics , Neuroblastoma/drug therapy , Neuroblastoma/pathology , Anaplastic Lymphoma Kinase/genetics , Anaplastic Lymphoma Kinase/antagonists & inhibitors , Clonal Evolution/genetics , Male , Female , Child , Child, Preschool , Cell-Free Nucleic Acids/genetics , Aminopyridines/therapeutic use , Pyrazoles/therapeutic use , Lactams , Infant , Adolescent , Exome Sequencing , Protein Kinase Inhibitors/therapeutic use , Neoplasm Recurrence, Local/genetics , Neoplasm Recurrence, Local/drug therapy , Neoplasm Recurrence, Local/pathology , Molecular Targeted Therapy/methods , Biomarkers, Tumor/genetics , Whole Genome Sequencing/methodsABSTRACT
BACKGROUND: Non-small cell lung cancer (NSCLC) harboring ROS1 rearrangements is a molecular subset that exhibits favorable responses to tyrosine kinase inhibitor (TKI) treatment than chemotherapy. This study investigated real-world treatment patterns and survival outcomes among patients with ROS1-rearranged advanced NSCLC. METHODS: We conducted a retrospective analysis of patients with ROS1-rearranged advanced NSCLC treated in four different hospitals in China from August 2018 to March 2022. The study analyzed gene fusion distribution, resistance patterns, and survival outcomes. RESULTS: ROS1 rearrangement occurs in 1.8 % (550/31,225) of our study cohort. CD74 was the most common ROS1 fusion partner, accounting for 45.8 %. Crizotinib was used in 73.9 % of patients in the first-line treatment, and an increased use of chemotherapy, ceritinib, and lorlatinib was seen in the second-line setting. Lung (43.2 %) and brain (27.6 %) were the most common sites of progression in first-line setting, while brain progression (39.2 %) was the most common site of progression in second-line. Median overall survival was 46 months (95 % confidence intervals: 39.6-52.4). First-line crizotinib use yielded significantly superior survival outcomes over chemotherapy in terms of progression-free (18.5 vs. 6.0; p < 0.001) and overall survival (49.8 vs. 37; p = 0.024). The choice of treatment in the latter line also had survival implications, wherein survival outcomes were better when first-line crizotinib was followed by sequential TKI therapy than first-line chemotherapy followed by TKI therapy. CONCLUSIONS: Our study provided insights into the real-world treatment, drug resistance patterns, and survival outcomes among patients with ROS1-rearranged NSCLC. This information serves as a valuable reference for guiding the treatment of this molecular subset of NSCLC.