RESUMO
The presence of heterogeneity in responses to oncolytic virotherapy poses a barrier to clinical effectiveness, as resistance to this treatment can occur through the inhibition of viral spread within the tumor, potentially leading to treatment failures. Here we show that 4-octyl itaconate (4-OI), a chemical derivative of the Krebs cycle-derived metabolite itaconate, enhances oncolytic virotherapy with VSVΔ51 in various models including human and murine resistant cancer cell lines, three-dimensional (3D) patient-derived colon tumoroids and organotypic brain tumor slices. Furthermore, 4-OI in combination with VSVΔ51 improves therapeutic outcomes in a resistant murine colon tumor model. Mechanistically, we find that 4-OI suppresses antiviral immunity in cancer cells through the modification of cysteine residues in MAVS and IKKß independently of the NRF2/KEAP1 axis. We propose that the combination of a metabolite-derived drug with an oncolytic virus agent can greatly improve anticancer therapeutic outcomes by direct interference with the type I IFN and NF-κB-mediated antiviral responses.
Assuntos
Terapia Viral Oncolítica , Vírus Oncolíticos , Succinatos , Animais , Humanos , Terapia Viral Oncolítica/métodos , Succinatos/farmacologia , Camundongos , Linhagem Celular Tumoral , Interferon Tipo I/metabolismo , Fator 2 Relacionado a NF-E2/metabolismo , Neoplasias do Colo/terapia , Neoplasias do Colo/imunologia , Neoplasias do Colo/tratamento farmacológico , Antivirais/farmacologia , NF-kappa B/metabolismo , Quinase I-kappa B/metabolismo , Proteína 1 Associada a ECH Semelhante a Kelch/metabolismo , Inflamação/tratamento farmacológico , Feminino , Vírus da Estomatite Vesicular Indiana/fisiologia , Vírus da Estomatite Vesicular Indiana/efeitos dos fármacos , Transdução de Sinais/efeitos dos fármacosRESUMO
A novel strategy that combines oxidative aminocatalysis and gold catalysis allows the preparation of chiral α-quaternary isochromanes, a motif that is prevalent in natural products and synthetic bioactive compounds. In the first step, α-branched aldehydes and propargylic alcohols are transformed into α-quaternary ethers with excellent optical purities (>90 % ee) via oxidative umpolung with DDQ and an amino acid-derived primary amine catalyst. Subsequent gold(I)-catalyzed intramolecular hydroarylation affords the isochromane products with retention of the quaternary stereocenter. A second approach explores the use of allylic alcohols as reaction partners for the oxidative coupling to furnish α-quaternary ethers with generally lower enantiopurities. Stereoretentive cyclization to isochromane products is achieved via intramolecular Friedel-Crafts type alkylation with allylic acetates as a reactive handle. A number of synthetic elaborations and a biological study on these α-quaternary isochromanes highlight the potential applicability of the presented method.
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Carboxylic polyether ionophores (CPIs) are among the most prevalent agricultural antibiotics (notably in the US) and these compounds have been in use for decades. The potential to reposition CPIs beyond veterinary use, e. g. through chemical modifications to enhance their selectivity window, is an exciting challenge and opportunity, considering their general resilience towards resistance development. Given the very large societal impact of these somewhat controversial compounds, it is surprising that many aspects of their mechanisms and activities in cells remain unclear. Here, we report comparative biological activities of the CPI routiennocin and two stereoisomers, including its enantiomer. We used an efficient convergent synthesis strategy to access the compounds and conducted a broad survey of antibacterial activities against planktonic cells and biofilms as well as the compounds' effects on mammalian cells, the latter assessed both via standard cell viability assays and broad morphological profiling. Interestingly, similar bioactivity of the enantiomeric pair was observed across all assays, strongly suggesting that chiral interactions do not play a decisive role in the mode of action. Overall, our findings are consistent with a mechanistic model involving highly dynamic behaviour of CPIs in biological membranes.
Assuntos
Antibacterianos , Policetídeos de Poliéter , Animais , Antibacterianos/farmacologia , Ionóforos/química , Mamíferos/metabolismoRESUMO
Electrophilic groups are one of the key pillars of contemporary chemical biology and medicinal chemistry. For instance, 3-membered N-heterocyclic compounds-such as aziridines, azirines, and oxaziridines-possess unique electronic and structural properties which underlie their potential and applicability as covalent tools. The α-lactams are also members of this group of compounds, however, their utility within the field remains unexplored. Here, we demonstrate an α-lactam reagent (AM2) that is tolerant to aqueous buffers while being reactive towards biologically relevant nucleophiles. Interestingly, carboxylesterasesâ 1 and 2 (CES1/2), both serine hydrolases with key roles in endo- and xenobiotic metabolism, were found as primary covalent targets for AM2 in HepG2 liver cancer cells. All in all, this study constitutes the starting point for the further development and exploration of α-lactam-based electrophilic probes in covalent chemical biology.
Assuntos
Azirinas , Compostos Heterocíclicos , Lactamas , BiologiaRESUMO
Covalently acting compounds experience a strong interest within chemical biology both as molecular probes in studies of fundamental biological mechanisms and/or as novel drug candidates. In this context, the identification of new classes of reactive groups is particularly important as these can expose novel reactivity modes and, consequently, expand the ligandable proteome. Here, we investigated the electrophilic reactivity of the 3-acyl-5-hydroxy-1,5-dihydro-2H-pyrrole-2-one (AHPO) scaffold, a heterocyclic motif that is e.g. present in various bioactive natural products. Our investigations were focused on the compound MT-21 - a simplified structural analogue of the natural product epolactaene - which is known to have both neurotrophic activity and ability to trigger apoptotic cell death. We found that the central N-acyl hemiaminal group of MT-21 can function as an electrophilic centre enabling divergent reactivity with both amine- and thiol-based nucleophiles, which furthermore translated to reactivity with proteins in both cell lysates and live cells. We found that in live cells MT-21 strongly engaged the lipid transport protein fatty acid-binding protein 5 (FABP5) by direct binding to a cysteine residue in the bottom of the ligand binding pocket. Through preparation of a series of MT-21 derivatives, we probed the specificity of this interaction which was found to be strongly dependent on subtle structural changes. Our study suggests that MT-21 may be employed as a tool compound in future studies of the biology of FABP5, which remains incompletely understood. Furthermore, our study has also made clear that other natural products containing the AHPO-motif may likewise possess covalent reactivity and that this property may underlie their biological activity.
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The transcription factor BACH1 is a potential therapeutic target for a variety of chronic conditions linked to oxidative stress and inflammation, as well as cancer metastasis. However, only a few BACH1 degraders/inhibitors have been described. BACH1 is a transcriptional repressor of heme oxygenase 1 (HMOX1), which is positively regulated by transcription factor NRF2 and is highly inducible by derivatives of the synthetic oleanane triterpenoid 2-cyano-3,12-dioxooleana-1,9(11)-dien-28-oic acid (CDDO). Most of the therapeutic activities of these compounds are due to their anti-inflammatory and antioxidant properties, which are widely attributed to their ability to activate NRF2. However, with such a broad range of action, these compounds have other molecular targets that have not been fully identified and could also be of importance for their therapeutic profile. Herein we identified BACH1 as a target of two CDDO-derivatives (CDDO-Me and CDDO-TFEA), but not of CDDO. While both CDDO and CDDO-derivatives activate NRF2 similarly, only CDDO-Me and CDDO-TFEA inhibit BACH1, which explains the much higher potency of these CDDO-derivatives as HMOX1 inducers compared with unmodified CDDO. Notably, we demonstrate that CDDO-Me and CDDO-TFEA inhibit BACH1 via a novel mechanism that reduces BACH1 nuclear levels while accumulating its cytoplasmic form. In an in vitro model, both CDDO-derivatives impaired lung cancer cell invasion in a BACH1-dependent and NRF2-independent manner, while CDDO was inactive. Altogether, our study identifies CDDO-Me and CDDO-TFEA as dual KEAP1/BACH1 inhibitors, providing a rationale for further therapeutic uses of these drugs.
Assuntos
Ácido Oleanólico , Triterpenos , Proteína 1 Associada a ECH Semelhante a Kelch , Fator 2 Relacionado a NF-E2/genética , Fator 2 Relacionado a NF-E2/metabolismo , Ácido Oleanólico/análogos & derivados , Ácido Oleanólico/farmacologia , Estresse Oxidativo , Triterpenos/farmacologiaRESUMO
The development of new immunomodulatory agents can impact various areas of medicine. In particular, compounds with the ability to modulate innate immunological pathways hold significant unexplored potential. Herein, we report a modular synthetic approach to the macrodiolide natural product (-)-vermiculine, an agent previously shown to possess diverse biological effects, including cytotoxic and immunosuppressive activity. The synthesis allows for a high degree of flexibility in modifying the macrocyclic framework, including the formation of all possible stereoisomers. In total, 18 analogues were prepared. Two analogues with minor structural modifications showed clearly enhanced cancer cell line selectivity and reduced toxicity. Moreover, these compounds possessed broad inhibitory activity against innate immunological pathways in human PBMCs, including the DNA-sensing cGAS-STING pathway. Initial mechanistic characterization suggests a surprising impairment of the STING-TBK1 interaction.
Assuntos
Fatores Imunológicos/farmacologia , Proteínas de Membrana/antagonistas & inibidores , Nucleotidiltransferases/antagonistas & inibidores , DNA/efeitos dos fármacos , DNA/metabolismo , Humanos , Fatores Imunológicos/síntese química , Fatores Imunológicos/química , Lactonas/síntese química , Lactonas/química , Lactonas/farmacologia , Leucócitos Mononucleares/efeitos dos fármacos , Leucócitos Mononucleares/metabolismo , Proteínas de Membrana/metabolismo , Conformação Molecular , Nucleotidiltransferases/metabolismoRESUMO
Isobenzopyrylium ions are unique, highly reactive, aromatic intermediates which are largely unexplored in asymmetric catalysis despite their high potential synthetic utility. In this study, an organocatalytic asymmetric multicomponent cascade via dienamine catalysis, involving a cycloaddition, a nucleophilic addition, and a ring-opening reaction, is disclosed. The reaction furnishes chiral tetrahydronaphthols containing four contiguous stereocenters in good to high yield, high diastereoselectivity (up to >20:1), and excellent enantioselectivity (93-98% ee). The obtained products are important synthetic intermediates, and it is demonstrated that they can be used for the generation of frameworks such as octahydrobenzo[h]isoquinoline and [2.2.2]octane scaffolds. Furthermore, mechanistic experiments involving oxygen-18-labeling studies and density functional theory calculations provide a vivid picture of the reaction mechanism. Finally, the bioactivity of 16 representative tetrahydronaphthol compounds has been evaluated in U-2OS cancer cells with some compounds showing a unique profile and a clear morphological change.
RESUMO
Polyether ionophores are complex natural products capable of transporting cations across biological membranes. Many polyether ionophores possess potent antimicrobial activity and a few selected compounds have the ability to target aggressive cancer cells. Nevertheless, ionophore function is believed to be associated with idiosyncratic cellular toxicity and, consequently, human clinical development has not been pursued. Here, we demonstrate that structurally novel polyether ionophores can be efficiently constructed by recycling components of highly abundant polyethers to afford analogues with enhanced antibacterial selectivity compared to a panel of natural polyether ionophores. We used classic degradation reactions of the natural polyethers lasalocid and monensin and combined the resulting fragments with building blocks provided by total synthesis, including halogen-functionalized tetronic acids as cation-binding groups. Our results suggest that structural optimization of polyether ionophores is possible and that this area represents a potential opportunity for future methodological innovation.
Assuntos
Antibacterianos/síntese química , Éteres/química , Ionóforos/química , Aldeídos/química , Antibacterianos/química , Antibacterianos/farmacologia , Linhagem Celular , Sobrevivência Celular/efeitos dos fármacos , Cristalografia por Raios X , Furanos/síntese química , Furanos/química , Humanos , Ionóforos/síntese química , Ionóforos/farmacologia , Lasalocida/síntese química , Lasalocida/química , Conformação Molecular , Monensin/síntese química , Monensin/química , OxirreduçãoRESUMO
Pandemic spread of emerging human pathogenic viruses, such as the current SARS-CoV-2, poses both an immediate and future challenge to human health and society. Currently, effective treatment of infection with SARS-CoV-2 is limited and broad spectrum antiviral therapies to meet other emerging pandemics are absent leaving the World population largely unprotected. Here, we have identified distinct members of the family of polyether ionophore antibiotics with potent ability to inhibit SARS-CoV-2 replication and cytopathogenicity in cells. Several compounds from this class displayed more than 100-fold selectivity between viral-induced cytopathogenicity and inhibition of cell viability, however the compound X-206 displayed >500-fold selectivity and was furthermore able to inhibit viral replication even at sub-nM levels. The antiviral mechanism of the polyether ionophores is currently not understood in detail. We demonstrate, e.g. through unbiased bioactivity profiling, that their effects on the host cells differ from those of cationic amphiphiles such as hydroxychloroquine. Collectively, our data suggest that polyether ionophore antibiotics should be subject to further investigations as potential broad-spectrum antiviral agents.
Assuntos
Antibacterianos/farmacologia , Antivirais/farmacologia , Tratamento Farmacológico da COVID-19 , Éteres Cíclicos/farmacologia , Ionóforos/farmacologia , SARS-CoV-2/efeitos dos fármacos , Animais , Chlorocebus aethiops , Humanos , Células Vero , Replicação Viral/efeitos dos fármacosRESUMO
An amendment to this paper has been published and can be accessed via a link at the top of the paper.
RESUMO
Antiviral strategies to inhibit Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV2) and the pathogenic consequences of COVID-19 are urgently required. Here, we demonstrate that the NRF2 antioxidant gene expression pathway is suppressed in biopsies obtained from COVID-19 patients. Further, we uncover that NRF2 agonists 4-octyl-itaconate (4-OI) and the clinically approved dimethyl fumarate (DMF) induce a cellular antiviral program that potently inhibits replication of SARS-CoV2 across cell lines. The inhibitory effect of 4-OI and DMF extends to the replication of several other pathogenic viruses including Herpes Simplex Virus-1 and-2, Vaccinia virus, and Zika virus through a type I interferon (IFN)-independent mechanism. In addition, 4-OI and DMF limit host inflammatory responses to SARS-CoV2 infection associated with airway COVID-19 pathology. In conclusion, NRF2 agonists 4-OI and DMF induce a distinct IFN-independent antiviral program that is broadly effective in limiting virus replication and in suppressing the pro-inflammatory responses of human pathogenic viruses, including SARS-CoV2.
Assuntos
Anti-Inflamatórios/farmacologia , Antivirais/farmacologia , Betacoronavirus/efeitos dos fármacos , Infecções por Coronavirus/tratamento farmacológico , Fumarato de Dimetilo/agonistas , Fator 2 Relacionado a NF-E2/metabolismo , Pneumonia Viral/tratamento farmacológico , Succinatos/agonistas , Adulto , Antioxidantes/farmacologia , Betacoronavirus/metabolismo , COVID-19 , Infecções por Coronavirus/virologia , Fumarato de Dimetilo/farmacologia , Feminino , Expressão Gênica , Técnicas de Silenciamento de Genes , Humanos , Interferon Tipo I , Pulmão/patologia , Masculino , Fator 2 Relacionado a NF-E2/genética , Pandemias , Pneumonia Viral/virologia , SARS-CoV-2 , Transdução de Sinais/efeitos dos fármacos , Succinatos/farmacologia , Replicação Viral/efeitos dos fármacosRESUMO
Phenomycin is a bacterial mini-protein of 89 amino acids discovered more than 50 years ago with toxicity in the nanomolar regime toward mammalian cells. The protein inhibits the function of the eukaryotic ribosome in cell-free systems and appears to target translation initiation. Several fundamental questions concerning the cellular activity of phenomycin, however, have remained unanswered. In this paper, we have used morphological profiling to show that direct inhibition of translation underlies the toxicity of phenomycin in cells. We have performed studies of the cellular uptake mechanism of phenomycin, showing that endosomal escape is the toxicity-limiting step, and we have solved a solution phase high-resolution structure of the protein using NMR spectroscopy. Through bioinformatic as well as functional comparisons between phenomycin and two homologs, we have identified a peptide segment, which constitutes one of two loops in the structure that is critical for the toxicity of phenomycin.
Assuntos
Peptídeos e Proteínas de Sinalização Intercelular/química , Peptídeos e Proteínas de Sinalização Intercelular/toxicidade , Animais , Toxinas Bacterianas/química , Toxinas Bacterianas/metabolismo , Toxinas Bacterianas/toxicidade , Bacteriocinas/farmacocinética , Bacteriocinas/toxicidade , Linhagem Celular , Endossomos/efeitos dos fármacos , Endossomos/metabolismo , Humanos , Peptídeos e Proteínas de Sinalização Intercelular/genética , Peptídeos e Proteínas de Sinalização Intercelular/metabolismo , Células MCF-7 , Camundongos , Mutação , Ressonância Magnética Nuclear Biomolecular , Conformação Proteica , Inibidores da Síntese de Proteínas/química , Inibidores da Síntese de Proteínas/toxicidade , Relação Estrutura-AtividadeRESUMO
In this paper we will outline the efforts we have made recently to establish the profiling platform known as cell painting in our laboratory. This platform, which is based on fluorescence microscopy, allows rapid and cheap access to bioactivity fingerprints for small molecules and thereby can contribute with important information in many experimental situations that is faced in laboratories involved in molecular probe design, mode-of-action studies or that perform focused phenotypic screens. We have tried to achieve the following two objectives: (1) provide a detailed description of the hurdles that we had to overcome during establishment and describe our final protocol; (2) provide a more pedagogical description of the different methods used to analyse and represent data from this experiment. Finally, we provide an example of how the method can be used to clarify mechanistic dichotomies.
Assuntos
Microscopia de Fluorescência/métodos , Sondas Moleculares/química , Linhagem Celular Tumoral , Corantes Fluorescentes/química , Humanos , Piperidonas/química , Piperidonas/farmacologia , Biossíntese de Proteínas/efeitos dos fármacos , Streptomyces/química , Streptomyces/metabolismoRESUMO
The natural products pantomycin and stendomycin were both reported as antimicrobial agents. We demonstrate by gene cluster analysis, LC-MS analysis, and isolation that these polypeptides are identical, and we identify previously unknown congeners. We show that stendomycin can be chemically modified at its electrophilic dehydrobutyrine moiety yielding the first bioactive analogue of this natural product which can undergo additional functionalization. This compound may be a valuable starting point for molecular probe development, and we invite its distribution to the scientific community.